A nonproliferating parvovirus vaccine vector elicits sustained, protective humoral immunity following a single intravenous or intranasal inoculation.

PubMed

Palmer, Gene A; Brogdon, Jennifer L; Constant, Stephanie L; Tattersall, Peter

2004-02-01

An ideal vaccine delivery system would elicit persistent protection following a single administration, preferably by a noninvasive route, and be safe even in the face of immunosuppression, either inherited or acquired, of the recipient. We have exploited the unique life cycle of the autonomous parvoviruses to develop a nonproliferating vaccine platform that appears to both induce priming and continually boost a protective immune response following a single inoculation. A crippled parvovirus vector was constructed, based on a chimera between minute virus of mice (MVM) and LuIII, which expresses Borrelia burgdorferi outer surface protein A (OspA) instead of its coat protein. The vector was packaged into an MVM lymphotropic capsid and inoculated into naive C3H/HeNcr mice. Vaccination with a single vector dose, either intravenously or intranasally, elicited high-titer anti-OspA-specific antibody that provided protection from live spirochete challenge and was sustained over the lifetime of the animal. Both humoral and cell-mediated Th(1) immunity was induced, as shown by anti-OspA immunoglobulin G2a antibody and preferential gamma interferon production by OspA-specific CD4(+) T cells.

A Nonproliferating Parvovirus Vaccine Vector Elicits Sustained, Protective Humoral Immunity following a Single Intravenous or Intranasal Inoculation

PubMed Central

Palmer, Gene A.; Brogdon, Jennifer L.; Constant, Stephanie L.; Tattersall, Peter

2004-01-01

An ideal vaccine delivery system would elicit persistent protection following a single administration, preferably by a noninvasive route, and be safe even in the face of immunosuppression, either inherited or acquired, of the recipient. We have exploited the unique life cycle of the autonomous parvoviruses to develop a nonproliferating vaccine platform that appears to both induce priming and continually boost a protective immune response following a single inoculation. A crippled parvovirus vector was constructed, based on a chimera between minute virus of mice (MVM) and LuIII, which expresses Borrelia burgdorferi outer surface protein A (OspA) instead of its coat protein. The vector was packaged into an MVM lymphotropic capsid and inoculated into naive C3H/HeNcr mice. Vaccination with a single vector dose, either intravenously or intranasally, elicited high-titer anti-OspA-specific antibody that provided protection from live spirochete challenge and was sustained over the lifetime of the animal. Both humoral and cell-mediated Th1 immunity was induced, as shown by anti-OspA immunoglobulin G2a antibody and preferential gamma interferon production by OspA-specific CD4+ T cells. PMID:14722265

Tick receptor for outer surface protein A from Ixodes ricinus — the first intrinsically disordered protein involved in vector-microbe recognition

NASA Astrophysics Data System (ADS)

Urbanowicz, Anna; Lewandowski, Dominik; Szpotkowski, Kamil; Figlerowicz, Marek

2016-04-01

The tick receptor for outer surface protein A (TROSPA) is the only identified factor involved in tick gut colonization by various Borrelia species. TROSPA is localized in the gut epithelium and can recognize and bind the outer surface bacterial protein OspA via an unknown mechanism. Based on earlier reports and our latest observations, we considered that TROSPA would be the first identified intrinsically disordered protein (IDP) involved in the interaction between a vector and a pathogenic microbe. To verify this hypothesis, we performed structural studies of a TROSPA mutant from Ixodes ricinus using both computational and experimental approaches. Irrespective of the method used, we observed that the secondary structure content of the TROSPA polypeptide chain is low. In addition, the collected SAXS data indicated that this protein is highly extended and exists in solution as a set of numerous conformers. These features are all commonly considered hallmarks of IDPs. Taking advantage of our SAXS data, we created structural models of TROSPA and proposed a putative mechanism for the TROSPA-OspA interaction. The disordered nature of TROSPA may explain the ability of a wide spectrum of Borrelia species to colonize the tick gut.

Chimeric OspA genes, proteins and methods of use thereof

DOE Office of Scientific and Technical Information (OSTI.GOV)

Crowe, Brian A.; Livey, Ian; O'Rourke, Maria

The invention relates to the development of chimeric OspA molecules for use in a new Lyme vaccine. More specifically, the chimeric OspA molecules comprise the proximal portion from one OspA serotype, together with the distal portion from another OspA serotype, while retaining antigenic properties of both of the parent polypeptides. The chimeric OspA molecules are delivered alone or in combination to provide protection against a variety of Borrelia genospecies. The invention also provides methods for administering the chimeric OspA molecules to a subject in the prevention and treatment of Lyme disease or borreliosis.

Differential response of oyster shell powder on enzyme profile and nutritional value of oyster mushroom Pleurotus florida PF05.

PubMed

Naraian, Ram; Narayan, Om Prakash; Srivastava, Jatin

2014-01-01

Oyster mushroom Pleurotus florida was cultivated on different combinations of wheat straw (WS) as basal substrate and oyster shell powder (OSP) supplement. The OSP supplementation considerably responded to different cultivation phases. The mycelium grew fast and showed rapid growth rate (8.91 mmd(-1)) in WS + OSP (97 + 3) combination while WS + OSP (92 + 8) showed maximum laccase (3.133 U/g) and Mn peroxidase (MnP) activities (0.091 U/g). The climax level of laccase (5.433 U/g) and MnP (0.097 U/g) was recorded during fruit body initiation in WS + OSP (97 + 3) and WS + OSP (98 + 2) combinations, respectively. The WS + OSP (97 + 3) combination represented the best condition for mushroom cultivation and produced the highest biological efficiency (147%). In addition, protein and lipid contents in fruit bodies were slightly improved in response to OSP. The carbohydrate was significantly increased by raising concentration of OSP. The highest values of protein, carbohydrate, and lipid noted were 31.3 μg/g, 0.0639 (g/g), and 0.373 (g/g) correspondingly. Conclusively it was evident that lower concentrations of OSP acted positively and relatively to higher concentrations and improved nutritional content which may suitably be used to enhance both yield and nutritional values of mushroom.

Crystal Structure of Neurotropism-Associated Variable Surface Protein 1 (VSP1) of Borrelia Turicatae

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lawson,C.; Yung, B.; Barbour, A.

2006-01-01

Vsp surface lipoproteins are serotype-defining antigens of relapsing fever spirochetes that undergo multiphasic antigenic variation to allow bacterial persistence in spite of an immune response. Two isogenic serotypes of Borrelia turicatae strain Oz1 differ in their Vsp sequences and in disease manifestations in infected mice: Vsp1 is associated with the selection of a neurological niche, while Vsp2 is associated with blood and skin infection. We report here crystal structures of the Vsp1 dimer at 2.7 and 2.2 Angstroms. The structures confirm that relapsing fever Vsp proteins share a common helical fold with OspCs of Lyme disease-causing Borrelia. The fold featuresmore » an inner stem formed by highly conserved N and C termini and an outer 'dome' formed by the variable central residues. Both Vsp1 and OspC structures possess small water-filled cavities, or pockets, that are lined largely by variable residues and are thus highly variable in shape. These features appear to signify tolerance of the Vsp-OspC fold for imperfect packing of residues at its antigenic surface. Structural comparison of Vsp1 with a homology model for Vsp2 suggests that observed differences in disease manifestation may arise in part from distinct differences in electrostatic surface properties; additional predicted positively charged surface patches on Vsp2 compared to Vsp1 may be sufficient to explain the relative propensity of Vsp2 to bind to acidic glycosaminoglycans.« less

Reservoir targeted vaccine for lyme borreliosis induces a yearlong, neutralizing antibody response to OspA in white-footed mice.

PubMed

Meirelles Richer, Luciana; Aroso, Miguel; Contente-Cuomo, Tania; Ivanova, Larisa; Gomes-Solecki, Maria

2011-11-01

Lyme disease is caused by the spirochete Borrelia burgdorferi. The enzootic cycle of this pathogen requires that Ixodes spp. acquire B. burgdorferi from infected wildlife reservoirs and transmit it to other uninfected wildlife. At present, there are no effective measures to control B. burgdorferi; there is no human vaccine available, and existing vector control measures are generally not acceptable to the public. However, if B. burgdorferi could be eliminated from its reservoir hosts or from the ticks that feed on them, the enzootic cycle would be broken, and the incidence of Lyme disease would decrease. We developed OspA-RTV, a reservoir targeted bait vaccine (RTV) based on the immunogenic outer surface protein A (OspA) of B. burgdorferi aimed at breaking the natural cycle of this spirochete. White-footed mice, the major reservoir species for this spirochete in nature developed a systemic OspA-specific IgG response as a result of ingestion of the bait formulation. This immune response protected white-footed mice against B. burgdorferi infection upon tick challenge and cleared B. burgdorferi from the tick vector. In performing extensive studies to optimize the OspA-RTV for field deployment, we determined that mice that consumed the vaccine over periods of 1 or 4 months developed a yearlong, neutralizing anti-OspA systemic IgG response. Furthermore, we defined the minimum number of OspA-RTV units needed to induce a protective immune response.

Cross-reactive acquired immunity influences transmission success of the Lyme disease pathogen, Borrelia afzelii.

PubMed

Jacquet, Maxime; Durand, Jonas; Rais, Olivier; Voordouw, Maarten J

2015-12-01

Cross-reactive acquired immunity in the vertebrate host induces indirect competition between strains of a given pathogen species and is critical for understanding the ecology of mixed infections. In vector-borne diseases, cross-reactive antibodies can reduce pathogen transmission at the vector-to-host and the host-to-vector lifecycle transition. The highly polymorphic, immunodominant, outer surface protein C (OspC) of the tick-borne spirochete bacterium Borrelia afzelii induces a strong antibody response in the vertebrate host. To test how cross-immunity in the vertebrate host influences tick-to-host and host-to-tick transmission, mice were immunized with one of two strain-specific recombinant OspC proteins (A3, A10), challenged via tick bite with one of the two B. afzelii ospC strains (A3, A10), and infested with xenodiagnostic ticks. Immunization with a given rOspC antigen protected mice against homologous strains carrying the same major ospC group allele but provided little or no cross-protection against heterologous strains carrying a different major ospC group allele. There were cross-immunity effects on the tick spirochete load but not on the probability of host-to-tick transmission. The spirochete load in ticks that had fed on mice with cross-immune experience was reduced by a factor of two compared to ticks that had fed on naive control mice. In addition, strain-specific differences in mouse spirochete load, host-to-tick transmission, tick spirochete load, and the OspC-specific IgG response revealed the mechanisms that determine variation in transmission success between strains of B. afzelii. This study shows that cross-immunity in infected vertebrate hosts can reduce pathogen load in the arthropod vector with potential consequences for vector-to-host pathogen transmission. Copyright © 2015 Elsevier B.V. All rights reserved.

First characterization of immunogenic conjugates of Vi negative Salmonella Typhi O-specific polysaccharides with rEPA protein for vaccine development.

PubMed

Salman, M; St Michael, F; Ali, A; Jabbar, A; Cairns, C; Hayes, A C; Rahman, M; Iqbal, M; Haque, A; Cox, A D

2017-11-01

Efficacious typhoid vaccines for young children will significantly reduce the disease burden in developing world. The Vi polysaccharide based conjugate vaccines (Vi-rEPA) against Salmonella Typhi Vi positive strains has shown high efficacy but may be ineffective against Vi negative S. Typhi. In this study, for the first time, we report the synthesis and evaluation of polysaccharide-protein conjugates of Vi negative S. Typhi as potential vaccine candidates. Four different conjugates were synthesized using recombinant exoprotein A of Pseudomonas aeruginosa (rEPA) and human serum albumin (HSA) as the carrier proteins, using either direct reductive amination or an intermediate linker molecule, adipic acid dihydrazide (ADH). Upon injection into mice, a significantly higher antibody titer was observed in mice administrated with conjugate-1 (OSP-HSA) (P=0.0001) and conjugate 2 (OSP-rEPA) (P≤0.0001) as compared to OSP alone. In contrast, the antibody titer elicited by conjugate 3 (OSP ADH -HSA) and conjugate 4 (OSP ADH -rEPA) were insignificant (P=0.1684 and P=0.3794, respectively). We conclude that reductive amination is the superior method to prepare the S. Typhi OSP glycoconjugate. Moreover, rEPA was a better carrier protein than HSA. Thus OSP-rEPA conjugate seems to be efficacious typhoid vaccines candidate, it may be evaluated further and recommended for the clinical trials. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

Crystal structures of the Erp protein family members ErpP and ErpC from Borrelia burgdorferi reveal the reason for different affinities for complement regulator factor H.

PubMed

Brangulis, Kalvis; Petrovskis, Ivars; Kazaks, Andris; Akopjana, Inara; Tars, Kaspars

2015-05-01

Borrelia burgdorferi is the causative agent of Lyme disease, which can be acquired after the bite of an infected Ixodes tick. As a strategy to resist the innate immunity and to successfully spread and proliferate, B. burgdorferi expresses a set of outer membrane proteins that are capable of binding complement regulator factor H (CFH), factor H-like protein 1 (CFHL-1) and factor H-related proteins (CFHR) to avoid complement-mediated killing. B. burgdorferi B31 contains three proteins that belong to the Erp (OspE/F-related) protein family and are capable of binding CFH and some CFHRs, namely ErpA, ErpC and ErpP. We have determined the crystal structure of ErpP at 2.53Å resolution and the crystal structure of ErpC at 2.15Å resolution. Recently, the crystal structure of the Erp family member OspE from B. burgdorferi N40 was determined in complex with CFH domains 19-20, revealing the residues involved in the complex formation. Despite the high sequence conservation between ErpA, ErpC, ErpP and the homologous protein OspE (78-80%), the affinity for CFH and CFHRs differs markedly among the Erp family members, suggesting that ErpC may bind only CFHRs but not CFH. A comparison of the binding site in OspE with those of ErpC and ErpP revealed that the extended loop region, which is only observed in the potential binding site of ErpC, plays an important role by preventing the binding of CFH. These results can explain the inability of ErpC to bind CFH, whereas ErpP and ErpA still possess the ability to bind CFH. Copyright © 2015 Elsevier B.V. All rights reserved.

Recombinant constructs of Borrelia burgdorferi

DOEpatents

Dattwyler, Raymond J.; Gomes-Solecki, Maria J. C.; Luft, Benjamin J.; Dunn, John J.

2007-02-20

Novel chimeric nucleic acids, encoding chimeric Borrelia proteins comprising OspC or an antigenic fragment thereof and OspA or an antigenic fragment thereof, are disclosed. Chimeric proteins encoded by the nucleic acid sequences are also disclosed. The chimeric proteins are useful as vaccine immunogens against Lyme borreliosis, as well as for immunodiagnostic reagents.

Effect of Plasma Surface Finish on Wettability and Mechanical Properties of SAC305 Solder Joints

NASA Astrophysics Data System (ADS)

Kim, Kyoung-Ho; Koike, Junichi; Yoon, Jeong-Won; Yoo, Sehoon

2016-12-01

The wetting behavior, interfacial reactions, and mechanical reliability of Sn-Ag-Cu solder on a plasma-coated printed circuit board (PCB) substrate were evaluated under multiple heat-treatments. Conventional organic solderability preservative (OSP) finished PCBs were used as a reference. The plasma process created a dense and highly cross-linked polymer coating on the Cu substrates. The plasma finished samples had higher wetting forces and shorter zero-cross times than those with OSP surface finish. The OSP sample was degraded after sequential multiple heat treatments and reflow processes, whereas the solderability of the plasma finished sample was retained after multiple heat treatments. After the soldering process, similar microstructures were observed at the interfaces of the two solder joints, where the development of intermetallic compounds was observed. From ball shear tests, it was found that the shear force for the plasma substrate was consistently higher than that for the OSP substrate. Deterioration of the OSP surface finish was observed after multiple heat treatments. Overall, the plasma surface finish was superior to the conventional OSP finish with respect to wettability and joint reliability, indicating that it is a suitable material for the fabrication of complex electronic devices.

Chromosomal localization of murine and human oligodendrocyte-specific protein genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bronstein, J.M.; Wu, S.; Korenberg, J.R.

1996-06-01

Oligodendrocyte-specific protein (OSP) is a recently described protein present only in myelin of the central nervous system. Several inherited disorders of myelin are caused by mutations in myelin genes but the etiology of many remain unknown. We mapped the location of the mouse OSP gene to the proximal region of chromosome 3 using two sets of multilocus crosses and to human chromosome 3 using somatic cell hybrids. Fine mapping with fluorescence in situ hybridization placed the OSP gene at human chromosome 3q26.2-q26.3. To date, there are no known inherited neurological disorders that localize to these regions. 24 refs., 2 figs.

Pre-exposure Prophylaxis With OspA-Specific Human Monoclonal Antibodies Protects Mice Against Tick Transmission of Lyme Disease Spirochetes.

PubMed

Wang, Yang; Kern, Aurélie; Boatright, Naomi K; Schiller, Zachary A; Sadowski, Andrew; Ejemel, Monir; Souders, Colby A; Reimann, Keith A; Hu, Linden; Thomas, William D; Klempner, Mark S

2016-07-15

Tick transmission of Borrelia spirochetes to humans results in significant morbidity from Lyme disease worldwide. Serum concentrations of antibodies against outer surface protein A (OspA) were shown to correlate with protection from infection with Borrelia burgdorferi, the primary cause of Lyme disease in the United States. Mice transgenic for human immunoglobulin genes were immunized with OspA from B. burgdorferi to generate human monoclonal antibodies (HuMabs) against OspA. HuMabs were generated and tested in in vitro borreliacidal assays and animal protection assays. Nearly 100 unique OspA-specific HuMabs were generated, and 4 HuMabs (221-7, 857-2, 319-44, and 212-55) were selected as lead candidates on the basis of borreliacidal activity. HuMabs 319-44, 857-2, and 212-55 were borreliacidal against 1 or 2 Borrelia genospecies, whereas 221-7 was borreliacidal (half maximal inhibitory concentration, < 1 nM) against B. burgdorferi, Borrelia afzelii, and Borrelia garinii, the 3 main genospecies endemic in the United States, Europe, and Asia. All 4 HuMabs completely protected mice from infection at 10 mg/kg in a murine model of tick-mediated transmission of B. burgdorferi  Our study indicates that OspA-specific HuMabs can prevent the transmission of Borrelia and that administration of these antibodies could be employed as preexposure prophylaxis for Lyme disease. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

E2~Ub conjugates regulate the kinase activity of Shigella effector OspG during pathogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pruneda, Jonathan N.; Smith, F. Donelson; Daurie, Angela

Pathogenic bacteria introduce effector proteins directly into the cytosol of eukaryotic cells to promote invasion and colonization. OspG, a Shigella spp. effector kinase, plays a role in this process by helping to suppress the host inflammatory response. OspG has been reported to bind host E2 ubiquitin-conjugating enzymes activated with ubiquitin (E2~Ub), a key enzyme complex in ubiquitin transfer pathways. A cocrystal structure of the OspG/UbcH5c~Ub complex reveals that complex formation has important ramifications for the activity of both OspG and the UbcH5c~Ub conjugate. OspG is a minimal kinase domain containing only essential elements required for catalysis. UbcH5c~Ub binding stabilizes anmore » active conformation of the kinase, greatly enhancing OspG kinase activity. In contrast, interaction with OspG stabilizes an extended, less reactive form of UbcH5c~Ub. Recognizing conserved E2 features, OspG can interact with at least ten distinct human E2s~Ub. Mouse oral infection studies indicate that E2~Ub conjugates act as novel regulators of OspG effector kinase function in eukaryotic host cells.« less

Electromigration Failure Mechanism in Sn-Cu Solder Alloys with OSP Cu Surface Finish

NASA Astrophysics Data System (ADS)

Chu, Ming-Hui; Liang, S. W.; Chen, Chih; Huang, Annie T.

2012-09-01

Organic solderable preservative (OSP) has been adopted as the Cu substrate surface finish in flip-chip solder joints for many years. In this study, the electromigration behavior of lead-free Sn-Cu solder alloys with thin-film under bump metallization and OSP surface finish was investigated. The results showed that severe damage occurred on the substrate side (cathode side), whereas the damage on the chip side (cathode side) was not severe. The damage on the substrate side included void formation, copper dissolution, and formation of intermetallic compounds (IMCs). The OSP Cu interface on the substrate side became the weakest point in the solder joint even when thin-film metallization was used on the chip side. Three-dimensional simulations were employed to investigate the current density distribution in the area between the OSP Cu surface finish and the solder. The results indicated that the current density was higher along the periphery of the bonding area between the solder and the Cu pad, consistent with the area of IMC and void formation in our experimental results.

Whole-Chain Tick Saliva Proteins Presented on Hepatitis B Virus Capsid-Like Particles Induce High-Titered Antibodies with Neutralizing Potential

PubMed Central

Kolb, Philipp; Wallich, Reinhard; Nassal, Michael

2015-01-01

Ticks are vectors for various, including pathogenic, microbes. Tick saliva contains multiple anti-host defense factors that enable ticks their bloodmeals yet also facilitate microbe transmission. Lyme disease-causing borreliae profit specifically from the broadly conserved tick histamine release factor (tHRF), and from cysteine-rich glycoproteins represented by Salp15 from Ixodes scapularis and Iric-1 from Ixodes ricinus ticks which they recruit to their outer surface protein C (OspC). Hence these tick proteins are attractive targets for anti-tick vaccines that simultaneously impair borrelia transmission. Main obstacles are the tick proteins´ immunosuppressive activities, and for Salp15 orthologs, the lack of efficient recombinant expression systems. Here, we exploited the immune-enhancing properties of hepatitis B virus core protein (HBc) derived capsid-like particles (CLPs) to generate, in E. coli, nanoparticulate vaccines presenting tHRF and, as surrogates for the barely soluble wild-type proteins, cysteine-free Salp15 and Iric-1 variants. The latter CLPs were exclusively accessible in the less sterically constrained SplitCore system. Mice immunized with tHRF CLPs mounted a strong anti-tHRF antibody response. CLPs presenting cysteine-free Salp15 and Iric-1 induced antibodies to wild-type, including glycosylated, Salp15 and Iric-1. The broadly distributed epitopes included the OspC interaction sites. In vitro, the anti-Salp15 antibodies interfered with OspC binding and enhanced human complement-mediated killing of Salp15 decorated borreliae. A mixture of all three CLPs induced high titered antibodies against all three targets, suggesting the feasibility of combination vaccines. These data warrant in vivo validation of the new candidate vaccines´ protective potential against tick infestation and Borrelia transmission. PMID:26352137

Strain-specific antibodies reduce co-feeding transmission of the Lyme disease pathogen, Borrelia afzelii.

PubMed

Jacquet, Maxime; Durand, Jonas; Rais, Olivier; Voordouw, Maarten J

2016-03-01

Vector-borne pathogens use a diversity of strategies to evade the vertebrate immune system. Co-feeding transmission is a potential immune evasion strategy because the vector-borne pathogen minimizes the time spent in the vertebrate host. We tested whether the Lyme disease pathogen, Borrelia afzelii, can use co-feeding transmission to escape the acquired immune response in the vertebrate host. We induced a strain-specific, protective antibody response by immunizing mice with one of two variants of OspC (A3 and A10), the highly variable outer surface protein C of Borrelia pathogens. Immunized mice were challenged via tick bite with B. afzelii strains A3 or A10 and infested with larval ticks at days 2 and 34 post-infection to measure co-feeding and systemic transmission respectively. Antibodies against a particular OspC variant significantly reduced co-feeding transmission of the targeted (homologous) strain but not the non-targeted (heterologous) strain. Cross-immunity between OspC antigens had no effect in co-feeding ticks but reduced the spirochaete load twofold in ticks infected via systemic transmission. In summary, OspC-specific antibodies reduced co-feeding transmission of a homologous but not a heterologous strain of B. afzelii. Co-feeding transmission allowed B. afzelii to evade the negative consequences of cross-immunity on the tick spirochaete load. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

Shigella flexneri type III secreted effector OspF reveals new crosstalks of proinflammatory signaling pathways during bacterial infection.

PubMed

Reiterer, Veronika; Grossniklaus, Lars; Tschon, Therese; Kasper, Christoph Alexander; Sorg, Isabel; Arrieumerlou, Cécile

2011-07-01

Shigella flexneri type III secreted effector OspF harbors a phosphothreonine lyase activity that irreversibly dephosphorylates MAP kinases (MAPKs) p38 and ERK in infected epithelial cells and thereby, dampens innate immunity. Whereas this activity has been well characterized, the impact of OspF on other host signaling pathways that control inflammation was unknown. Here we report that OspF potentiates the activation of the MAPK JNK and the transcription factor NF-κB during S. flexneri infection. This unexpected effect of OspF was dependent on the phosphothreonine lyase activity of OspF on p38, and resulted from the disruption of a negative feedback loop regulation between p38 and TGF-beta activated kinase 1 (TAK1), mediated via the phosphorylation of TAK1-binding protein 1. Interestingly, potentiated JNK activation was not associated with enhanced c-Jun signaling as OspF also inhibits c-Jun expression at the transcriptional level. Altogether, our data reveal the impact of OspF on the activation of NF-κB, JNK and c-Jun, and demonstrate the existence of a negative feedback loop regulation between p38 and TAK1 during S. flexneri infection. Furthermore, this study validates the use of bacterial effectors as molecular tools to identify the crosstalks that connect important host signaling pathways induced upon bacterial infection. Copyright © 2011 Elsevier Inc. All rights reserved.

HLA type and immune response to Borrelia burgdorferi outer surface protein a in people in whom arthritis developed after Lyme disease vaccination.

PubMed

Ball, Robert; Shadomy, Sean V; Meyer, Abbie; Huber, Brigitte T; Leffell, Mary S; Zachary, Andrea; Belotto, Michael; Hilton, Eileen; Bryant-Genevier, Marthe; Schriefer, Martin E; Miller, Frederick W; Braun, M Miles

2009-04-01

To investigate whether persons with treatment-resistant Lyme arthritis-associated HLA alleles might develop arthritis as a result of an autoimmune reaction triggered by Borrelia burgdorferi outer surface protein A (OspA), the Lyme disease vaccine antigen. Persons in whom inflammatory arthritis had developed after Lyme disease vaccine (cases) were compared with 3 control groups: 1) inflammatory arthritis but not Lyme disease vaccine (arthritis controls), 2) Lyme disease vaccine but not inflammatory arthritis (vaccine controls), and 3) neither Lyme disease vaccine nor inflammatory arthritis (normal controls). HLA-DRB1 allele typing, Western blotting for Lyme antigen, and T cell reactivity testing were performed. Twenty-seven cases were matched with 162 controls (54 in each control group). Odds ratios (ORs) for the presence of 1 or 2 treatment-resistant Lyme arthritis alleles were 0.8 (95% confidence interval [95% CI] 0.3-2.1), 1.6 (95% CI 0.5-4.4), and 1.75 (95% CI 0.6-5.3) in cases versus arthritis controls, vaccine controls, and normal controls, respectively. There were no significant differences in the frequency of DRB1 alleles. T cell response to OspA was similar between cases and vaccine controls, as measured using the stimulation index (OR 1.6 [95% CI 0.5-5.1]) or change in uptake of tritiated thymidine (counts per minute) (OR 0.7 [95% CI 0.2-2.3]), but cases were less likely to have IgG antibodies to OspA (OR 0.3 [95% CI 0.1-0.8]). Cases were sampled closer to the time of vaccination (median 3.59 years versus 5.48 years), and fewer cases had received 3 doses of vaccine (37% versus 93%). Treatment-resistant Lyme arthritis alleles were not found more commonly in persons who developed arthritis after Lyme disease vaccination, and immune responses to OspA were not significantly more common in arthritis cases. These results suggest that Lyme disease vaccine is not a major factor in the development of arthritis in these cases.

HLA Type and Immune Response to Borrelia burgdorferi Outer Surface Protein A in People in Whom Arthritis Developed After Lyme Disease Vaccination

PubMed Central

Ball, Robert; Shadomy, Sean V.; Meyer, Abbie; Huber, Brigitte T.; Leffell, Mary S.; Zachary, Andrea; Belotto, Michael; Hilton, Eileen; Bryant-Genevier, Marthe; Schriefer, Martin E.; Miller, Frederick W.; Braun, M. Miles

2009-01-01

Objective To investigate whether persons with treatment-resistant Lyme arthritis–associated HLA alleles might develop arthritis as a result of an autoimmune reaction triggered by Borrelia burgdorferi outer surface protein A (OspA), the Lyme disease vaccine antigen. Methods Persons in whom inflammatory arthritis had developed after Lyme disease vaccine (cases) were compared with 3 control groups: 1) inflammatory arthritis but not Lyme disease vaccine (arthritis controls), 2) Lyme disease vaccine but not inflammatory arthritis (vaccine controls), and 3) neither Lyme disease vaccine nor inflammatory arthritis (normal controls). HLA–DRB1 allele typing, Western blotting for Lyme antigen, and T cell reactivity testing were performed. Results Twenty-seven cases were matched with 162 controls (54 in each control group). Odds ratios (ORs) for the presence of 1 or 2 treatment-resistant Lyme arthritis alleles were 0.8 (95% confidence interval [95% CI] 0.3–2.1), 1.6 (95% CI 0.5–4.4), and 1.75 (95% CI 0.6–5.3) in cases versus arthritis controls, vaccine controls, and normal controls, respectively. There were no significant differences in the frequency of DRB1 alleles. T cell response to OspA was similar between cases and vaccine controls, as measured using the stimulation index (OR 1.6 [95% CI 0.5–5.1]) or change in uptake of tritiated thymidine (counts per minute) (OR 0.7 [95% CI 0.2–2.3]), but cases were less likely to have IgG antibodies to OspA (OR 0.3 [95% CI 0.1–0.8]). Cases were sampled closer to the time of vaccination (median 3.59 years versus 5.48 years), and fewer cases had received 3 doses of vaccine (37% versus 93%). Conclusion Treatment-resistant Lyme arthritis alleles were not found more commonly in persons who developed arthritis after Lyme disease vaccination, and immune responses to OspA were not significantly more common in arthritis cases. These results suggest that Lyme disease vaccine is not a major factor in the development of arthritis in these cases. PMID:19333928

The OsPS1-F gene regulates growth and development in rice by modulating photosynthetic electron transport rate.

PubMed

Ramamoorthy, Rengasamy; Vishal, Bhushan; Ramachandran, Srinivasan; Kumar, Prakash P

2018-02-01

Ds insertion in rice OsPS1-F gene results in semi-dwarf plants with reduced tiller number and grain yield, while genetic complementation with OsPS1-F rescued the mutant phenotype. Photosynthetic electron transport is regulated in the chloroplast thylakoid membrane by multi-protein complexes. Studies about photosynthetic machinery and its subunits in crop plants are necessary, because they could be crucial for yield enhancement in the long term. Here, we report the characterization of OsPS1-F (encoding Oryza sativa PHOTOSYSTEM 1-F subunit) using a single copy Ds insertion rice mutant line. The homozygous mutant (osps1-f) showed striking difference in growth and development compared to the wild type (WT), including, reduction in plant height, tiller number, grain yield as well as pale yellow leaf coloration. Chlorophyll concentration and electron transport rate were significantly reduced in the mutant compared to the WT. OsPS1-F gene was highly expressed in rice leaves compared to other tissues at different developmental stages tested. Upon complementation of the mutant with proUBI::OsPS1-F, the observed mutant phenotypes were rescued. Our results illustrate that OsPS1-F plays an important role in regulating proper growth and development of rice plants.

Bacterin That Induces Anti-OspA and Anti-OspC Borreliacidal Antibodies Provides a High Level of Protection against Canine Lyme Disease▿

PubMed Central

LaFleur, Rhonda L.; Dant, Jennifer C.; Wasmoen, Terri L.; Callister, Steven M.; Jobe, Dean A.; Lovrich, Steven D.; Warner, Thomas F.; Abdelmagid, O.; Schell, Ronald F.

2009-01-01

Groups of 15 laboratory-bred beagles were vaccinated and boosted with either a placebo or adjuvanted bivalent bacterin comprised of a traditional Borrelia burgdorferi strain and a unique ospA- and ospB-negative B. burgdorferi strain that expressed high levels of OspC and then challenged with B. burgdorferi-infected Ixodes scapularis ticks. The vaccinated dogs produced high titers of anti-OspA and anti-OspC borreliacidal antibodies, including borreliacidal antibodies specific for an epitope within the last seven amino acids at the OspC carboxy terminus (termed OspC7) that was conserved among pathogenic Borrelia genospecies. In addition, spirochetes were eliminated from the infected ticks that fed on the bacterin recipients, B. burgdorferi was not isolated from the skin or joints, and antibody responses associated specifically with canine infection with B. burgdorferi were not produced. In contrast, B. burgdorferi was recovered from engorged ticks that fed on 13 (87%) placebo-vaccinated dogs (P < 0.0001), skin biopsy specimens from 14 (93%) dogs (P < 0.0001), and joint tissue specimens from 8 (53%) dogs (P = 0.0022). In addition, 14 (93%) dogs developed specific antibody responses against B. burgdorferi proteins, including 11 (73%) with C6 peptide antibodies (P < 0.0001). Moreover, 10 (67%) dogs developed Lyme disease-associated joint abnormalities (P < 0.0001), including 4 (27%) dogs that developed joint stiffness or lameness and 6 (40%) that developed chronic joint inflammation (synovitis). The results therefore confirmed that the bacterin provided a high level of protection against Lyme disease shortly after immunization. PMID:19052162

A host-restricted viral vector for antigen-specific immunization against Lyme disease pathogen.

PubMed

Xiao, Sa; Kumar, Manish; Yang, Xiuli; Akkoyunlu, Mustafa; Collins, Peter L; Samal, Siba K; Pal, Utpal

2011-07-18

Newcastle disease virus (NDV) is an avian virus that is attenuated in primates and is a potential vaccine vector for human use. We evaluated NDV as a vector for expressing selected antigens of the Lyme disease pathogen Borrelia burgdorferi. A series of recombinant NDVs were generated that expressed intracellular or extracellular forms of two B. burgdorferi antigens: namely, the basic membrane protein A (BmpA) and the outer surface protein C (OspC). Expression of the intracellular and extracellular forms of these antigens was confirmed in cultured chicken cells. C3H or Balb/C mice that were immunized intranasally with the NDV vectors mounted vigorous serum antibody responses against the NDV vector, but failed to mount a robust response against either the intracellular or extracellular forms of BmpA or OspC. By contrast, a single immunization of hamsters with the NDV vectors via the intranasal, intramuscular, or intraperitoneal route resulted in rapid and rigorous antibody responses against the intracellular or extracellular forms of BmpA and OspC. When groups of hamsters were separately inoculated with various NDV vectors and challenged with B. burgdorferi (10(8)cells/animal), immunization with vector expressing either intracellular or extracellular BmpA was associated with a significant reduction of the pathogen load in the joints. Taken together, our studies highlighted the importance of NDV as vaccine vector that can be used for simple yet effective immunization of hosts against bacterial infections including Lyme disease. Copyright © 2011 Elsevier Ltd. All rights reserved.

Effect of Epoxy on Mechanical Property of SAC305 Solder Joint with Various Surface Finishes Under 3-Point Bend Test.

PubMed

Jeong, Haksan; Myung, Woo-Ram; Sung, Yong-Gue; Kim, Kyung-Yeol; Jung, Seung-Boo

2018-09-01

Microstructures and mechanical property of Sn-3.0Ag-0.5Cu (SAC305) and epoxy Sn-3.0Ag-0.5Cu (epoxy SAC) solder joints were investigated with various surface finishes; organic solderability preservative (OSP), electroless nickel immersion gold (ENIG) and electroless nickel electroless palladium immersion gold (ENEPIG). Bending property of solder joints was evaluated by 3-point bend test method. Microstructure and chemical composition of solder joints was characterized by scanning electron microscope (SEM) and energy dispersive X-ray spectroscopy (EDX), respectively. Epoxy did not effect on intermetallic compound (IMC) morphology. Scalloped shaped Cu6Sn5 IMC was observed at OSP surface finish. Chunky-like shaped and needle-like shaped (Ni,Cu)6Sn5 IMC were observed at the solder/ENIG joint and solder/ENEPIG joint, respectively. The bending cycles of SAC305/OSP joint, SAC305/ENIG joints and SAC305/ENEPIG joints were 720, 440 and 481 cycle numbers. The bending cycles of epoxy SAC and three types surface finished solder joints were over 1000 bending cycles. Under OSP surface finish, bending cycles of epoxy SAC solder was approximately 1.5 times higher than those of SAC305 solder joint. Bending cycles of epoxy SAC solder was over twice times higher than those of SAC305 solder with ENIG and ENEPIG surface finishes. The bending property of epoxy solder joint was enhanced due to epoxy fillet held the solder joint.

Improved Serodiagnostic Performance for Lyme Disease by Use of Two Recombinant Proteins in Enzyme-Linked Immunosorbent Assay Compared to Standardized Two-Tier Testing.

PubMed

Bradshaw, Gary L; Thueson, R Kelley; Uriona, Todd J

2017-10-01

The most reliable test method for the serological confirmation of Lyme disease (LD) is a 2-tier method recommended by the CDC in 1995. The first-tier test is a low-specificity enzyme-linked immunosorbent assay (ELISA), and the second-tier tests are higher-specificity IgG and IgM Western blots. This study describes the selection of two Borrelia burgdorferi recombinant proteins and evaluation of their performance in a simple 1-tier test for the serological confirmation of LD. These two proteins were generated from (i) the full-length dbpA gene combined with the invariable region 6 of the vlsE gene (DbpA/C6) and (b) the full-length ospC gene (OspC). The expressed DbpA/C6 and OspC proteins were useful in detecting anti- Borrelia IgG and IgM antibodies, respectively. A blind study was conducted on a well-characterized panel of 279 human sera from the CDC, comparing ELISAs using these two recombinant antigens with the 2-tier test method. The two methods (DbpA/C6-OspC versus 2-tier test) were equivalent in identifying sera from negative-control subjects (99% and 100% specificity, respectively) and in detecting stage II and III LD patient sera (100% and 100% sensitivity). However, the DbpA/C6-OspC ELISA was markedly better (80% versus 63%) than the 2-tier test method in detecting anti- Borrelia antibodies in stage I LD patients. The findings suggest that these antigens could be used in a simple 1-tier ELISA that is faster to perform, easier to interpret, and less expensive than the 2-tier test method and which is better at detecting Borrelia -specific antibodies in sera from patients with stage I LD. Copyright © 2017 Bradshaw et al.

The effect of surface characteristics on the transport of multiple Escherichia coli isolates in large scale columns of quartz sand.

PubMed

Lutterodt, G; Basnet, M; Foppen, J W A; Uhlenbrook, S

2009-02-01

Bacteria properties play an important role in the transport of bacteria in groundwater, but their role, especially for longer transport distances (>0.5 m) has not been studied. Thereto, we studied the effects of cell surface hydrophobicity, outer surface potential (OSP), cell sphericity, motility, and Ag43 protein expression on the outer cell surface for a number of E. coli strains, obtained from the environment on their transport behavior in columns of saturated quartz sand of 5 m height in two solutions: demineralized (DI) water and artificial groundwater (AGW). In DI water, sticking efficiencies ranged between 0.1 and 0.4 at the column inlet, and then decreased with transport distance to 0.02-0.2. In AGW, sticking efficiencies were on average 1log-unit higher than those in DI (water). Bacteria motility and Ag43 expression affected attachment with a (high) statistical significance. In contrast, hydrophobicity, OSP and cell sphericity did not significantly correlate with sticking efficiency. However, for transport distances more than 0.33 m, the correlation between sticking efficiency, Ag43 expression, and motility became insignificant. We concluded that Ag43 and motility played an important role in E. coli attachment to quartz grain surfaces, and that the transport distance dependent sticking efficiency reductions were caused by motility and Ag43 expression variations within a population. The implication of our findings is that less motile bacteria with little or no Ag43 expression may travel longer distances once they enter groundwater environments. In future studies, the possible effect of bacteria surface structures, like fimbriae, pili and surface proteins on bacteria attachment need to be considered more systematically in order to arrive at more meaningful inter-population comparisons of the transport behavior of E. coli strains in aquifers.

Discovery and Targeted Proteomics on Cutaneous Biopsies Infected by Borrelia to Investigate Lyme Disease*

PubMed Central

Schnell, Gilles; Boeuf, Amandine; Westermann, Benoît; Jaulhac, Benoît; Lipsker, Dan; Carapito, Christine; Boulanger, Nathalie; Ehret-Sabatier, Laurence

2015-01-01

Lyme disease is the most important vector-borne disease in the Northern hemisphere and represents a major public health challenge with insufficient means of reliable diagnosis. Skin is rarely investigated in proteomics but constitutes in the case of Lyme disease the key interface where the pathogens can enter, persist, and multiply. Therefore, we investigated proteomics on skin samples to detect Borrelia proteins directly in cutaneous biopsies in a robust and specific way. We first set up a discovery gel prefractionation-LC-MS/MS approach on a murine model infected by Borrelia burgdorferi sensu stricto that allowed the identification of 25 Borrelia proteins among more than 1300 mouse proteins. Then we developed a targeted gel prefractionation-LC-selected reaction monitoring (SRM) assay to detect 9/33 Borrelia proteins/peptides in mouse skin tissue samples using heavy labeled synthetic peptides. We successfully transferred this assay from the mouse model to human skin biopsies (naturally infected by Borrelia), and we were able to detect two Borrelia proteins: OspC and flagellin. Considering the extreme variability of OspC, we developed an extended SRM assay to target a large set of variants. This assay afforded the detection of nine peptides belonging to either OspC or flagellin in human skin biopsies. We further shortened the sample preparation and showed that Borrelia is detectable in mouse and human skin biopsies by directly using a liquid digestion followed by LC-SRM analysis without any prefractionation. This study thus shows that a targeted SRM approach is a promising tool for the early direct diagnosis of Lyme disease with high sensitivity (<10 fmol of OspC/mg of human skin biopsy). PMID:25713121

Induction of Experimental Arthritis by Borrelial Lipoprotein and CpG Motifs: Are Toll-Like Receptors 2, 4, 9 or CD-14 Involved?

DOE Office of Scientific and Technical Information (OSTI.GOV)

Batsford, S.; Dunn, J.; Mihatsch, M.

Bacterial lipoproteins and CpG-DNA are ligands for Toll-Like-Receptors (TLR) 2 and 9 respectively. Both classes of molecules were reported to induce experimental arthritis in rodents following direct intra-articular injection. Here we studied: (1) whether arthritis induction by Outer surface (Lipo)protein A (OspA) (B.burgdorferi) involved the TLR-2 as well as the TLR-4 or the CD-14 receptors in addition, and (2) re-examined the arthritogenic potential of CpG-DNA motifs in mice. Following intra-articular injection of the test substances [20 {micro}g recombinant, lipidated OspA; 1nM(6 {micro}g) to 10nM(60 {micro}g) synthetic CpG-DNA], inflammation was monitored by {sup 99}Tc scintigraphy (ratio left/right knee joint uptake >more » 1.1 indicates inflammation) and by histology. Lipoprotein OspA induced severe, acute arthritis in TLR-2{sup +/+} w.t. but not in TLR-2{sup -/-} mice (p<0.01). There were no significant differences in the severity of arthritis induced in TLR-4{sup +/+} w.t. and TLR-4{sup -/-} mutant mice, or between CD14{sup +/+} w.t. and CD14{sup -/-} mice. CpG-DNA (1or 10 nM) did not cause notable inflammation in C57BL/6 mice; {sup 99}Tc ratios were < 1.0 and histology showed only minimal changes. Induction of arthritis by the OspA lipoprotein of B.burgdorferi involves the TLR-2 receptor, no evidence for additional participation of TLR-4 or CD14 receptors was found. Intra-articular injection of CpG-DNA did not produce manifest joint injury in mice, at variance with previous reports.« less

Dust remobilization tests in DIII-D divertor

NASA Astrophysics Data System (ADS)

Bykov, I.; Rudakov, D.; Moyer, R.; Ratynskaia, S.; Tolias, P.; Deangeli, M.; McLean, A.; Bystrov, K.

2015-11-01

Accumulation of dust on hot surfaces is a safety concern for ITER operation. We studied the life cycle of pre-deposited dust under ITER-relevant conditions by exposing W samples with W, C and Al (surrogate for Be) dust at the outer strike point (OSP) in a few ELMy H-mode discharges using DiMES. The maxima in the dust ejection rate correspond to ELM crashes under both attached and detached OSP conditions, as confirmed by a fast camera monitoring DiMES. SEM mapping of dust before and after exposures shows that >95 % of C and <5 % of metal dust gets remobilized in a few shots. In discharges with detached OSP, remaining Al particles melt and fuse together, forming larger spherical grains. At elevated heat flux with attached OSP, they melt, destruct and fuse with W substrate, which is not thermally affected. In this mode W grains partly melt and adjacent particles can weld together, forming larger asymmetric agglomerates with increased adhesion to the surface. We show that these results are consistent with recent observations from Pilot-PSI. Work supported by the US DOE under DE-FC02-04ER54698, DE-FG02-07ER54917 and DE-AC52-07NA27344.

Low prevalence of Borrelia bavariensis in Ixodes ricinus ticks in southeastern Austria.

PubMed

Glatz, Martin; Muellegger, Robert R; Hizo-Teufel, Cecilia; Fingerle, Volker

2014-10-01

Borrelia bavariensis was recently described as a distinct genospecies among the B. burgdorferi sensu lato complex. The prevalence of B. bavariensis in Austria, a highly endemic area for tick-transmitted pathogens, is scarcely characterized. To investigate the prevalence of B. bavariensis in Ixodes ricinus ticks we reevaluated the results of a study conducted in 518 ticks from southeastern Austria collected in 2002 and 2003. The presence of B. burgdorferi s.l.-specific DNA in ticks was analyzed by a PCR for the outer surface protein A (ospA) gene. Borrelia species were differentiated by restriction fragment length polymorphism (RFLP) analysis, and samples positive for B. bavariensis were further analyzed by multilocus sequence analysis. Two of 133 (1.5%) B. burgdorferi s.l.-positive I. ricinus ticks were infected with B. bavariensis. Both specimens were coinfected with the OspA serotype 5 of B. garinii. Borrelia bavariensis is present; however, seem to be rare in I. ricinus ticks in southeastern Austria. Copyright © 2014 Elsevier GmbH. All rights reserved.

Soluble cysteine-rich tick saliva proteins Salp15 and Iric-1 from E. coli

PubMed Central

Kolb, Philipp; Vorreiter, Jolanta; Habicht, Jüri; Bentrop, Detlef; Wallich, Reinhard; Nassal, Michael

2014-01-01

Ticks transmit numerous pathogens, including borreliae, which cause Lyme disease. Tick saliva contains a complex mix of anti-host defense factors, including the immunosuppressive cysteine-rich secretory glycoprotein Salp15 from Ixodes scapularis ticks and orthologs like Iric-1 from Ixodesricinus. All tick-borne microbes benefit from the immunosuppression at the tick bite site; in addition, borreliae exploit the binding of Salp15 to their outer surface protein C (OspC) for enhanced transmission. Hence, Salp15 proteins are attractive targets for anti-tick vaccines that also target borreliae. However, recombinant Salp proteins are not accessible in sufficient quantity for either vaccine manufacturing or for structural characterization. As an alternative to low-yield eukaryotic systems, we investigated cytoplasmic expression in Escherichia coli, even though this would not result in glycosylation. His-tagged Salp15 was efficiently expressed but insoluble. Among the various solubility-enhancing protein tags tested, DsbA was superior, yielding milligram amounts of soluble, monomeric Salp15 and Iric-1 fusions. Easily accessible mutants enabled epitope mapping of two monoclonal antibodies that, importantly, cross-react with glycosylated Salp15, and revealed interaction sites with OspC. Free Salp15 and Iric-1 from protease-cleavable fusions, despite limited solubility, allowed the recording of 1H–15N 2D NMR spectra, suggesting partial folding of the wild-type proteins but not of Cys-free variants. Fusion to the NMR-compatible GB1 domain sufficiently enhanced solubility to reveal first secondary structure elements in 13C/15N double-labeled Iric-1. Together, E. coli expression of appropriately fused Salp15 proteins may be highly valuable for the molecular characterization of the function and eventually the 3D structure of these medically relevant tick proteins. PMID:25628987

Soluble cysteine-rich tick saliva proteins Salp15 and Iric-1 from E. coli.

PubMed

Kolb, Philipp; Vorreiter, Jolanta; Habicht, Jüri; Bentrop, Detlef; Wallich, Reinhard; Nassal, Michael

2015-01-01

Ticks transmit numerous pathogens, including borreliae, which cause Lyme disease. Tick saliva contains a complex mix of anti-host defense factors, including the immunosuppressive cysteine-rich secretory glycoprotein Salp15 from Ixodes scapularis ticks and orthologs like Iric-1 from Ixodes ricinus. All tick-borne microbes benefit from the immunosuppression at the tick bite site; in addition, borreliae exploit the binding of Salp15 to their outer surface protein C (OspC) for enhanced transmission. Hence, Salp15 proteins are attractive targets for anti-tick vaccines that also target borreliae. However, recombinant Salp proteins are not accessible in sufficient quantity for either vaccine manufacturing or for structural characterization. As an alternative to low-yield eukaryotic systems, we investigated cytoplasmic expression in Escherichia coli, even though this would not result in glycosylation. His-tagged Salp15 was efficiently expressed but insoluble. Among the various solubility-enhancing protein tags tested, DsbA was superior, yielding milligram amounts of soluble, monomeric Salp15 and Iric-1 fusions. Easily accessible mutants enabled epitope mapping of two monoclonal antibodies that, importantly, cross-react with glycosylated Salp15, and revealed interaction sites with OspC. Free Salp15 and Iric-1 from protease-cleavable fusions, despite limited solubility, allowed the recording of (1)H-(15)N 2D NMR spectra, suggesting partial folding of the wild-type proteins but not of Cys-free variants. Fusion to the NMR-compatible GB1 domain sufficiently enhanced solubility to reveal first secondary structure elements in (13)C/(15)N double-labeled Iric-1. Together, E. coli expression of appropriately fused Salp15 proteins may be highly valuable for the molecular characterization of the function and eventually the 3D structure of these medically relevant tick proteins.

Experimental infections of the reservoir species Peromyscus leucopus with diverse strains of Borrelia burgdorferi, a Lyme disease agent.

PubMed

Baum, Elisabeth; Hue, Fong; Barbour, Alan G

2012-12-04

The rodent Peromyscus leucopus is a major natural reservoir for the Lyme disease agent Borrelia burgdorferi and a host for its vector Ixodes scapularis. At various locations in northeastern United States 10 to 15 B. burgdorferi strains coexist at different prevalences in tick populations. We asked whether representative strains of high or low prevalence differed in their infections of P. leucopus. After 5 weeks of experimental infection of groups with each of 6 isolates, distributions and burdens of bacteria in tissues were measured by quantitative PCR, and antibodies to B. burgdorferi were evaluated by immunoblotting and protein microarray. All groups of animals were infected in their joints, ears, tails, and hearts, but overall spirochete burdens were lower in animals infected with low-prevalence strains. Animals were similar regardless of the infecting isolate in their levels of antibodies to whole cells, FlaB, BmpA, and DbpB proteins, and the conserved N-terminal region of the serotype-defining OspC proteins. But there were strain-specific antibody responses to full-length OspC and to plasmid-encoded VlsE, BBK07, and BBK12 proteins. Sequencing of additional VlsE genes revealed substantial diversity within some pairs of strains but near-identical sequences within other pairs, which otherwise differed in their ospC alleles. The presence or absence of full-length bbk07 and bbk12 genes accounted for the differences in antibody responses. We propose that for B. burgdorferi, there is selection in reservoir species for (i) sequence diversity, as for OspC and VlsE, and (ii) the presence or absence of polymorphisms, as for BBK07 and BBK12. Humans are dead-end hosts for Borrelia agents of Lyme disease (LD), and, thus, irrelevant for the pathogens' maintenance. Many reports of human cases and laboratory mouse infections exist, but less is known about infection and immunity in natural reservoirs, such as the rodent Peromyscus leucopus. We observed that high- and low-prevalence strains of Borrelia burgdorferi were capable of infecting P. leucopus but elicited different patterns of antibody responses. Antibody reactivities to the VlsE protein were as type-specific as previously characterized reactivities to serotype-defining OspC proteins. In addition, the low-prevalence strains lacked full-length genes for two proteins that (i) are encoded by a virulence-associated plasmid in some high-prevalence strains and (ii) LD patients and field-captured rodents commonly have antibodies to. Immune selection against these genes may have led to null phenotype lineages that can infect otherwise immune hosts but at the cost of reduced fitness and lower prevalence.

Lyme disease risk not amplified in a species-poor vertebrate community: similar Borrelia burgdorferi tick infection prevalence and OspC genotype frequencies.

PubMed

States, S L; Brinkerhoff, R J; Carpi, G; Steeves, T K; Folsom-O'Keefe, C; DeVeaux, M; Diuk-Wasser, M A

2014-10-01

The effect of biodiversity declines on human health is currently debated, but empirical assessments are lacking. Lyme disease provides a model system to assess relationships between biodiversity and human disease because the etiologic agent, Borrelia burgdorferi, is transmitted in the United States by the generalist black-legged tick (Ixodes scapularis) among a wide range of mammalian and avian hosts. The 'dilution effect' hypothesis predicts that species-poor host communities dominated by white-footed mice (Peromyscus leucopus) will pose the greatest human risk because P. leucopus infects the largest numbers of ticks, resulting in higher human exposure to infected I. scapularis ticks. P. leucopus-dominated communities are also expected to maintain a higher frequency of those B. burgdorferi outer surface protein C (ospC) genotypes that this host species more efficiently transmits ('multiple niche polymorphism' hypothesis). Because some of these genotypes are human invasive, an additive increase in human disease risk is expected in species-poor settings. We assessed these theoretical predictions by comparing I. scapularis nymphal infection prevalence, density of infected nymphs and B. burgdorferi genotype diversity at sites on Block Island, RI, where P. leucopus dominates the mammalian host community, to species-diverse sites in northeastern Connecticut. We found no support for the dilution effect hypothesis; B. burgdorferi nymphal infection prevalence was similar between island and mainland and the density of B. burgdorferi infected nymphs was higher on the mainland, contrary to what is predicted by the dilution effect hypothesis. Evidence for the multiple niche polymorphism hypothesis was mixed: there was lower ospC genotype diversity at island than mainland sites, but no overrepresentation of genotypes with higher fitness in P. leucopus or that are more invasive in humans. We conclude that other mechanisms explain similar nymphal infection prevalence in both communities and that high ospC genotype diversity can be maintained in both species-poor and species-rich communities. Copyright © 2014 Elsevier B.V. All rights reserved.

The downward flux of biogenic material in the NE subarctic Pacific: importance of algal sinking and mesozooplankton herbivory

NASA Astrophysics Data System (ADS)

Thibault, D.; Roy, S.; Wong, C. S.; Bishop, J. K.

1999-11-01

In the present study we examine factors that affect the downward flux of biogenic carbon in the NE subarctic Pacific, one of the important high-nutrient-low-chlorophyll (HNLC) regions in the open ocean. We focus on the role of mesozooplankton, since their seasonal peaks in biomass and growth are in phase with the seasonal variations in the downward POC fluxes, whereas phytoplankton biomass is more-or-less uniform year-round. The relative importance of mesozooplankton and algal sinking was examined using the pigment composition of material accumulated in short-term free-drifting sediment traps positioned just below the upper stratified surface layer (ca. 100-200 m). This was compared with the phytoplankton composition in the surface waters, and with the grazing activity (gut pigments and fecal pellet production rates) of the most abundant large copepods. We also examined whether the relationships between the downward flux of carbon and pelagic processes were similar in the coastal, continental margin and offshore HNLC regions of the NE subarctic Pacific, the latter represented by Ocean Station Papa (OSP). Our results show that grazing had a variable impact on the downward flux of biogenic carbon. Carbon-transformed pheopigments (particularly pyropheophorbide a, frequently associated with copepod grazing) represented up to 13% of the total downward POC flux inshore (in May 1996) and 8-9% at OSP in May and February 1996, respectively. This flux of pheopigments was accompanied by a large potential input of fecal pellets from large copepods (as estimated from defecation rates of freshly collected animals) only in May 1996 at OSP, suggesting that pheopigments came from other sources (other herbivores, senescing algae) in February. The larger flux of pheopigments in May was probably related to the abundance of mesozooplankton at that time of the year. During summer (August 1996), both the flux of pheopigments and the potential input of fecal pellets from large copepods were negligible at OSP, consistent with more intense pelagic recycling reported in other studies. Inshore, the flux of carbon-transformed pheopigments was slightly higher than at OSP, and its contribution to the downward POC flux in May 1996 was twice that in August 1996. In contrast, the potential input of feces carbon was higher in August than in May 1996, again suggesting other sources for pheopigments found in the traps. The contribution of sinking phytoplankton to the downward biogenic flux was negligible in summer, when prymnesiophytes (indicated by the presence of 19'-hexanoyloxyfucoxanthin) and pelagophytes (19'-butanoyloxyfucoxanthin-containing) dominated in surface offshore waters. The contribution of sinking algae was maximal (9%) in winter (February 1996) at OSP, when fucoxanthin (mainly a diatom marker) dominated the carotenoid composition in the traps and when the abundance of diatoms in surface waters showed its seasonal maximum for this station. Inshore, the low contribution of diatoms (fucoxanthin) to the sinking fluxes may have resulted from inadequate sampling (i.e. the spring bloom may have been missed). Overall, we conclude that: (a) large copepods significantly influenced the downward POC flux only during spring at OSP; (b) unidentified herbivores (e.g. salps, pteropods) producing pigmented, fast-sinking fecal material likely had an important impact during winter; (c) algal sinking made a small contribution to the downward POC flux (maximum in winter); and (d) neither algal sinking nor mesozooplankton grazing had a significant influence on the downward flux of biogenic material in summer at OSP.

A Dual Luciferase Reporter System for B. burgdorferi Measures Transcriptional Activity during Tick-Pathogen Interactions

PubMed Central

Adams, Philip P.; Flores Avile, Carlos; Jewett, Mollie W.

2017-01-01

Knowledge of the transcriptional responses of vector-borne pathogens at the vector-pathogen interface is critical for understanding disease transmission. Borrelia (Borreliella) burgdorferi, the causative agent of Lyme disease in the United States, is transmitted by the bite of infected Ixodes sp. ticks. It is known that B. burgdorferi has altered patterns of gene expression during tick acquisition, persistence and transmission. Recently, we and others have discovered in vitro expression of RNAs found internal, overlapping, and antisense to annotated open reading frames in the B. burgdorferi genome. However, there is a lack of molecular genetic tools for B. burgdorferi for quantitative, strand-specific, comparative analysis of these transcripts in distinct environments such as the arthropod vector. To address this need, we have developed a dual luciferase reporter system to quantify B. burgdorferi promoter activities in a strand-specific manner. We demonstrate that constitutive expression of a B. burgdorferi codon-optimized Renilla reniformis luciferase gene (rlucBb) allows normalization of the activity of a promoter of interest when fused to the B. burgdorferi codon-optimized Photinus pyralis luciferase gene (flucBb) on the same plasmid. Using the well characterized, differentially regulated, promoters for flagellin (flaBp), outer surface protein A (ospAp) and outer surface protein C (ospCp), we document the efficacy of the dual luciferase system for quantitation of promoter activities during in vitro growth and in infected ticks. Cumulatively, the dual luciferase method outlined herein is the first dual reporter system for B. burgdorferi, providing a novel and highly versatile approach for strand-specific molecular genetic analyses. PMID:28620587

A Dual Luciferase Reporter System for B. burgdorferi Measures Transcriptional Activity during Tick-Pathogen Interactions.

PubMed

Adams, Philip P; Flores Avile, Carlos; Jewett, Mollie W

2017-01-01

Knowledge of the transcriptional responses of vector-borne pathogens at the vector-pathogen interface is critical for understanding disease transmission. Borrelia ( Borreliella ) burgdorferi , the causative agent of Lyme disease in the United States, is transmitted by the bite of infected Ixodes sp . ticks. It is known that B. burgdorferi has altered patterns of gene expression during tick acquisition, persistence and transmission. Recently, we and others have discovered in vitro expression of RNAs found internal, overlapping, and antisense to annotated open reading frames in the B. burgdorferi genome. However, there is a lack of molecular genetic tools for B. burgdorferi for quantitative, strand-specific, comparative analysis of these transcripts in distinct environments such as the arthropod vector. To address this need, we have developed a dual luciferase reporter system to quantify B. burgdorferi promoter activities in a strand-specific manner. We demonstrate that constitutive expression of a B. burgdorferi codon-optimized Renilla reniformis luciferase gene ( rluc Bb ) allows normalization of the activity of a promoter of interest when fused to the B. burgdorferi codon-optimized Photinus pyralis luciferase gene ( fluc Bb ) on the same plasmid. Using the well characterized, differentially regulated, promoters for flagellin ( flaBp ), outer surface protein A ( ospAp ) and outer surface protein C ( ospCp ), we document the efficacy of the dual luciferase system for quantitation of promoter activities during in vitro growth and in infected ticks. Cumulatively, the dual luciferase method outlined herein is the first dual reporter system for B. burgdorferi , providing a novel and highly versatile approach for strand-specific molecular genetic analyses.

Role of a Novel Human Leukocyte Antigen-DQA1*01:02;DRB1*15:01 Mixed Isotype Heterodimer in the Pathogenesis of “Humanized” Multiple Sclerosis-like Disease*

PubMed Central

Kaushansky, Nathali; Eisenstein, Miriam; Boura-Halfon, Sigalit; Hansen, Bjarke Endel; Nielsen, Claus Henrik; Milo, Ron; Zeilig, Gabriel; Lassmann, Hans; Altmann, Daniel M.; Ben-Nun, Avraham

2015-01-01

Gene-wide association and candidate gene studies indicate that the greatest effect on multiple sclerosis (MS) risk is driven by the HLA-DRB1*15:01 allele within the HLA-DR15 haplotype (HLA-DRB1*15:01-DQA1*01:02-DQB1*0602-DRB5*01:01). Nevertheless, linkage disequilibrium makes it difficult to define, without functional studies, whether the functionally relevant effect derives from DRB1*15:01 only, from its neighboring DQA1*01:02-DQB1*06:02 or DRB5*01:01 genes of HLA-DR15 haplotype, or from their combinations or epistatic interactions. Here, we analyzed the impact of the different HLA-DR15 haplotype alleles on disease susceptibility in a new “humanized” model of MS induced in HLA-transgenic (Tg) mice by human oligodendrocyte-specific protein (OSP)/claudin-11 (hOSP), one of the bona fide potential primary target antigens in MS. We show that the hOSP-associated MS-like disease is dominated by the DRB1*15:01 allele not only as the DRA1*01:01;DRB1*15:01 isotypic heterodimer but also, unexpectedly, as a functional DQA1*01:02;DRB1*15:01 mixed isotype heterodimer. The contribution of HLA-DQA1/DRB1 mixed isotype heterodimer to OSP pathogenesis was revealed in (DRB1*1501xDQB1*0602)F1 double-Tg mice immunized with hOSP(142–161) peptide, where the encephalitogenic potential of prevalent DRB1*1501/hOSP(142–161)-reactive Th1/Th17 cells is hindered due to a single amino acid difference in the OSP(142–161) region between humans and mice; this impedes binding of DRB1*1501 to the mouse OSP(142–161) epitope in the mouse CNS while exposing functional binding of mouse OSP(142–161) to DQA1*01:02;DRB1*15:01 mixed isotype heterodimer. This study, which shows for the first time a functional HLA-DQA1/DRB1 mixed isotype heterodimer and its potential association with disease susceptibility, provides a rationale for a potential effect on MS risk from DQA1*01:02 through functional DQA1*01:02;DRB1*15:01 antigen presentation. Furthermore, it highlights a potential contribution to MS risk also from interisotypic combination between products of neighboring HLA-DR15 haplotype alleles, in this case the DQA1/DRB1 combination. PMID:25911099

Host Immune Evasion by Lyme and Relapsing Fever Borreliae: Findings to Lead Future Studies for Borrelia miyamotoi

PubMed Central

Stone, Brandee L.; Brissette, Catherine A.

2017-01-01

The emerging pathogen, Borrelia miyamotoi, is a relapsing fever spirochete vectored by the same species of Ixodes ticks that carry the causative agents of Lyme disease in the US, Europe, and Asia. Symptoms caused by infection with B. miyamotoi are similar to a relapsing fever infection. However, B. miyamotoi has adapted to different vectors and reservoirs, which could result in unique physiology, including immune evasion mechanisms. Lyme Borrelia utilize a combination of Ixodes-produced inhibitors and native proteins [i.e., factor H-binding proteins (FHBPs)/complement regulator-acquiring surface proteins, p43, BBK32, BGA66, BGA71, CD59-like protein] to inhibit complement, while some relapsing fever spirochetes use C4b-binding protein and likely Ornithodoros-produced inhibitors. To evade the humoral response, Borrelia utilize antigenic variation of either outer surface proteins (Osps) and the Vmp-like sequences (Vls) system (Lyme borreliae) or variable membrane proteins (Vmps, relapsing fever borreliae). B. miyamotoi possesses putative FHBPs and antigenic variation of Vmps has been demonstrated. This review summarizes and compares the common mechanisms utilized by Lyme and relapsing fever spirochetes, as well as the current state of understanding immune evasion by B. miyamotoi. PMID:28154563

Host Immune Evasion by Lyme and Relapsing Fever Borreliae: Findings to Lead Future Studies for Borrelia miyamotoi.

PubMed

Stone, Brandee L; Brissette, Catherine A

2017-01-01

The emerging pathogen, Borrelia miyamotoi , is a relapsing fever spirochete vectored by the same species of Ixodes ticks that carry the causative agents of Lyme disease in the US, Europe, and Asia. Symptoms caused by infection with B. miyamotoi are similar to a relapsing fever infection. However, B. miyamotoi has adapted to different vectors and reservoirs, which could result in unique physiology, including immune evasion mechanisms. Lyme Borrelia utilize a combination of Ixodes -produced inhibitors and native proteins [i.e., factor H-binding proteins (FHBPs)/complement regulator-acquiring surface proteins, p43, BBK32, BGA66, BGA71, CD59-like protein] to inhibit complement, while some relapsing fever spirochetes use C4b-binding protein and likely Ornithodoros -produced inhibitors. To evade the humoral response, Borrelia utilize antigenic variation of either outer surface proteins (Osps) and the Vmp-like sequences (Vls) system (Lyme borreliae) or variable membrane proteins (Vmps, relapsing fever borreliae). B. miyamotoi possesses putative FHBPs and antigenic variation of Vmps has been demonstrated. This review summarizes and compares the common mechanisms utilized by Lyme and relapsing fever spirochetes, as well as the current state of understanding immune evasion by B. miyamotoi .

Activating KRAS mutations are characteristic of oncocytic sinonasal papilloma and associated sinonasal squamous cell carcinoma.

PubMed

Udager, Aaron M; McHugh, Jonathan B; Betz, Bryan L; Montone, Kathleen T; Livolsi, Virginia A; Seethala, Raja R; Yakirevich, Evgeny; Iwenofu, O Hans; Perez-Ordonez, Bayardo; DuRoss, Kathleen E; Weigelin, Helmut C; Lim, Megan S; Elenitoba-Johnson, Kojo Sj; Brown, Noah A

2016-08-01

Oncocytic sinonasal papillomas (OSPs) are benign tumours of the sinonasal tract, a subset of which are associated with synchronous or metachronous sinonasal squamous cell carcinoma (SNSCC). Activating EGFR mutations were recently identified in nearly 90% of inverted sinonasal papillomas (ISPs) - a related tumour with distinct morphology. EGFR mutations were, however, not found in OSP, suggesting that different molecular alterations drive the oncogenesis of these tumours. In this study, tissue from 51 cases of OSP and five cases of OSP-associated SNSCC was obtained retrospectively from six institutions. Tissue was also obtained from 50 cases of ISP, 22 cases of ISP-associated SNSCC, ten cases of exophytic sinonasal papilloma (ESP), and 19 cases of SNSCC with no known papilloma association. Using targeted next-generation and conventional Sanger sequencing, we identified KRAS mutations in 51/51 (100%) OSPs and 5/5 (100%) OSP-associated SNSCCs. The somatic nature of KRAS mutations was confirmed in a subset of cases with matched germline DNA, and four matched pairs of OSP and concurrent associated SNSCC had concordant KRAS genotypes. In contrast, KRAS mutations were present in only one (5%) SNSCC with no known papilloma association and none of the ISPs, ISP-associated SNSCCs, or ESPs. This is the first report of somatic KRAS mutations in OSP and OSP-associated SNSCC. The presence of identical mutations in OSP and concurrent associated SNSCC supports the putative role of OSP as a precursor to SNSCC, and the high frequency and specificity of KRAS mutations suggest that OSP and OSP-associated SNSCC are biologically distinct from other similar sinonasal tumours. The identification of KRAS mutations in all studied OSP cases represents an important development in our understanding of the pathogenesis of this disease and may have implications for diagnosis and therapy. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Histopathology and immunophenotype of acrodermatitis chronica atrophicans correlated with ospA and ospC genotypes of Borrelia species.

PubMed

Brandt, Friederike C; Ertas, Beyhan; Falk, Thomas M; Metze, Dieter; Böer-Auer, Almut

2015-10-01

Chronic cutaneous borreliosis (acrodermatitis chronica atrophicans, ACA) is a relatively rare manifestation of borreliosis attributed mainly to Borrelia afzelii. Chronic borreliosis has been associated with ospA and ospC genotypes. Literature on molecular investigations of Borrelia in lesions of ACA is scant. Histopathological and immmunohistochemical features in 22 biopsies of ACA (16 patients) were examined. Paraffin-embedded biopsies were analyzed with polymerase chain reaction (PCR) assays targeting ospA and ospC genes, sequencing and phylogenetic analysis. Genotyping of ospA identified B. afzelii, serotype 2, in 12 of 16 patients. ospC-PCR was positive in seven patients revealing genotypes Af5 (n = 4), Af2 (n = 2) and Af6 (n = 1). Histopathologically, interstitial granulomatous infiltrates (CD68 positive) were common, combined with thickened collagen bundles and band-like infiltrates of CD4 positive T lymphocytes. Plasma cells were sparse/absent in 9 of 22 specimens even on staining with CD138. On CD34-staining, interstitial fibroblasts were often reduced akin to the situation in morphea. With assays targeting ospA and ospC genes we confirmed from paraffin-embedded biopsies that B. afzelii, serotype 2, osp C groups Af5, Af2 and Af6 is the main cause of ACA. Specimens commonly showed a combination of band-like T-cell-rich infiltrates with interstitial granulomatous features, a pattern previously under-recognized in ACA. This finding was particularly typical for lesions infected with ospC genotype Af5. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Human Lipopolysaccharide-binding Protein (LBP) and CD14 Independently Deliver Triacylated Lipoproteins to Toll-like Receptor 1 (TLR1) and TLR2 and Enhance Formation of the Ternary Signaling Complex*

PubMed Central

Ranoa, Diana Rose E.; Kelley, Stacy L.; Tapping, Richard I.

2013-01-01

Bacterial lipoproteins are the most potent microbial agonists for the Toll-like receptor 2 (TLR2) subfamily, and this pattern recognition event induces cellular activation, leading to host immune responses. Triacylated bacterial lipoproteins coordinately bind TLR1 and TLR2, resulting in a stable ternary complex that drives intracellular signaling. The sensitivity of TLR-expressing cells to lipoproteins is greatly enhanced by two lipid-binding serum proteins known as lipopolysaccharide-binding protein (LBP) and soluble CD14 (sCD14); however, the physical mechanism that underlies this increased sensitivity is not known. To address this, we measured the ability of LBP and sCD14 to drive ternary complex formation between soluble extracellular domains of TLR1 and TLR2 and a synthetic triacylated lipopeptide agonist. Importantly, addition of substoichiometric amounts of either LBP or sCD14 significantly enhanced formation of a TLR1·TLR2 lipopeptide ternary complex as measured by size exclusion chromatography. However, neither LBP nor sCD14 was physically associated with the final ternary complex. Similar results were obtained using outer surface protein A (OspA), a naturally occurring triacylated lipoprotein agonist from Borrelia burgdorferi. Activation studies revealed that either LBP or sCD14 sensitized TLR-expressing cells to nanogram levels of either the synthetic lipopeptide or OspA lipoprotein agonist. Together, our results show that either LBP or sCD14 can drive ternary complex formation and TLR activation by acting as mobile carriers of triacylated lipopeptides or lipoproteins. PMID:23430250

From osp(1|32)⊕osp(1|32) to the M-theory superalgebra: a contraction procedure

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fernández, J. J., E-mail: julio.j.fernandez@hotmail.es; Izquierdo, J. M., E-mail: izquierd@fta.uva.es; Olmo, M. A. del, E-mail: olmo@fta.uva.es

We show the impossibility to obtain the D’auria–Fré-type superalgebras that allow for an underlying gauge theoretical structure of D = 11 supergravity from the superalgebra osp(1|32)⊕osp(1|32)−, by means of aWeimar-Woods contraction.

Lusztig symmetries and Poincare-Birkhoff-Witt basis for wU{sub r,s}{sup d}(osp(1|2n))

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Junli; College of Applied Sciences, Beijing University of Technology, Beijing 100124; Yang, Shilin

2013-12-15

We investigate a new kind of two-parameter weak quantized superalgebra wU{sub r,s}{sup d}(osp(1|2n)), which is a weak Hopf superalgebra. It has a homomorphic image which is isomorphic to the usual two-parameter quantum superalgebra U{sub r,s}(osp(1|2n)) of osp(1|2n). We also discuss the basis of wU{sub r,s}{sup d}(osp(1|2n)) by Lusztig's symmetries.

Mechanical Reliability of the Epoxy Sn-58wt.%Bi Solder Joints with Different Surface Finishes Under Thermal Shock

NASA Astrophysics Data System (ADS)

Sung, Yong-Gue; Myung, Woo-Ram; Jeong, Haksan; Ko, Min-Kwan; Moon, Jeonghoon; Jung, Seung-Boo

2018-04-01

The effect of thermal shock on the mechanical reliability of epoxy Sn-58wt.%Bi composite (epoxy Sn-58wt.%Bi) solder joints was investigated with different surface-finished substrates. Sn-58wt.%Bi-based solder has been considered as a promising candidate for low-temperature solder among various lead-free solders. However, Sn-58wt.%Bi solder joints can be easily broken under impact conditions such as mechanical shock, drop tests, and bending tests because of their poor ductility. Therefore, previous researchers have tried to improve the mechanical property of Sn-58wt.%Bi solder by additional elements and mixtures of metal powder and epoxy resin. Epoxy Sn-58wt.%Bi solder paste was fabricated by mixing epoxy resin and Sn-58wt.%Bi solder powder to enhance the mechanical reliability of Sn-58wt.%Bi solder joints. The epoxy Sn-58wt.%Bi solder paste was screen-printed onto various printed circuit board surfaces finished with organic solder preservatives (OSP), electroless nickel immersion gold (ENIG), and electroless nickel electroless palladium immersion gold (ENEPIG). The test components were prepared by a reflow process at a peak temperature of 190°C. The thermal shock test was carried out under the temperature range of - 40 to 125°C to evaluate the reliability of Sn-58wt.%Bi and epoxy Sn-58wt.%Bi solder joints. The OSP-finished sample showed a relatively higher mechanical property than those of ENIG and ENEPIG after thermal shock. The average number of cycles for epoxy Sn-58wt.%Bi solder with the OSP surface finish were 6 times higher than that for Sn-58wt.%Bi solder with the same finish. The microstructures of the solder joints were investigated by scanning electron microscopy, and the composition of the intermetallic compound (IMC) layer was analyzed by using energy dispersive spectrometry. Cu6Sn5 IMC was formed by the reaction between Sn-58wt.%Bi solder and a OSP surface-finished Cu after the reflow process. Ni3Sn4 IMC and (Ni, Pd)3Sn4 IMC were formed at the solder joints between the ENIG and solder, and between ENEPIG surface finish and solders, respectively.

Mechanical Reliability of the Epoxy Sn-58wt.%Bi Solder Joints with Different Surface Finishes Under Thermal Shock

NASA Astrophysics Data System (ADS)

Sung, Yong-Gue; Myung, Woo-Ram; Jeong, Haksan; Ko, Min-Kwan; Moon, Jeonghoon; Jung, Seung-Boo

2018-07-01

The effect of thermal shock on the mechanical reliability of epoxy Sn-58wt.%Bi composite (epoxy Sn-58wt.%Bi) solder joints was investigated with different surface-finished substrates. Sn-58wt.%Bi-based solder has been considered as a promising candidate for low-temperature solder among various lead-free solders. However, Sn-58wt.%Bi solder joints can be easily broken under impact conditions such as mechanical shock, drop tests, and bending tests because of their poor ductility. Therefore, previous researchers have tried to improve the mechanical property of Sn-58wt.%Bi solder by additional elements and mixtures of metal powder and epoxy resin. Epoxy Sn-58wt.%Bi solder paste was fabricated by mixing epoxy resin and Sn-58wt.%Bi solder powder to enhance the mechanical reliability of Sn-58wt.%Bi solder joints. The epoxy Sn-58wt.%Bi solder paste was screen-printed onto various printed circuit board surfaces finished with organic solder preservatives (OSP), electroless nickel immersion gold (ENIG), and electroless nickel electroless palladium immersion gold (ENEPIG). The test components were prepared by a reflow process at a peak temperature of 190°C. The thermal shock test was carried out under the temperature range of - 40 to 125°C to evaluate the reliability of Sn-58wt.%Bi and epoxy Sn-58wt.%Bi solder joints. The OSP-finished sample showed a relatively higher mechanical property than those of ENIG and ENEPIG after thermal shock. The average number of cycles for epoxy Sn-58wt.%Bi solder with the OSP surface finish were 6 times higher than that for Sn-58wt.%Bi solder with the same finish. The microstructures of the solder joints were investigated by scanning electron microscopy, and the composition of the intermetallic compound (IMC) layer was analyzed by using energy dispersive spectrometry. Cu6Sn5 IMC was formed by the reaction between Sn-58wt.%Bi solder and a OSP surface-finished Cu after the reflow process. Ni3Sn4 IMC and (Ni, Pd)3Sn4 IMC were formed at the solder joints between the ENIG and solder, and between ENEPIG surface finish and solders, respectively.

[Recombinant OspC identification and antigenicity detection from Borrelia burgdorferi PD91 in China].

PubMed

Chen, Jian; Wan, Kang-Lin

2003-10-01

To recombine OspC gene from Borrelia burgdorferi PD91 of China and expressed it in E. coli for early diagnosis of Lyme disease. The OspC gene was amplified from the genome of Borrelia burgdorferi PD91 strain by polymerase chain reaction and recombined with plasmid PET-11D. The recombinant plasmid PET-11D-OspC was identified with PCR, restriction endonuclease analysis and sequencing. The antigenicity was verified with Western Blot. OspC gene was cloned correctly into vector PET-11D. The resultant sequence was definitely different from the published sequence. The recombinant OspC seemed to have had strong antigenicity. The findings laid basis for the studies on early diagnosis of Lyme disease.

The Shigella flexneri OspB effector: an early immunomodulator.

PubMed

Ambrosi, Cecilia; Pompili, Monica; Scribano, Daniela; Limongi, Dolores; Petrucca, Andrea; Cannavacciuolo, Sonia; Schippa, Serena; Zagaglia, Carlo; Grossi, Milena; Nicoletti, Mauro

2015-01-01

Through the action of the type three secretion system (T3SS) Shigella flexneri delivers several effectors into host cells to promote cellular invasion, multiplication and to exploit host-cell signaling pathways to modulate the host innate immune response. Although much progress has been made in the understanding of many type III effectors, the molecular and cellular mechanism of the OspB effector is still poorly characterized. In this study we present new evidence that better elucidates the role of OspB as pro-inflammatory factor at very early stages of infection. Indeed, we demonstrate that, during the first hour of infection, OspB is required for full activation of ERK1/2 and p38 MAPKs and the cytosolic phospholipase A(2) (cPLA(2)). Activation of cPLA(2) ultimately leads to the production and secretion of PMN chemoattractant metabolite(s) uncoupled with release of IL-8. Moreover, we also present evidence that OspB is required for the development of the full and promptly inflammatory reaction characteristic of S. flexneri wild-type infection in vivo. Based on OspB and OspF similarity (both effectors share similar transcription regulation, temporal secretion into host cells and nuclear localization) we hypothesized that OspB and OspF effectors may form a pair aimed at modulating the host cell response throughout the infection process, with opposite effects. A model is presented to illustrate how OspB activity would promote S. flexneri invasion and bacterial dissemination at early critical phases of infection. Copyright © 2014 Elsevier GmbH. All rights reserved.

Student's perspectives on objective structured practical examination (OSPE) in Forensic Medicine - a report from India.

PubMed

Pramod Kumar, G N; Sentitoshi; Nath, Dhritiman; Menezes, Ritesh G; Kanchan, Tanuj

2015-05-01

The objective of the present study was to know the perceptions of students regarding objective structured practical examination (OSPE) as a tool for assessment in Forensic Medicine. The present study was conducted in the Department of Forensic Medicine and Toxicology, Mahatma Gandhi Medical College and Research Institute (MGMCRI), Pondicherry, India. Undergraduate medical students of the 4th semester were enrolled in the study to know their perceptions regarding OSPE. The students were briefed regarding OSPE with a PowerPoint presentation and interaction. An examination was conducted using OSPE with10 stations and a total of 74 students participated in the study. The feedback was collected using a preformed proforma consisting of 12 items and analyzed. Most of the participants (82.4%) agreed that OSPE is a better method of examination than the conventional/traditional practical examination. The majority of the participants (77.0%) said that the OSPE covered wide range of knowledge than the conventional practical examination. A large number of students (63.5%) were of the opinion that the OSPE may be exhausting and stressful if number of stations are increased. Overall a larger proportion of the participants preferred OSPE over the conventional practical examination considering the various attributes examined in the study. Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

Effect of surface finish on the failure mechanisms of flip-chip solder joints under electromigration

NASA Astrophysics Data System (ADS)

Lin, Y. L.; Lai, Y. S.; Tsai, C. M.; Kao, C. R.

2006-12-01

Two substrate surface finishes, Au/Ni and organic solderable preservative (OSP), were used to study the effect of the surface finish on the reliability of flip-chip solder joints under electromigration at 150°C ambient temperature. The solder used was eutectic PbSn, and the applied current density was 5×103 A/cm2 at the contact window of the chip. The under bump metallurgy (UBM) on the chip was sputtered Cu/Ni. It was found that the mean-time-to-failure (MTTF) of the OSP joints was six times better than that of the Au/Ni joints (3080 h vs. 500 h). Microstructure examinations uncovered that the combined effect of current crowding and the accompanying local Joule heating accelerated the local Ni UBM consumption near the point of electron entrance. Once Ni was depleted at a certain region, this region became nonconductive, and the flow of the electrons was diverted to the neighboring region. This neighboring region then became the place where electrons entered the joint, and the local Ni UBM consumption was accelerated. This process repeated itself, and the Ni-depleted region extended further on, creating an ever-larger nonconductive region. The solder joint eventually, failed when the nonconductive region became too large, making the effective current density very high. Accordingly, the key factor determining the MTTF was the Ni consumption rate. The joints with the OSP surface finish had a longer MTTF because Cu released from the substrate was able to reduce the Ni consumption rate.

Regulatory protein BBD18 of the lyme disease spirochete: essential role during tick acquisition?

PubMed

Hayes, Beth M; Dulebohn, Daniel P; Sarkar, Amit; Tilly, Kit; Bestor, Aaron; Ambroggio, Xavier; Rosa, Patricia A

2014-04-01

The Lyme disease spirochete Borrelia burgdorferi senses and responds to environmental cues as it transits between the tick vector and vertebrate host. Failure to properly adapt can block transmission of the spirochete and persistence in either vector or host. We previously identified BBD18, a novel plasmid-encoded protein of B. burgdorferi, as a putative repressor of the host-essential factor OspC. In this study, we investigate the in vivo role of BBD18 as a regulatory protein, using an experimental mouse-tick model system that closely resembles the natural infectious cycle of B. burgdorferi. We show that spirochetes that have been engineered to constitutively produce BBD18 can colonize and persist in ticks but do not infect mice when introduced by either tick bite or needle inoculation. Conversely, spirochetes lacking BBD18 can persistently infect mice but are not acquired by feeding ticks. Through site-directed mutagenesis, we have demonstrated that abrogation of spirochete infection in mice by overexpression of BBD18 occurs only with bbd18 alleles that can suppress OspC synthesis. Finally, we demonstrate that BBD18-mediated regulation does not utilize a previously described ospC operator sequence required by B. burgdorferi for persistence in immunocompetent mice. These data lead us to conclude that BBD18 does not represent the putative repressor utilized by B. burgdorferi for the specific downregulation of OspC in the mammalian host. Rather, we suggest that BBD18 exhibits features more consistent with those of a global regulatory protein whose critical role occurs during spirochete acquisition by feeding ticks. IMPORTANCE Lyme disease, caused by Borrelia burgdorferi, is the most common arthropod-borne disease in North America. B. burgdorferi is transmitted to humans and other vertebrate hosts by ticks as they take a blood meal. Transmission between vectors and hosts requires the bacterium to sense changes in the environment and adapt. However, the mechanisms involved in this process are not well understood. By determining how B. burgdorferi cycles between two very different environments, we can potentially establish novel ways to interfere with transmission and limit infection of this vector-borne pathogen. We are studying a regulatory protein called BBD18 that we recently described. We found that too much BBD18 interferes with the spirochete's ability to establish infection in mice, whereas too little BBD18 appears to prevent colonization in ticks. Our study provides new insight into key elements of the infectious cycle of the Lyme disease spirochete.

Toll-like receptor cascade and gene polymorphism in host-pathogen interaction in Lyme disease.

PubMed

Rahman, Shusmita; Shering, Maria; Ogden, Nicholas H; Lindsay, Robbin; Badawi, Alaa

2016-01-01

Lyme disease (LD) risk occurs in North America and Europe where the tick vectors of the causal agent Borrelia burgdorferi sensu lato are found. It is associated with local and systemic manifestations, and has persistent posttreatment health complications in some individuals. The innate immune system likely plays a critical role in both host defense against B. burgdorferi and disease severity. Recognition of B. burgdorferi, activation of the innate immune system, production of proinflammatory cytokines, and modulation of the host adaptive responses are all initiated by Toll-like receptors (TLRs). A number of Borrelia outer-surface proteins (eg, OspA and OspB) are recognized by TLRs. Specifically, TLR1 and TLR2 were identified as the receptors most relevant to LD. Several functional single-nucleotide polymorphisms have been identified in TLR genes, and are associated with varying cytokines types and synthesis levels, altered pathogen recognition, and disruption of the downstream signaling cascade. These single-nucleotide polymorphism-related functional alterations are postulated to be linked to disease development and posttreatment persistent illness. Elucidating the role of TLRs in LD may facilitate a better understanding of disease pathogenesis and can provide an insight into novel therapeutic targets during active disease or postinfection and posttreatment stages.

Toll-like receptor cascade and gene polymorphism in host–pathogen interaction in Lyme disease

PubMed Central

Rahman, Shusmita; Shering, Maria; Ogden, Nicholas H; Lindsay, Robbin; Badawi, Alaa

2016-01-01

Lyme disease (LD) risk occurs in North America and Europe where the tick vectors of the causal agent Borrelia burgdorferi sensu lato are found. It is associated with local and systemic manifestations, and has persistent posttreatment health complications in some individuals. The innate immune system likely plays a critical role in both host defense against B. burgdorferi and disease severity. Recognition of B. burgdorferi, activation of the innate immune system, production of proinflammatory cytokines, and modulation of the host adaptive responses are all initiated by Toll-like receptors (TLRs). A number of Borrelia outer-surface proteins (eg, OspA and OspB) are recognized by TLRs. Specifically, TLR1 and TLR2 were identified as the receptors most relevant to LD. Several functional single-nucleotide polymorphisms have been identified in TLR genes, and are associated with varying cytokines types and synthesis levels, altered pathogen recognition, and disruption of the downstream signaling cascade. These single-nucleotide polymorphism-related functional alterations are postulated to be linked to disease development and posttreatment persistent illness. Elucidating the role of TLRs in LD may facilitate a better understanding of disease pathogenesis and can provide an insight into novel therapeutic targets during active disease or postinfection and posttreatment stages. PMID:27330321

Assessment for learning with Objectively Structured Practical Examination in Biochemistry

PubMed Central

Jaswal, Shivani; Chattwal, Jugesh; Kaur, Jasbinder; Gupta, Seema; Singh, Tejinder

2015-01-01

Context: Despite a radical shift in assessment methodologies over the last decade, the majority of medical colleges still follow the Traditional Practical Examination (TPE). TPE raises concerns about examiner variability, standardization, and uniformity of assessment. To address these issues and in line with the notion of assessments as motivating what and how students learn, Objectively Structured Practical Examination (OSPE) was introduced, as an assessment modality. Despite its usefulness, awareness and motivation to use the same, still needs to be probed. Aims: To implement OSPE in the assessment of practical skills in biochemistry, and to know student and faculty perspectives regarding OSPE. Settings and Design: OSPE was introduced at the stage of formative assessment of practical skills, for 94 year one MBBS students. Subjects and Methods: Students were divided into two groups; the first group was evaluated by the traditional method and the second by OSPE. Students were crossed over on a second examination. The mean score obtained by both the methods was compared statistically. Students and faculty perspectives regarding OSPE were obtained by a questionnaire. Student performance was compared using “Bland–Altman technique,” and Student's t-test. Results: The mean scores of students was found to be significantly higher (P < 0.0001) when assessed with OSPE as compared to TPE. Number of students achieving >70% marks was also significantly higher with OSPE. Validity was supported by a significant correlation coefficient of comparison of marks by the two methods. Feedback from students and faculty indicated that they endorsed OSPE. Conclusions: This evaluation demonstrated the need for a structured approach to assessment. Going in line with the notion that assessment drives learning, introducing OSPE would help tailoring teaching-learning to optimize student satisfaction and learning. PMID:26380217

Model Based Optimal Sensor Network Design for Condition Monitoring in an IGCC Plant

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kumar, Rajeeva; Kumar, Aditya; Dai, Dan

2012-12-31

This report summarizes the achievements and final results of this program. The objective of this program is to develop a general model-based sensor network design methodology and tools to address key issues in the design of an optimal sensor network configuration: the type, location and number of sensors used in a network, for online condition monitoring. In particular, the focus in this work is to develop software tools for optimal sensor placement (OSP) and use these tools to design optimal sensor network configuration for online condition monitoring of gasifier refractory wear and radiant syngas cooler (RSC) fouling. The methodology developedmore » will be applicable to sensing system design for online condition monitoring for broad range of applications. The overall approach consists of (i) defining condition monitoring requirement in terms of OSP and mapping these requirements in mathematical terms for OSP algorithm, (ii) analyzing trade-off of alternate OSP algorithms, down selecting the most relevant ones and developing them for IGCC applications (iii) enhancing the gasifier and RSC models as required by OSP algorithms, (iv) applying the developed OSP algorithm to design the optimal sensor network required for the condition monitoring of an IGCC gasifier refractory and RSC fouling. Two key requirements for OSP for condition monitoring are desired precision for the monitoring variables (e.g. refractory wear) and reliability of the proposed sensor network in the presence of expected sensor failures. The OSP problem is naturally posed within a Kalman filtering approach as an integer programming problem where the key requirements of precision and reliability are imposed as constraints. The optimization is performed over the overall network cost. Based on extensive literature survey two formulations were identified as being relevant to OSP for condition monitoring; one based on LMI formulation and the other being standard INLP formulation. Various algorithms to solve these two formulations were developed and validated. For a given OSP problem the computation efficiency largely depends on the “size” of the problem. Initially a simplified 1-D gasifier model assuming axial and azimuthal symmetry was used to test out various OSP algorithms. Finally these algorithms were used to design the optimal sensor network for condition monitoring of IGCC gasifier refractory wear and RSC fouling. The sensors type and locations obtained as solution to the OSP problem were validated using model based sensing approach. The OSP algorithm has been developed in a modular form and has been packaged as a software tool for OSP design where a designer can explore various OSP design algorithm is a user friendly way. The OSP software tool is implemented in Matlab/Simulink© in-house. The tool also uses few optimization routines that are freely available on World Wide Web. In addition a modular Extended Kalman Filter (EKF) block has also been developed in Matlab/Simulink© which can be utilized for model based sensing of important process variables that are not directly measured through combining the online sensors with model based estimation once the hardware sensor and their locations has been finalized. The OSP algorithm details and the results of applying these algorithms to obtain optimal sensor location for condition monitoring of gasifier refractory wear and RSC fouling profile are summarized in this final report.« less

Patatin-related phospholipase A, pPLAIIIα, modulates the longitudinal growth of vegetative tissues and seeds in rice

PubMed Central

Liu, Guangmeng; Zhang, Ke; Ai, Jun; Deng, Xianjun; Hong, Yueyun; wang, Xuemin

2015-01-01

Patatin-related phospholipase A (pPLA) hydrolyses glycerolipids to produce fatty acids and lysoglycerolipids. The Oryza sativa genome has 21 putative pPLAs that are grouped into five subfamilies. Overexpression of OspPLAIIIα resulted in a dwarf phenotype with decreased length of rice stems, roots, leaves, seeds, panicles, and seeds, whereas OspPLAIIIα-knockout plants had longer panicles and seeds. OspPLAIIIα-overexpressing plants were less sensitive than wild-type and knockout plants to gibberellin-promoted seedling elongation. OspPLAIIIα overexpression and knockout had an opposite effect on the expression of the growth repressor SLENDER1 in the gibberellin signalling process. OspPLAIIIα-overexpressing plants had decreased mechanical strength and cellulose content, but exhibited increases in the expression of several cellulose synthase genes. These results indicate that OspPLAIIIα plays a role in rice vegetative and reproductive growth and that the constitutive, high activity of OspPLAIIIα suppresses cell elongation. The decreased gibberellin response in overexpressing plants is probably a result of the decreased ability to make cellulose for anisotropic cell expansion. PMID:26290597

Measurement and modeling of surface temperature dynamics of the NSTX liquid lithium divertor

NASA Astrophysics Data System (ADS)

McLean, A. G.; Gan, K. F.; Ahn, J.-W.; Gray, T. K.; Maingi, R.; Abrams, T.; Jaworski, M. A.; Kaita, R.; Kugel, H. W.; Nygren, R. E.; Skinner, C. H.; Soukhanovskii, V. A.

2013-07-01

Dual-band infrared (IR) measurements of the National Spherical Torus eXperiment (NSTX) Liquid Lithium Divertor (LLD) are reported that demonstrate liquid Li is more effective at removing plasma heat flux than Li-conditioned graphite. Extended dwell of the outer strike point (OSP) on the LLD caused an incrementally larger area to be heated above the Li melting point through the discharge leading to enhanced D retention and plasma confinement. Measurement of Tsurface near the OSP demonstrates a significant reduction of the LLD surface temperature compared to that of Li-coated graphite at the same major radius. Modeling of these data with a 2-D simulation of the LLD structure in the DFLUX code suggests that the structure of the LLD was successful at handling up to q⊥,peak = 5 MW/m2 inter-ELM and up to 10 MW/m2 during ELMs from its plasma-facing surface as intended, and provide an innovative method for inferring the Li layer thickness.

Immunization with a recombinant subunit OspA vaccine markedly impacts the rate of newly acquired Borrelia burgdorferi infections in client-owned dogs living in a coastal community in Maine, USA.

PubMed

Eschner, Andrew K; Mugnai, Kristen

2015-02-10

In North America, Borrelia burgdorferi is the causative bacterial agent of canine Lyme borreliosis and is transmitted following prolonged attachment and feeding of vector ticks, Ixodes scapularis or Ixodes pacificus. Its prevention is predicated upon tick-avoidance, effective on-animal tick control and effective immunization strategies. The purpose of this study is to characterize dogs that are newly seropositive for Borrelia burgdorferi infection in relation to compliant use of a recombinant OspA canine Lyme borreliosis vaccine. Specifically, Preventive Fractions (PF) and Risk Ratios (RR) associated with the degree of vaccine compliancy (complete versus incomplete) are determined. 6,202 dogs were tested over a five year period in a single veterinary hospital utilizing a non-adjuvanted, recombinant OspA vaccine according to a 0, 1, 6 month (then yearly) protocol. Rates of newly acquired "Lyme-positive" antibody test results were compared between protocol compliant and poorly compliant (incompletely and/or non-vaccinated) dogs. Over the five-year span, one percent (range 0.39 - 1.3) of protocol compliant vaccinated, previously antibody negative dogs became seropositive for infection. Approximately twenty-one percent (range 16.8 - 33.3) of incompletely vaccinated dogs became positive for infection-specific antibodies. The Preventative Fraction for testing positive for antibodies specific for infection with Borrelia burgdorferi in any given year based on optimal vaccine compliance was, on average, 95.3% (range 93.29 - 98.08). The Risk Ratio for becoming infected with Borrelia burgdorferi antibodies in any given year if vaccine non-compliant was 21.41 (range 14.9 - 52.1). There was a high statistically significant relationship (p = <0.0001) in the observed data in terms of vaccination protocol compliance and the probability of Borrelia burgdorferi infection in each of the five years under study. The recombinant outer surface protein A (rOspA) vaccine for dogs is highly effective in preventing new seropositive cases of Borrelia burgdoferi infection over a five-year period in dogs living in an endemic area. Dogs that were vaccine protocol-compliant were significantly less likely to become infected (as indirectly assessed by antibody) with the agent of canine Lyme borreliosis as measured by Preventive Fraction and Risk Ratio calculations.

Detection of Borrelia burgdorferi sensu stricto ospC alleles associated with human lyme borreliosis worldwide in non-human-biting tick Ixodes affinis and rodent hosts in Southeastern United States.

PubMed

Rudenko, Nataliia; Golovchenko, Maryna; Hönig, Václav; Mallátová, Nadja; Krbková, Lenka; Mikulásek, Peter; Fedorova, Natalia; Belfiore, Natalia M; Grubhoffer, Libor; Lane, Robert S; Oliver, James H

2013-03-01

Comparative analysis of ospC genes from 127 Borrelia burgdorferi sensu stricto strains collected in European and North American regions where Lyme disease is endemic and where it is not endemic revealed a close relatedness of geographically distinct populations. ospC alleles A, B, and L were detected on both continents in vectors and hosts, including humans. Six ospC alleles, A, B, L, Q, R, and V, were prevalent in Europe; 4 of them were detected in samples of human origin. Ten ospC alleles, A, B, D, E3, F, G, H, H3, I3, and M, were identified in the far-western United States. Four ospC alleles, B, G, H, and L, were abundant in the southeastern United States. Here we present the first expanded analysis of ospC alleles of B. burgdorferi strains from the southeastern United States with respect to their relatedness to strains from other North American and European localities. We demonstrate that ospC genotypes commonly associated with human Lyme disease in European and North American regions where the disease is endemic were detected in B. burgdorferi strains isolated from the non-human-biting tick Ixodes affinis and rodent hosts in the southeastern United States. We discovered that some ospC alleles previously known only from Europe are widely distributed in the southeastern United States, a finding that confirms the hypothesis of transoceanic migration of Borrelia species.

3D printed porous polycaprolactone/oyster shell powder (PCL/OSP) scaffolds for bone tissue engineering

NASA Astrophysics Data System (ADS)

Luo, Wenfeng; Zhang, Shuangying; Lan, Yuewei; Huang, Chen; Wang, Chao; Lai, Xuexu; Chen, Hanwei; Ao, Ningjian

2018-04-01

In this work, oyster shell powder (OSP) was used as the bio-filler and combined with polycaprolactone (PCL) through melt blending methodology. The PCL and PCL/OSP scaffolds were prepared using additive manufacturing process. All the 3D printed scaffolds hold a highly porosity and interconnected pore structures. OSP particles are dispersed in the polymer matrix, which helped to improve the degree of crystallinity and mineralization ability of the scaffolds. There was no significant cytotoxicity of the prepared scaffolds towards MG-63 cells, and all the scaffolds showed a well ALP activity. Therefore, PCL/OSP scaffolds had a high potential to be employed in the bone tissue engineering.

Gauge supergravity in D = 2 + 2

NASA Astrophysics Data System (ADS)

Castellani, Leonardo

2017-10-01

We present an action for chiral N = (1 , 0) supergravity in 2 + 2 dimensions. The fields of the theory are organized into an OSp(1|4) connection supermatrix, and are given by the usual vierbein V a , spin connection ω ab , and Majorana gravitino ψ. In analogy with a construction used for D = 10 + 2 gauge supergravity, the action is given by ∫STr( R 2 Γ), where R is the OSp(1|4) curvature supermatrix two-form, and Γ a constant supermatrix containing γ 5. It is similar, but not identical to the MacDowell-Mansouri action for D = 2 + 2 supergravity. The constant supermatrix breaks OSp(1|4) gauge invariance to a subalgebra OSp(1|2) ⊕ Sp(2), including a Majorana-Weyl supercharge. Thus half of the OSp(1|4) gauge supersymmetry survives. The gauge fields are the selfdual part of ω ab and the Weyl projection of ψ for OSp(1|2), and the antiselfdual part of ω ab for Sp(2). Supersymmetry transformations, being part of a gauge superalgebra, close off-shell. The selfduality condition on the spin connection can be consistently imposed, and the resulting "projected" action is OSp(1|2) gauge invariant.

NASA's Orbital Space Plane Risk Reduction Strategy

NASA Technical Reports Server (NTRS)

Dumbacher, Dan

2003-01-01

This paper documents the transformation of NASA s Space Launch Initiative (SLI) Second Generation Reusable Launch Vehicle Program under the revised Integrated Space Transportation Plan, announced November 2002. Outlining the technology development approach followed by the original SLI, this paper gives insight into the current risk-reduction strategy that will enable confident development of the Nation s first orbital space plane (OSP). The OSP will perform an astronaut and contingency cargo transportation function, with an early crew rescue capability, thus enabling increased crew size and enhanced science operations aboard the International Space Station. The OSP design chosen for full-scale development will take advantage of the latest innovations American industry has to offer. The OSP Program identifies critical technologies that must be advanced to field a safe, reliable, affordable space transportation system for U.S. access to the Station and low-Earth orbit. OSP flight demonstrators will test crew safety features, validate autonomous operations, and mature thermal protection systems. Additional enabling technologies may be identified during the OSP design process as part of an overall risk-management strategy. The OSP Program uses a comprehensive and evolutionary systems acquisition approach, while applying appropriate lessons learned.

Patatin-related phospholipase A, pPLAIIIα, modulates the longitudinal growth of vegetative tissues and seeds in rice.

PubMed

Liu, Guangmeng; Zhang, Ke; Ai, Jun; Deng, Xianjun; Hong, Yueyun; Wang, Xuemin

2015-11-01

Patatin-related phospholipase A (pPLA) hydrolyses glycerolipids to produce fatty acids and lysoglycerolipids. The Oryza sativa genome has 21 putative pPLAs that are grouped into five subfamilies. Overexpression of OspPLAIIIα resulted in a dwarf phenotype with decreased length of rice stems, roots, leaves, seeds, panicles, and seeds, whereas OspPLAIIIα-knockout plants had longer panicles and seeds. OspPLAIIIα-overexpressing plants were less sensitive than wild-type and knockout plants to gibberellin-promoted seedling elongation. OspPLAIIIα overexpression and knockout had an opposite effect on the expression of the growth repressor SLENDER1 in the gibberellin signalling process. OspPLAIIIα-overexpressing plants had decreased mechanical strength and cellulose content, but exhibited increases in the expression of several cellulose synthase genes. These results indicate that OspPLAIIIα plays a role in rice vegetative and reproductive growth and that the constitutive, high activity of OspPLAIIIα suppresses cell elongation. The decreased gibberellin response in overexpressing plants is probably a result of the decreased ability to make cellulose for anisotropic cell expansion. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology.

The Debate on the Moral Responsibilities of Online Service Providers.

PubMed

Taddeo, Mariarosaria; Floridi, Luciano

2016-12-01

Online service providers (OSPs)-such as AOL, Facebook, Google, Microsoft, and Twitter-significantly shape the informational environment (infosphere) and influence users' experiences and interactions within it. There is a general agreement on the centrality of OSPs in information societies, but little consensus about what principles should shape their moral responsibilities and practices. In this article, we analyse the main contributions to the debate on the moral responsibilities of OSPs. By endorsing the method of the levels of abstract (LoAs), we first analyse the moral responsibilities of OSPs in the web (LoA IN ). These concern the management of online information, which includes information filtering, Internet censorship, the circulation of harmful content, and the implementation and fostering of human rights (including privacy). We then consider the moral responsibilities ascribed to OSPs on the web (LoA ON ) and focus on the existing legal regulation of access to users' data. The overall analysis provides an overview of the current state of the debate and highlights two main results. First, topics related to OSPs' public role-especially their gatekeeping function, their corporate social responsibilities, and their role in implementing and fostering human rights-have acquired increasing relevance in the specialised literature. Second, there is a lack of an ethical framework that can (a) define OSPs' responsibilities, and (b) provide the fundamental sharable principles necessary to guide OSPs' conduct within the multicultural and international context in which they operate. This article contributes to the ethical framework necessary to deal with (a) and (b) by endorsing a LoA enabling the definition of the responsibilities of OSPs with respect to the well-being of the infosphere and of the entities inhabiting it (LoA For ).

Effect of Electromigration on the Type of Drop Failure of Sn-3.0Ag-0.5Cu Solder Joints in PBGA Packages

NASA Astrophysics Data System (ADS)

Huang, M. L.; Zhao, N.

2015-10-01

Board-level drop tests of plastic ball grid array (PBGA) packages were performed in accordance with the Joint Electron Devices Engineering Council standard to investigate the effect of electromigration (EM) on the drop reliability of Sn-3.0Ag-0.5Cu solder joints with two substrate surface finishes, organic solderability preservative (OSP) and electroless nickel electroless palladium immersion gold (ENEPIG). In the as-soldered state, drop failures occurred at the substrate sides only, with cracks propagating within the interfacial intermetallic compound (IMC) layer for OSP solder joints and along the IMC/Ni-P interface for ENEPIG solder joints. The drop lifetime of OSP solder joints was approximately twice that of ENEPIG joints. EM had an important effect on crack formation and drop lifetime of the PBGA solder joints. ENEPIG solder joints performed better in drop reliability tests after EM, that is, the drop lifetime of ENEPIG joints decreased by 43% whereas that of OSP solder joints decreased by 91%, compared with the as-soldered cases. The more serious polarity effect, i.e., excessive growth of the interfacial IMC at the anode, was responsible for the sharper decrease in drop lifetime. The different types of drop failure of PBGA solder joints before and after EM, including the position of initiation and the propagation path of cracks, are discussed on the basis of the growth behavior of interfacial IMC.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sloan, J.M.; Pergantis, C.G.

Organic and organo-metallic coatings are presently being applied over bare copper as an approach to improve the co-planarity of circuit boards. Conformal organic solderability preservative coatings (OSP) are environmentally and economically advantageous over the more commonly used lead based coatings. Problems arise in assessing the solderability of the bare copper and the integrity of the organic coating. Specular reflectance Fourier transform infrared spectroscopy (FT-IR) was utilized to monitor and evaluate the formation of Cu oxides occurring on copper substrates used in the manufacturing of electronic circuit boards. Previous studies reported the utility of this technique. By measuring the oxide andmore » protective coating characteristics of these surfaces, their solderability performance can rapidly be evaluated in a manufacturing environment. OSP coated test specimens were subjected to hot-dry and hot-wet environmental conditions using MIL-STD-202F and MIL-STD-883E as guides. The resultant FT-IR spectra provided clear evidence for the formation of various Cu oxides at the Cu/OSP interface over exposure time, for the samples subjected to the hot-dry environment. IR spectral bands consistent with O-Cu-O and Cu{sub 2}O{sub 2} formation appear, while very minimal deterioration to the OSP coating was observed. The appearance of the Cu oxide layers grew steadily with increased environmental exposure. Specimens subjected to the hot-wet conditions showed no significant signs of deterioration. The IR data can be directly correlated to solderability performance as evaluated by wet balance testing.« less

Objective structured practical examination in biochemistry: An experience in Medical College, Kolkata.

PubMed

Kundu, Dipankar; Das, H N; Sen, Gargi; Osta, Manish; Mandal, T; Gautam, Divyendu

2013-01-01

Undergraduate medical examination is undergoing extensive re evaluation with new core educational objectives being defined. Consequently, new exam systems have also been designed to test the objectives. Objective structured practical examination (OSPE) is one of them. To introduce OSPE as a method of assessment of practical skills and learning and to determine student satisfaction regarding the OSPE. Furthermore, to explore the faculty perception of OSPE as a learning and assessment tool. The first M.B.B.S students of 2011 12 batch of Medical College, Kolkata, were the subjects for the study. OSPE was organized and conducted on "Identification of Unknown Abnormal Constituents in Urine." Coefficient of reliability of questions administered was done by calculating Cronbach's alpha. A questionnaire on various components of the OSPE was administered to get the feedback. 16 students failed to achieve an average of 50% or above in the assessment. However, 49 students on an average achieved >75%, 52 students achieved between 65% and 75%, and 29 students scored between 50% and 65%. Cronbach's alpha of the questions administered showed to be having high internal consistency with a score of 0.80. Ninety nine percent of students believed that OSPE helps them to improve and 81% felt that this type of assessment fits in as both learning and evaluation tools. Faculty feedback reflected that such assessment tested objectivity, measured practical skills better, and eliminated examiner bias to a greater extent. OSPE tests different desired components of competence better and eliminated examiner bias. Student feedback reflects that such assessment helps them to improve as it is effective both as teaching and evaluation tools.

Molecular characterization, genomic arrangement, and expression of bmpD, a new member of the bmp class of genes encoding membrane proteins of Borrelia burgdorferi.

PubMed Central

Ramamoorthy, R; Povinelli, L; Philipp M, T

1996-01-01

An expression library made with Borrelia burgdorferi DNA in the vector lambda ZapII was screened with serum from a monkey infected with the Lyme disease agent. This serum killed B. burgdorferi in vitro by an antibody-dependent, complement-mediated mechanism and contained antibodies to at least seven spirochetal antigens, none of which were the major outer surface proteins OspA or OspB. Among several positive clones, a clone containing the B. burgdorferi bmpA gene encoding the immunodominant antigen P39 was obtained. Chromosome walking and DNA sequence analysis permitted the identification of two additional upstream genes homologous to the bmpA gene and its related companion, bmpB. The first of these was the recently characterized bmpC gene, and adjacent to it was the fourth and new member of this class, which has been designated bmpD. The gene product encoded by bmpD is 34l residues long, contains a signal sequence with a potential signal peptidase II cleavage site, and has 26% identity with TmpC of Treponema pallidum. Southern blotting confirmed the tandem arrangement of all four bmp genes in the chromosome of B. burgdorferi JD1. However, Northern (RNA) blotting revealed that bmpD is expressed as a monocistronic transcript, which indicates that it is not part of an operon at the bmp locus. The bmpD gene was found to be conserved in representative members of the three species of the B. burgdorferi sensu lato complex, suggesting that it serves an important biological function in the spirochete. PMID:8606088

Synthesis, folding, structure and activity of a predicted peptide from the sea anemone Oulactis sp. with an ShKT fold.

PubMed

Krishnarjuna, Bankala; Villegas-Moreno, Jessica; Mitchell, Michela L; Csoti, Agota; Peigneur, Steve; Amero, Carlos; Pennington, Michael W; Tytgat, Jan; Panyi, Gyorgy; Norton, Raymond S

2018-05-19

Sea anemone venom is rich in bioactive compounds, including peptides containing multiple disulfide bridges. In a transcriptomic study on Oulactis sp., we identified the putative 36-residue peptide, OspTx2b, which is an isoform of the K V channel blocker OspTx2a (Sunanda P et al. [2018] Identification, chemical synthesis, structure and function of a new K V 1 channel blocking peptide from Oulactis sp. Peptide Science, in press). As OspTx2b contains a ShK/BgK-like cysteine framework, with high amino acid sequence similarity to BgK, we were interested to investigate its structure and function. The solution structure of OspTx2b was determined using nuclear magnetic resonance spectroscopy. OspTx2b does indeed possess a BgK-like scaffold, with the same disulfide bond connectivities. The orientation of the Lys-Tyr dyad in OspTx2b is more similar to that in ShK than in BgK. However, it failed to show against a range of voltage-gated potassium channels in Xenopus oocytes and human T lymphocytes. OspTx2b also showed no growth inhibitory activity against several strains of bacteria and fungi. Having a BgK-like fold with the Lys-Tyr dyad but no BgK-like activity highlights the importance of key amino acid residues in BgK that are missing in OspTx2b. The lack of activity against the K V channels assessed in this study emphasises that the ShK/BgK scaffold is capable of supporting functional activity beyond potassium channel blockade. Copyright © 2018. Published by Elsevier Ltd.

Immunoblot analysis of immunoglobulin G response to the Lyme disease agent (Borrelia burgdorferi) in experimentally and naturally exposed dogs.

PubMed Central

Greene, R T; Walker, R L; Nicholson, W L; Heidner, H W; Levine, J F; Burgess, E C; Wyand, M; Breitschwerdt, E B; Berkhoff, H A

1988-01-01

Immunoblots were used to study the immunoglobulin G response to Borrelia burgdorferi in experimentally and naturally exposed dogs. Adsorption studies confirmed that the antibodies were specific for B. burgdorferi. Experimentally exposed dogs were asymptomatic. Naturally exposed dogs included both asymptomatic animals and animals showing signs compatible with Lyme disease. Naturally exposed dogs were from four geographic regions of the country. No differences were detected between immunoblot patterns of naturally exposed symptomatic or asymptomatic dogs from different areas of the country. The immunoblot patterns obtained with sera from experimentally exposed dogs were different from those obtained with sera from naturally exposed dogs and were characterized by reactivity to fewer and different protein bands. Immunoblot analysis using an OspA-protein-producing Escherichia coli recombinant showed that experimentally exposed dogs produced antibodies to OspA, whereas naturally exposed dogs did not. Modifications of the immune response over time, different routes of antigen presentation, and strain variation are factors postulated to account for the observed differences. Images PMID:3366860

Objective structured practical examination (OSPE) in Forensic Medicine: students' point of view.

PubMed

Menezes, Ritesh G; Nayak, Vinod C; Binu, V S; Kanchan, Tanuj; Rao, P P Jagadish; Baral, Prakash; Lobo, Stany W

2011-11-01

The purpose of this study was to assess the attitudes of undergraduate medical students towards the objective structured practical examination (OSPE) in Forensic Medicine, in a medical college in Nepal. Participants included 59 undergraduate medical students of the 7th semester. Findings indicated that the OSPE was an acceptable tool considering the conduct of practical examination in Forensic Medicine at the undergraduate level. The overall mean attitude score was towards the favourable side. Students strongly agreed that the OSPE tested a wide range of skills. They also strongly agreed that it was a good form of examination as well as a learning experience. The introduction of the OSPE replacing the conventional method of practical examination in Forensic Medicine is a step in the right direction taken to objectively assess undergraduate medical students. Copyright © 2011 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

Application of Probabilistic Risk Assessment (PRA) During Conceptual Design for the NASA Orbital Space Plane (OSP)

NASA Technical Reports Server (NTRS)

Rogers, James H.; Safie, Fayssal M.; Stott, James E.; Lo, Yunnhon

2004-01-01

In order to meet the space transportation needs for a new century, America's National Aeronautics and Space Administration (NASA) has implemented an Integrated Space Transportation Plan to produce safe, economical, and reliable access to space. One near term objective of this initiative is the design and development of a next-generation vehicle and launch system that will transport crew and cargo to and from the International Space Station (ISS), the Orbital Space Plane (OSP). The OSP system is composed of a manned launch vehicle by an existing Evolved Expendable Launch Vehicle (EELV). The OSP will provide emergency crew rescue from the ISS by 2008, and provide crew and limited cargo transfer to and from the ISS by 2012. A key requirement is for the OSP to be safer and more reliable than the Soyuz and Space Shuttle, which currently provide these capabilities.

Persistently declining oxygen levels in the interior waters of the eastern subarctic Pacific

NASA Astrophysics Data System (ADS)

Whitney, Frank A.; Freeland, Howard J.; Robert, Marie

2007-10-01

Fifty years of measurements at Ocean Station Papa (OSP, 50°N, 145°W) show trends in the interior waters of the subarctic Pacific that are both impacted by short term (few years to bi-decadal) atmospheric or ocean circulation oscillations and by persistent climate trends. Between 1956 and 2006, waters below the ocean mixed layer to a depth of at least 1000 m have been warming and losing oxygen. On density surfaces found in the depth range 100-400 m ( σθ = 26.3-27.0), the ocean is warming at 0.005-0.012 °C y -1, whereas oxygen is declining at 0.39-0.70 μmol kg -1 y -1 or at an integrated rate of 123 mmol m -2 y -1 (decrease of 22% over 50 years). During this time, the hypoxic boundary (defined as 60 μmol O 2 kg -1) has shoaled from ∼400 to 300 m. In the Alaska Gyre, the 26.2 isopycnal occasionally ventilates, whereas at OSP 26.0 σθ has not been seen at the ocean surface since 1971 as the upper ocean continues to stratify. To interpret the 50 year record at OSP, the isopycnal transport of oxygenated waters within the interior of the subarctic Pacific is assessed by using a slightly modified “NO” parameter [Broecker, W., 1974. “NO” a conservative water-mass tracer. Earth and Planetary Science Letters 23, 100-107]. The highest nitrate-oxygen signature in interior waters of the North Pacific is found in the Bering Sea Gyre, Western Subarctic Gyre and East Kamchatka Current region as a consequence of winter mixing to the ∼26.6 isopycnal. By mixing with low NO waters found in the subtropics and Okhotsk Sea, this signature is diluted as waters flow eastward across the Pacific. Evidence of low NO waters flowing north from California is seen along the coasts of British Columbia and SE Alaska. Oxygen in the subsurface waters of the Alaskan Gyre was supplied ∼60% by subarctic and 40% by subtropical waters during WOCE surveys, whereas such estimates are shown to periodically vary by 20% at OSP. Other features discernable in the OSP data include periods of increased ventilation of deeper isopycnals on an ∼18 year cycle and strong, short term (few month) variability caused by passing mesoscale eddies. The potential impacts of declining oxygen on coastal ecosystems are discussed.

Study of Electromigration-Induced Failures on Cu Pillar Bumps Joined to OSP and ENEPIG Substrates

NASA Astrophysics Data System (ADS)

Hsiao, Yu-Hsiang; Lin, Kwang-Lung; Lee, Chiu-Wen; Shao, Yu-Hsiu; Lai, Yi-Shao

2012-12-01

This work studies electromigration (EM)-induced failures on Cu pillar bumps joined to organic solderability preservative (OSP) on Cu substrates (OSP-bumps) and electroless Ni(P)/electroless Pd/immersion Au (ENEPIG) under bump metallurgy (UBM) on Cu substrates (ENEPIG-bumps). Two failure modes (Cu pad consumption and gap formation) were found with OSP-bumps, but only one failure mode (gap formation) was found with ENEPIG-bumps. The main interfacial compound layer was the Cu6Sn5 compound, which suffered significant EM-induced dissolution, eventually resulting in severe Cu pad consumption at the cathode side for OSP-bumps. A (Cu,Ni)6Sn5 layer with strong resistance to EM-induced dissolution exists at the joint interface when a nickel barrier layer is incorporated at the cathode side (Ni or ENEPIG), and these imbalanced atomic fluxes result in the voids and gap formation. OSP-bumps showed better lifetime results than ENEPIG-bumps for several current stressing conditions. The inverse Cu atomic flux ( J Cu,chem) which diffuses from the Cu pad to cathode side retards the formation of voids. The driving force for J Cu,chem comes from the difference in chemical potential between the (Cu,Ni)6Sn5 and Cu6Sn5 phases.

Preparation and Properties of Nanocellulose from Organosolv Straw Pulp

NASA Astrophysics Data System (ADS)

Barbash, V. A.; Yaschenko, O. V.; Shniruk, O. M.

2017-03-01

The object of this work is to present a study of nanocellulose preparation from organosolv straw pulp (OSP) and its properties. OSP was obtained through thermal treatment in the system of isobutyl alcohol-H2O-KOH-hydrazine followed by processing in the mixture of acetic acid and hydrogen peroxide for bleaching and removal of residual non-cellulosic components. We have obtained nanocellulose from OSP through acid hydrolysis with lower consumption of sulfuric acid and followed by ultrasound treatment. The structural change and crystallinity degree of OSP and nanocellulose were studied by means of SEM and XRD techniques. It has been established that nanocellulose has a density up to 1.3 g/cm3, transparency up to 70%, crystallinity degree 72.5%. The TEM and AFM methods shown that nanocellulose have diameter of particles in the range from 10 to 40 nm. Thermogravimetric analysis confirmed that nanocellulose films have more dense structure and smaller mass loss in the temperature range 220-260 °C compared with OSP. The obtained nanocellulose films had high Young's modulus up to 11.45 GPa and tensile strength up to 42.3 MPa. The properties of obtained nanocellulose from OSP exhibit great potential in its application for the preparation of new nanocomposite materials.

Population structure of the lyme borreliosis spirochete Borrelia burgdorferi in the western black-legged tick (Ixodes pacificus) in Northern California.

PubMed

Girard, Yvette A; Travinsky, Bridgit; Schotthoefer, Anna; Fedorova, Natalia; Eisen, Rebecca J; Eisen, Lars; Barbour, Alan G; Lane, Robert S

2009-11-01

Factors potentially contributing to the lower incidence of Lyme borreliosis (LB) in the far-western than in the northeastern United States include tick host-seeking behavior resulting in fewer human tick encounters, lower densities of Borrelia burgdorferi-infected vector ticks in peridomestic environments, and genetic variation among B. burgdorferi spirochetes to which humans are exposed. We determined the population structure of B. burgdorferi in over 200 infected nymphs of the primary bridging vector to humans, Ixodes pacificus, collected in Mendocino County, CA. This was accomplished by sequence typing the spirochete lipoprotein ospC and the 16S-23S rRNA intergenic spacer (IGS). Thirteen ospC alleles belonging to 12 genotypes were found in California, and the two most abundant, ospC genotypes H3 and E3, have not been detected in ticks in the Northeast. The most prevalent ospC and IGS biallelic profile in the population, found in about 22% of ticks, was a new B. burgdorferi strain defined by ospC genotype H3. Eight of the most common ospC genotypes in the northeastern United States, including genotypes I and K that are associated with disseminated human infections, were absent in Mendocino County nymphs. ospC H3 was associated with hardwood-dominated habitats where western gray squirrels, the reservoir host, are commonly infected with LB spirochetes. The differences in B. burgdorferi population structure in California ticks compared to the Northeast emphasize the need for a greater understanding of the genetic diversity of spirochetes infecting California LB patients.

Architecture and Chemical Coding of the Inner and Outer Submucous Plexus in the Colon of Piglets

PubMed Central

Petto, Carola; Gäbel, Gotthold; Pfannkuche, Helga

2015-01-01

In the porcine colon, the submucous plexus is divided into an inner submucous plexus (ISP) on the epithelial side and an outer submucous plexus (OSP) on the circular muscle side. Although both plexuses are probably involved in the regulation of epithelial functions, they might differ in function and neurochemical coding according to their localization. Therefore, we examined expression and co-localization of different neurotransmitters and neuronal markers in both plexuses as well as in neuronal fibres. Immunohistochemical staining was performed on wholemount preparations of ISP and OSP and on cryostat sections. Antibodies against choline acetyltransferase (ChAT), substance P (SP), somatostatin (SOM), neuropeptide Y (NPY), vasoactive intestinal peptide (VIP), neuronal nitric oxide synthase (nNOS) and the pan-neuronal markers Hu C/D and neuron specific enolase (NSE) were used. The ISP contained 1,380 ± 131 ganglia per cm2 and 122 ± 12 neurons per ganglion. In contrast, the OSP showed a wider meshwork (215 ± 33 ganglia per cm2) and smaller ganglia (57 ± 3 neurons per ganglion). In the ISP, 42% of all neurons expressed ChAT. About 66% of ChAT-positive neurons co-localized SP. A small number of ISP neurons expressed SOM. Chemical coding in the OSP was more complex. Besides the ChAT/±SP subpopulation (32% of all neurons), a nNOS-immunoreactive population (31%) was detected. Most nitrergic neurons were only immunoreactive for nNOS; 10% co-localized with VIP. A small subpopulation of OSP neurons was immunoreactive for ChAT/nNOS/±VIP. All types of neurotransmitters found in the ISP or OSP were also detected in neuronal fibres within the mucosa. We suppose that the cholinergic population in the ISP is involved in the control of epithelial functions. Regarding neurochemical coding, the OSP shares some similarities with the myenteric plexus. Because of its location and neurochemical characteristics, the OSP may be involved in controlling both the mucosa and circular muscle. PMID:26230272

Net community production and calcification from 7 years of NOAA Station Papa Mooring measurements

NASA Astrophysics Data System (ADS)

Fassbender, Andrea J.; Sabine, Christopher L.; Cronin, Meghan F.

2016-02-01

Seven years of near-continuous observations from the Ocean Station Papa (OSP) surface mooring were used to evaluate drivers of marine carbon cycling in the eastern subarctic Pacific. Processes contributing to mixed layer carbon inventory changes throughout each deployment year were quantitatively assessed using a time-dependent mass balance approach in which total alkalinity and dissolved inorganic carbon were used as tracers. By using two mixed layer carbon tracers, it was possible to isolate the influences of net community production (NCP) and calcification. Our results indicate that the annual NCP at OSP is 2 ± 1 mol C m-2 yr-1 and the annual calcification is 0.3 ± 0.3 mol C m-2 yr-1. Piecing together evidence for potentially significant dissolved organic carbon cycling in this region, we estimate a particulate inorganic carbon to particulate organic carbon ratio between 0.15 and 0.25. This is at least double the global average, adding to the growing evidence that calcifying organisms play an important role in carbon export at this location. These results, coupled with significant seasonality in the NCP, suggest that carbon cycling near OSP may be more complex than previously thought and highlight the importance of continuous observations for robust assessments of biogeochemical cycling.

Climatology of sediment flux and composition in the subarctic Northeast Pacific Ocean with biogeochemical implications

NASA Astrophysics Data System (ADS)

Timothy, D. A.; Wong, C. S.; Barwell-Clarke, J. E.; Page, J. S.; White, L. A.; Macdonald, R. W.

2013-09-01

Sequentially sampling conical sediment traps were maintained at Ocean Station Papa (OSP; 50°N, 145°W) in the Alaska Gyre from September 1982 to June 2006. The time series began with a single trap at 3800 m and traps were added at 1000 m and 200 m in March 1983 and May 1989, respectively. A trap at 3500-3700 m also was moored 5° north of OSP from May 1990 to August 1992. Total mass, biogenic silica (BSi), calcium carbonate (CaCO3), particulate organic carbon (POC) and particulate nitrogen (PN) were routinely measured. In this paper, we develop climatologies of sediment flux and composition at OSP, describing the characteristic features for comparison to sedimentary conditions globally. We then expand our use of the climatologies to arrive at four main conclusions regarding ecology and geochemistry at OSP. Fluxes of BSi and CaCO3 at 200 m and 1000 m lag by one month the annual cycle of irradiance and arrive at 3800 m ∼16 d later, with maximum export occurring several months later for POC. Next, the annual cycle of BSi flux shows that diatom production in late winter and spring is higher than indicated by spring decline of surface nutrients. We then show that the annual cycle of POC flux implies a net community production of organic carbon (NCPOC) of 3.6-5.3 mol m-2 y-1, double estimates based on mixed layer tracers but similar to estimates unaffected by mixing. NCPOC, combined with a CaCO3:POC production ratio of 0.18 determined from trap fluxes, gives a net community production of CaCO3 (NCPIC) of 0.65-0.95 mol m-2 y-1, in agreement with CaCO3 dissolution in the water column plus abyssal CaCO3 flux. Lastly, the flux climatologies at 1000 m and 3800 m are used to infer particle transformations in the bathypelagic zone including disaggregation and remineralization. Fluxes at 3800 m are best described as the sum of a primary flux sinking rapidly and a slowly-sinking secondary flux. Disaggregation of the primary flux is the likely source of secondary fluxes, with a lithogenic component transported horizontally also reaching the 3800-m traps. A detailed description of the sampling also is provided so future experiments can benefit from the successes and failures encountered at OSP. Fluxes normalized to 2000 m are 2.7, 1.3 and 1.1 times higher than the global averages for BSi, CaCO3 and POC, respectively. The Alaska Gyre is thus a siliceous basin with unusually high calcareous fluxes. Lithogenic fluxes are minor at OSP, making this site ideal to detect dust-fall events. Fluxes of BSi and CaCO3 lag surface solar irradiance by about one month at 200 m and 1000 m, and by another ∼two weeks at 3800 m. POM is preferentially retained and recycled in the mixed layer, with maximum export occurring several months after maximum fluxes of BSi and CaCO3. Export fluxes are episodic at OSP despite perennially low chlorophyll concentrations showing little seasonality. As a result of episodically high fluxes, 40-50% of MARK7 traps with narrow sampling bottles became clogged during deployment at 3800 m. Given the common occurrence of this problem globally, traps with larger bottom orifices should become standard protocol. Sediment traps provide an excellent opportunity to test sedimentary tracers of past ocean conditions. In this regard, POC content at OSP is a poor indicator of mass or POC flux because POC is diluted by BSi and CaCO3 when mass flux is high. The annual cycle of BSi flux and a reanalysis of surface nutrient data show the spring delay in [Si(OH)4] decline, based on mixed-layer nutrient climatology, results from intense mixing in spring rather than delayed diatom growth as previously proposed. The annual cycle of POC flux, normalized to measures of net community production of organic carbon (NCPOC) in summer-fall (Emerson and Stump, 2010), implies an annual NCPOC of 3.6-5.3 mol m-2 y-1. This rate is similar to estimates of new production and of water-column OC remineralization plus deep POC flux, two equivalencies to NCPOC. It is also similar to estimates of export production at OSP made from global modeling, but it is double estimates of NCPOC based on mass balance of mixed-layer tracers. The estimate of NCPOC and a CaCO3:OC export ratio of 0.18 determined from trap data gives a net community production of CaCO3 (NCPIC) of 0.65-0.95 mol m-2 y-1 in agreement with water-column CaCO3 dissolution plus deep CaCO3 flux. The similarity between the CaCO3:POC flux ratio at 50 m and the CaCO3:POC production ratio from bottle incubations (Lipsen et al., 2007) requires that ∼70% of CaCO3 production must dissolve in the euphotic zone to match the rate of POC recycling at OSP. Flux climatologies at 1000 m and 3800 m imply sediments caught at 3800 m include a component sinking rapidly (the primary flux; ∼120-350 m d-1) and another component sinking slowly (the secondary flux; ∼10-20 m d-1). A mass-balance model finds that secondary fluxes contribute ∼40% to the annual mass flux at 3800 m. Based on compositional evidence and on the arrival times at 3800 m, the secondary flux likely derives from disaggregated primary fluxes with an additional lithogenic component transported horizontally to the bathypelagic zone at OSP. Remineralization of BSi, CaCO3, OC and N estimated from decreasing flux with depth in the bathypelagic zone agrees with estimates for the Pacific Ocean based on water-column tracers provided trapping efficiency at 1000 m is 0.6-0.8 and at 3800 m is 1. Alternatively, the estimates based on tracers may include a component of seafloor remineralization. In this case, remineralization in the water column at OSP is at least 30-45% of the remineralization determined by tracers, with the remainder occurring at the seafloor.

Borreliacidal OspC Antibody Response of Canines with Lyme Disease Differs Significantly from That of Humans with Lyme Disease▿

PubMed Central

Lovrich, Steven D.; La Fleur, Rhonda L.; Jobe, Dean A.; Johnson, Jennifer C.; Asp, Krista E.; Schell, Ronald F.; Callister, Steven M.

2007-01-01

Humans reliably produce high concentrations of borreliacidal OspC antibodies specific for the seven C-terminal amino acids shortly after infection with Borrelia burgdorferi. We show that dogs also produce OspC borreliacidal antibodies but that their frequencies, intensities, and antigenicities differ significantly. The findings therefore confirm a major difference between the borreliacidal antibody responses of humans and canines with Lyme disease. PMID:17344346

Lyme neuroborreliosis in 2 horses.

PubMed

Imai, D M; Barr, B C; Daft, B; Bertone, J J; Feng, S; Hodzic, E; Johnston, J M; Olsen, K J; Barthold, S W

2011-11-01

Lyme neuroborreliosis--characterized as chronic, necrosuppurative to nonsuppurative, perivascular to diffuse meningoradiculoneuritis--was diagnosed in 2 horses with progressive neurologic disease. In 1 horse, Borrelia burgdorferi sensu stricto was identified by polymerase chain reaction amplification of B burgdorferi sensu stricto-specific gene targets (ospA, ospC, flaB, dbpA, arp). Highest spirochetal burdens were in tissues with inflammation, including spinal cord, muscle, and joint capsule. Sequence analysis of ospA, ospC, and flaB revealed 99.9% sequence identity to the respective genes in B burgdorferi strain 297, an isolate from a human case of neuroborreliosis. In both horses, spirochetes were visualized in affected tissues with Steiner silver impregnation and by immunohistochemistry, predominantly within the dense collagenous tissue of the dura mater and leptomeninges.

Demonstration of the ability of a canine Lyme vaccine to reduce the incidence of histological synovial lesions following experimentally-induced canine Lyme borreliosis.

PubMed

Grosenbaugh, Deborah A; Rissi, Daniel R; Krimer, Paula M

2016-11-01

Lyme disease in dogs can be effectively prevented by vaccination against antigens expressed by the spirochete Borrelia burgdorferi during transmission by the tick vector Ixodes sp. Lyme vaccine efficacy has traditionally been based on indicators of infection following wild-caught tick challenge whereas most other types of vaccine are required to demonstrate protection from clinical signs of disease. In this vaccination-challenge study we sought to demonstrate the ability of a nonadjuvanted, outer surface protein A (OspA) vaccine to protect from infection and to prevent synovial lesions consistent with Borreliosis. Thirty, purpose-bred beagles were randomly divided into vaccinated and unvaccinated groups. The vaccinated group was administered two subcutaneous doses of a nonadjuvanted, purified, Borrelia burgdorferi OspA vaccine at a 21- day interval. Dogs were challenged by wild-caught, B. burgdorferi-infected ticks (Ixodes scapularis). Clinical signs, serology, Borrelia isolation and PCR evaluated antemortem vaccine efficacy. Postmortem histopathological analysis of synovial tissue was compared to antemortem infection status. Borreliosis was demonstrated by Borrelia isolation from skin biopsies in 13 out of 15 unvaccinated dogs. All unvaccinated dogs' Western blot profiles were consistent with infection. Two of 15 vaccinated dogs had at least one positive spirochete culture which cleared 91days post-challenge, and Western blot profiles were consistent with vaccination alone. No dogs, vaccinated or unvaccinated, exhibited clinical signs consistent with borreliosis. Based on a histopathological cumulative joint scoring system (CJS), all unvaccinated dogs had synovial lesions indicative of Lyme disease. Only one of the vaccinated dogs had a CJS that was greater than the statistical cut off score for the absence of synovial lesions. There was high correlation between clinical histopathology and spirochete isolation. Infection with B burgdorferi may produce inconsistent clinical signs of lameness. Histopathological changes in joints from infected dogs are reliable indicators of borreliosis and correlate well with other indicators of infection. This model provides support that vaccination with a nonadjuvanted, purified OspA vaccine offers protection from Borrelia infection and the resulting synovial lesions that can lead to clinical signs of lameness. Copyright © 2016 Elsevier B.V. All rights reserved.

A large-scale intervention to introduce orange sweet potato in rural Mozambique increases vitamin A intakes among children and women.

PubMed

Hotz, Christine; Loechl, Cornelia; de Brauw, Alan; Eozenou, Patrick; Gilligan, Daniel; Moursi, Mourad; Munhaua, Bernardino; van Jaarsveld, Paul; Carriquiry, Alicia; Meenakshi, J V

2012-07-14

β-Carotene-rich orange sweet potato (OSP) has been shown to improve vitamin A status of infants and young children in controlled efficacy trials and in a small-scale effectiveness study with intensive exposure to project inputs. However, the potential of this important food crop to reduce the risk of vitamin A deficiency in deficient populations will depend on the ability to distribute OSP vines and promote its household production and consumption on a large scale. In rural Mozambique, we conducted a randomised, controlled effectiveness study of a large-scale intervention to promote household-level OSP production and consumption using integrated agricultural, demand creation/behaviour change and marketing components. The following two intervention models were compared: a low-intensity (1 year) and a high-intensity (nearly 3 years) training model. The primary nutrition outcomes were OSP and vitamin A intakes by children 6-35 months and 3-5·5 years of age, and women. The intervention resulted in significant net increases in OSP intakes (model 1: 46, 48 and 97 g/d) and vitamin A intakes (model 1: 263, 254 and 492 μg retinol activity equivalents/d) among the younger children, older children and women, respectively. OSP accounted for 47-60 % of all sweet potato consumed and, among reference children, provided 80 % of total vitamin A intakes. A similar magnitude of impact was observed for both models, suggesting that group-level trainings in nutrition and agriculture could be limited to the first project year without compromising impact. Introduction of OSP to rural, sweet potato-producing communities in Mozambique is an effective way to improve vitamin A intakes.

Quantum Superalgebras at Roots of Unity and Topological Invariants of Three-manifolds

NASA Astrophysics Data System (ADS)

Blumen, Sacha C.

2006-01-01

The general method of Reshetikhin and Turaev is followed to develop topological invariants of closed, connected, orientable 3-manifolds from a new class of algebras called pseudo-modular Hopf algebras. Pseudo-modular Hopf algebras are a class of Z_2-graded ribbon Hopf algebras that generalise the concept of a modular Hopf algebra. The quantum superalgebra U_q(osp(1|2n)) over C is considered with q a primitive N^th root of unity for all integers N >= 3. For such a q, a certain left ideal I of U_q(osp(1|2n)) is also a two-sided Hopf ideal, and the quotient algebra U_q^(N)(osp(1|2n)) = U_q(osp(1|2n)) / I is a Z_2-graded ribbon Hopf algebra. For all n and all N >= 3, a finite collection of finite dimensional representations of U_q^(N)(osp(1|2n)) is defined. Each such representation of U_q^(N)(osp(1|2n)) is labelled by an integral dominant weight belonging to the truncated dominant Weyl chamber. Properties of these representations are considered: the quantum superdimension of each representation is calculated, each representation is shown to be self-dual, and more importantly, the decomposition of the tensor product of an arbitrary number of such representations is obtained for even N. It is proved that the quotient algebra U_q^(N)(osp(1|2n)), together with the set of finite dimensional representations discussed above, form a pseudo-modular Hopf algebra when N >= 6 is twice an odd number. Using this pseudo-modular Hopf algebra, we construct a topological invariant of 3-manifolds. This invariant is shown to be different to the topological invariants of 3-manifolds arising from quantum so(2n+1) at roots of unity.

A locally supersymmetric SO(10, 2) invariant action for D = 12 supergravity

NASA Astrophysics Data System (ADS)

Castellani, Leonardo

2017-06-01

We present an action for N = 1 supergravity in 10 + 2 dimensions, containing the gauge fields of the OSp(1|64) superalgebra, i.e. one-forms B ( n) with n=1,2,5,6,9,10 antisymmetric D=12 Lorentz indices and a Majorana gravitino ψ. The vielbein and spin connection correspond to B (1) and B (2) respectively. The action is not gauge invariant under the full OSp(1|64) superalgebra, but only under a subalgebra \\tilde{F} (containing the F algebra OSp(1|32)), whose gauge fields are B (2), B (6), B (10) and the Weyl projected Majorana gravitino 1/2(1+{Γ}_{13})ψ . Supersymmetry transformations are therefore generated by a Majorana-Weyl supercharge and, being part of a gauge superalgebra, close off-shell. The action is simply ∫ STr( R 6 Γ) where R is the OSp(1|64) curvature supermatrix two-form, and Γ is a constant supermatrix involving Γ13 and breaking OSp(1|64) to its \\tilde{F} subalgebra. The usual Einstein-Hilbert term is included in the action.

Effect of Interfacial Microstructures on the Bonding Strength of Sn-3.0Ag-0.5Cu Pb-Free Solder Bump

NASA Astrophysics Data System (ADS)

Kim, Jae-Myeong; Jeong, Myeong-Hyeok; Yoo, Sehoon; Park, Young-Bae

2012-05-01

The effect of interfacial microstructures on the bonding strength of Sn-3.0Ag-0.5Cu Pb-free solder bumps with respect to the loading speed, annealing time, and surface finish was investigated. The shear strength increased and the ductility decreased with increasing shear speed, primarily because of the time-independent plastic hardening and time-dependent strain-rate sensitivity of the solder alloy. The shear strength and toughness decreased for all surface finishes under the high-speed shear test of 500 mm/s as a result of increasing intermetallic compound (IMC) growth and pad interface weakness associated with increased annealing time. The immersion Sn and organic solderability preservative (OSP) finishes showed lower shear strength compared to the electroless nickel immersion gold (ENIG) finish. With increasing annealing time, the ENIG finish exhibited the pad open fracture mode, whereas the immersion Sn and OSP finishes exhibited the brittle fracture mode. In addition, the shear strength of the solder joints was correlated with each fracture mode.

Flight Demonstrations of Orbital Space Plane (OSP) Technologies

NASA Technical Reports Server (NTRS)

Turner, Susan

2003-01-01

The Orbital Space Plane (OSP) Program embodies NASA s priority to transport Space Station crews safely, reliably, and affordably, while it empowers the Nation s greater strategies for scientific exploration and space leadership. As early in the development cycle as possible, the OSP will provide crew rescue capability, offering an emergency ride home from the Space Station, while accommodating astronauts who are deconditioned due to long- duration missions, or those that may be ill or injured. As the OSP Program develops a fully integrated system, it will use existing technologies and employ computer modeling and simulation. Select flight demonstrator projects will provide valuable data on launch, orbital, reentry, and landing conditions to validate thermal protection systems, autonomous operations, and other advancements, especially those related to crew safety and survival.

Estimation and Validation of Oceanic Mass Circulation from the GRACE Mission

NASA Technical Reports Server (NTRS)

Boy, J.-P.; Rowlands, D. D.; Sabaka, T. J.; Luthcke, S. B.; Lemoine, F. G.

2011-01-01

Since the launch of the Gravity Recovery And Climate Experiment (GRACE) in March 2002, the Earth's surface mass variations have been monitored with unprecedented accuracy and resolution. Compared to the classical spherical harmonic solutions, global high-resolution mascon solutions allows the retrieval of mass variations with higher spatial and temporal sampling (2 degrees and 10 days). We present here the validation of the GRACE global mascon solutions by comparing mass estimates to a set of about 100 ocean bottom pressure (OSP) records, and show that the forward modelling of continental hydrology prior to the inversion of the K-band range rate data allows better estimates of ocean mass variations. We also validate our GRACE results to OSP variations modelled by different state-of-the-art ocean general circulation models, including ECCO (Estimating the Circulation and Climate of the Ocean) and operational and reanalysis from the MERCATOR project.

A lattice approach to the conformal OSp(2S+2|2S) supercoset sigma model. Part I: Algebraic structures in the spin chain. The Brauer algebra

NASA Astrophysics Data System (ADS)

Candu, Constantin; Saleur, Hubert

2009-02-01

We define and study a lattice model which we argue is in the universality class of the OSp(2S+2|2S) supercoset sigma model for a large range of values of the coupling constant gσ2. In this first paper, we analyze in details the symmetries of this lattice model, in particular the decomposition of the space of the quantum spin chain V as a bimodule over OSp(2S+2|2S) and its commutant, the Brauer algebra B(2). It turns out that V is a nonsemisimple module for both OSp(2S+2|2S) and B(2). The results are used in the companion paper to elucidate the structure of the (boundary) conformal field theory.

Interactions of Cd and Cu in anaerobic estuarine sediments. 1: Partitioning in geochemical fractions of sediments

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rule, J.H.; Alden, R.W. III

1996-04-01

Partitioning of Cd and Cu between geochemical fractions of an anaerobic estuarine sediment was determined after equilibrating fine-sandy sediment with different combinations of added Cd (0, 2.5, 5 mg/kg) and Cu(0, 12.5, 25 mg/kg). Sediments were placed in aquaria with 20 ppt seawater where bioassay test organisms were exposed for 14 d. At the start and conclusion of the experimental period, sediments were sequentially extracted to determine the elemental content of the exchangeable (EP), easily reducible (ERP), organic- sulfide (OSP), moderately reducible (MRP), and acid extractable (AEP) phases. Partitioning of the metals in both the native and treated sediments was,more » for Cd: OSP {much_gt} ERP > AEP > EP (MRP was below detection) and for Cu: OSP {much_gt} AEP > ERP > MRP > EP. Cadmium extracted in all phases and Cu in the EP, RP, and OSP were proportional to the respective treatments. The EP-Cd, ERP-Cd, and OSP-Cd were affected by the Cu treatment and significant interactions occurred between Cd and Cu for the EP-Cd, ERP-Cd, OSP-Cd, EP-Cu, and ERP-Cu. Increasing levels of applied Cd and Cu resulted in greater amounts of EP-Cd and ERP-Cd, fractions that are the most bioavailable and the most readily available for desorption into the water column. A significant conclusion is that the input of nontoxic metals may affect the geochemical phase distribution, potential bioavailability, and toxicity of native sediment metals.« less

Oral squamous papilloma occurring on the palate with review of literature.

PubMed

Nayak, Anjali; Nayak, Meghanand T

2016-11-01

Squamous papillomas are common lesions occurring on skin, oral and nasal mucosa and male and female genital organs. Oral squamous cell papilloma (OSP) is a benign proliferation of the stratified squamous epithelium and is generally believed to be caused by Human Papilloma Viruses (HPV). It constitutes around 2.5% of all oral verruco-papillary lesions. We here, report a case of palatal OSP occurring in a 55-year-old male. The aetiological, clinical, diagnostic and treatment aspects of OSP are discussed here. © 2016 Old City Publishing, Inc.

Plasma and memory B cell responses targeting O-specific polysaccharide (OSP) are associated with protection against Vibrio cholerae O1 infection among household contacts of cholera patients in Bangladesh.

PubMed

Aktar, Amena; Rahman, M Arifur; Afrin, Sadia; Akter, Aklima; Uddin, Taher; Yasmin, Tahirah; Sami, Md Israk Nur; Dash, Pinki; Jahan, Sultana Rownok; Chowdhury, Fahima; Khan, Ashraful I; LaRocque, Regina C; Charles, Richelle C; Bhuiyan, Taufiqur Rahman; Mandlik, Anjali; Kelly, Meagan; Kováč, Pavol; Xu, Peng; Calderwood, Stephen B; Harris, Jason B; Qadri, Firdausi; Ryan, Edward T

2018-04-01

The mediators of protection against cholera, a severe dehydrating illness of humans caused by Vibrio cholerae, are unknown. We have previously shown that plasma IgA as well as memory B IgG cells targeting lipopolysaccharide (LPS) of Vibrio cholerae O1 correlate with protection against V. cholerae O1 infection among household contacts of cholera patients. Protection against cholera is serogroup specific, and serogroup specificity is defined by the O-specific polysaccharide (OSP) component of LPS. Therefore, we prospectively followed household contacts of cholera patients to determine whether OSP-specific immune responses present at the time of enrollment are associated with protection against V. cholerae infection. In this study, we enrolled two hundred forty two household contacts of one hundred fifty index patients who were infected with Vibrio cholerae. We determined OSP-specific memory B cells and plasma IgA, IgG and IgM antibody responses on study entry (day 2). The presence of OSP-specific plasma IgA, IgM, and IgG antibody responses on study entry were associated with a decrease in the risk of infection in household contacts (IgA, p = 0.015; IgM, p = 0.01, and IgG, p = 0.024). In addition, the presence of OSP-specific IgG memory B cell responses in peripheral blood on study entry was also associated with a decreased risk of infection (44% reduction; 95% CI: 31.1 to 99.8) in contacts. No protection was associated with cholera toxin B subunit (CtxB)-specific memory B cell responses. These results suggest that immune responses that target OSP, both in plasma and memory responses, may be important in mediating protection against infection with V. cholerae O1.

Plasma and memory B cell responses targeting O-specific polysaccharide (OSP) are associated with protection against Vibrio cholerae O1 infection among household contacts of cholera patients in Bangladesh

PubMed Central

Aktar, Amena; Rahman, M. Arifur; Afrin, Sadia; Akter, Aklima; Uddin, Taher; Yasmin, Tahirah; Sami, Md. Israk Nur; Dash, Pinki; Jahan, Sultana Rownok; Chowdhury, Fahima; Khan, Ashraful I.; LaRocque, Regina C.; Charles, Richelle C.; Bhuiyan, Taufiqur Rahman; Mandlik, Anjali; Kelly, Meagan; Kováč, Pavol; Xu, Peng; Calderwood, Stephen B.; Harris, Jason B.; Ryan, Edward T.

2018-01-01

Background The mediators of protection against cholera, a severe dehydrating illness of humans caused by Vibrio cholerae, are unknown. We have previously shown that plasma IgA as well as memory B IgG cells targeting lipopolysaccharide (LPS) of Vibrio cholerae O1 correlate with protection against V. cholerae O1 infection among household contacts of cholera patients. Protection against cholera is serogroup specific, and serogroup specificity is defined by the O-specific polysaccharide (OSP) component of LPS. Therefore, we prospectively followed household contacts of cholera patients to determine whether OSP-specific immune responses present at the time of enrollment are associated with protection against V. cholerae infection. Methodology In this study, we enrolled two hundred forty two household contacts of one hundred fifty index patients who were infected with Vibrio cholerae. We determined OSP-specific memory B cells and plasma IgA, IgG and IgM antibody responses on study entry (day 2). Principle findings The presence of OSP-specific plasma IgA, IgM, and IgG antibody responses on study entry were associated with a decrease in the risk of infection in household contacts (IgA, p = 0.015; IgM, p = 0.01, and IgG, p = 0.024). In addition, the presence of OSP-specific IgG memory B cell responses in peripheral blood on study entry was also associated with a decreased risk of infection (44% reduction; 95% CI: 31.1 to 99.8) in contacts. No protection was associated with cholera toxin B subunit (CtxB)-specific memory B cell responses. Conclusion These results suggest that immune responses that target OSP, both in plasma and memory responses, may be important in mediating protection against infection with V. cholerae O1. PMID:29684006

A step towards measuring the fetal head circumference with the use of obstetric ultrasound in a low resource setting

NASA Astrophysics Data System (ADS)

van den Heuvel, Thomas L. A.; Petros, Hezkiel; Santini, Stefano; de Korte, Chris L.; van Ginneken, Bram

2017-03-01

Worldwide, 99% of all maternal deaths occur in low-resource countries. Ultrasound imaging can be used to detect maternal risk factors, but requires a well-trained sonographer to obtain the biometric parameters of the fetus. One of the most important biometric parameters is the fetal Head Circumference (HC). The HC can be used to estimate the Gestational Age (GA) and assess the growth of the fetus. In this paper we propose a method to estimate the fetal HC with the use of the Obstetric Sweep Protocol (OSP). With the OSP the abdomen of pregnant women is imaged with the use of sweeps. These sweeps can be taught to somebody without any prior knowledge of ultrasound within a day. Both the OSP and the standard two-dimensional ultrasound image for HC assessment were acquired by an experienced gynecologist from fifty pregnant women in St. Luke's Hospital in Wolisso, Ethiopia. The reference HC from the standard two-dimensional ultrasound image was compared to both the manually measured HC and the automatically measured HC from the OSP data. The median difference between the estimated GA from the manual measured HC using the OSP and the reference standard was -1.1 days (Median Absolute Deviation (MAD) 7.7 days). The median difference between the estimated GA from the automatically measured HC using the OSP and the reference standard was -6.2 days (MAD 8.6 days). Therefore, it can be concluded that it is possible to estimate the fetal GA with simple obstetric sweeps with a deviation of only one week.

Effects of Surface Finishes and Current Stressing on Interfacial Reaction Characteristics of Sn-3.0Ag-0.5Cu Solder Bumps

NASA Astrophysics Data System (ADS)

Kim, Jae-Myeong; Jeong, Myeong-Hyeok; Yoo, Sehoon; Park, Young-Bae

2012-04-01

The effects of surface finishes on the in situ interfacial reaction characteristics of ball grid array (BGA) Sn-3.0Ag-0.5Cu lead-free solder bumps were investigated under annealing and electromigration (EM) test conditions of 130°C to 175°C with 5.0 × 103 A/cm2. During reflow and annealing, (Cu,Ni)6Sn5 intermetallic compound (IMC) formed at the interface of electroless nickel immersion gold (ENIG) finish. In the case of both immersion Sn and organic solderability preservative (OSP) finishes, Cu6Sn5 and Cu3Sn IMCs formed. Overall, the IMC growth velocity of ENIG was much lower than that of the other finishes. The activation energies of total IMCs were found to be 0.52 eV for ENIG, 0.78 eV for immersion Sn, and 0.72 eV for OSP. The ENIG finish appeared to present an effective diffusion barrier between the Cu substrate and the solder, which leads to better EM reliability in comparison with Cu-based pad systems. The failure mechanisms were explored in detail via in situ EM tests.

Reflection matrices with U q [osp(2) (2|2m)] symmetry

NASA Astrophysics Data System (ADS)

Vieira, R. S.; Lima-Santos, A.

2017-09-01

We propose a classification of the reflection K-matrices (solutions of the boundary Yang-Baxter equation) for the Uq[osp(2)(2\\vert 2m)]=Uq[C(2)(m+1)] vertex-model. We found four families of solutions, namely, the complete solutions, in which no elements of the reflection K-matrix is null, the block-diagonal solutions, the X-shape solutions and the diagonal solutions. We highlight that these diagonal K-matrices also hold for the Uq[osp(2)(2n+2\\vert 2m)]=Uq[D(2)(n+1, m)] vertex-model.

On representations of U{sub q}osp(1{vert_bar}2) when q is a root of unity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chung, W.; Suzuki, T.

1997-06-01

The infinite dimensional highest weight representations of U{sub q}osp(1{vert_bar}2) for the deformation parameter q being a root of unity are investigated. As in the cases of q-deformed nongraded Lie algebras, we find that every irreducible representation is isomorphic to the tensor product of a highest weight representation of sl{sub 2}(R) and a finite dimensional one of U{sub q}osp(1{vert_bar}2). The structure is investigated in detail. {copyright} {ital 1997 American Institute of Physics.}

A biochip-based combined immunoassay for detection of serological status of Borrelia burgdorferi in Lyme borreliosis.

PubMed

Huang, Na-Li; Ye, Lei; Lv, Hui; Du, Yi-Xin; Schneider, Marion; Fan, Li-Bin; Du, Wei-Dong

2017-09-01

Dithiobis (succinimidyl undecanoate) modified gold surface biochip were used as a combined immunoassay platform for concurrently detecting immune responses to Borrelia burgdorferi (B. burgdorferi) sensu lato antigens, flagellin, outer surface protein C, variable major protein-like sequence proteins, and 3 VlsE protein IR 6 peptides. The peptides represented intrinsic Borrelia genospecies: B. burgdorferi sensu stricto, B. garinii, and B. afzelii, respectively. Fourier transform infrared spectroscopy was utilized to validate the surface chemical characteristics on the modified gold surface. The limits in detection of IgG antibody on the biochips were as little as 0.39μg/ml for anti-VlsE and 0.78μg/ml for anti-flagellin and anti-OspC, respectively. Samples from 56 neuroborreliosis (NB) patients and 114 healthy individuals were analyzed by the combined biochip. We found that the seroprevalences of IgM or IgG antibody against the 6 antigens were contributed to increased overall sensitivity by the multiplex immunobiochip assay. Serum combined positive rates of the 6 antigens in the patients were 92.86% for IgM antibody and 91.07% for IgG antibody. Part of the patients bore antibody responses against the 3 VlsE IR 6 variant peptides, indicating that Lyme borreliosis would attribute to consequence of multiple infections by one or more Borrelia burgdorferi strains. Concurrent assessment for both IgM and IgG antibodies against the protein antigens and B. burgdorferi IR 6 peptides in the sera of NB patients was beneficial from the biochip format, enabling detection of expanded serologic infection status and therapy strategy-making more efficiently. The combined biochip-based immunoassay, as a potential substitution of ELISA, provided a promising approach to extend the detection spectrum of infectious antibodies against a panel of Borrelia antigens. Copyright © 2017 Elsevier B.V. All rights reserved.

EspO1-2 Regulates EspM2-Mediated RhoA Activity to Stabilize Formation of Focal Adhesions in Enterohemorrhagic Escherichia coli-Infected Host Cells

PubMed Central

Iyoda, Sunao; Izumiya, Hidemasa; Watanabe, Haruo; Ohnishi, Makoto; Terajima, Jun

2013-01-01

Enterohemorrhagic Escherichia coli (EHEC) Sakai strain encodes two homologous type III effectors, EspO1-1 and EspO1-2. These EspO1s have amino acid sequence homology with Shigella OspE, which targets integrin-linked kinase to stabilize formation of focal adhesions (FAs). Like OspE, EspO1-1 was localized to FAs in EHEC-infected cells, but EspO1-2 was localized in the cytoplasm. An EHEC ΔespO1-1ΔespO1-2 double mutant induced cell rounding and FA loss in most of infected cells, but neither the ΔespO1-1 nor ΔespO1-2 single mutant did. These results suggested that EspO1-2 functioned in the cytoplasm by a different mechanism from EspO1-1 and OspE. Since several type III effectors modulate Rho GTPase, which contributes to FA formation, we investigated whether EspO1-2 modulates the function of these type III effectors. We identified a direct interaction between EspO1-2 and EspM2, which acts as a RhoA guanine nucleotide exchange factor. Upon ectopic co-expression, EspO1-2 co-localized with EspM2 in the cytoplasm and suppressed EspM2-mediated stress fiber formation. Consistent with these findings, an ΔespO1-1ΔespO1-2ΔespM2 triple mutant did not induce cell rounding in epithelial cells. These results indicated that EspO1-2 interacted with EspM2 to regulate EspM2-mediated RhoA activity and stabilize FA formation during EHEC infection. PMID:23409096

Sweet taste preference in binge-eating disorder: A preliminary investigation.

PubMed

Goodman, Erica L; Breithaupt, Lauren; Watson, Hunna J; Peat, Christine M; Baker, Jessica H; Bulik, Cynthia M; Brownley, Kimberly A

2018-01-01

Research suggests that individuals with high liking for sweets are at increased risk for binge eating, which has been minimally investigated in individuals with binge-eating disorder (BED). Forty-one adults (85% female, 83% white) with binge eating concerns completed a sweet taste test and measures of eating disorder behaviors and food cravings. A subset of participants with BED completed an oral glucose tolerance test (OGTT; N=21) and a 24-hour dietary recall (N=26). Regression models were used to compare highest sweet preferers (HSP [N=18]) to other sweet preferers (OSP [N=23]) and were used to assess associations between sweet taste preference and outcome variables. Effect sizes (ηp 2 ) for differences between HSP and OSP ranged from small (≤0.01) to large (≥0.24); group differences were statistically nonsignificant except for 24-hour caloric intake (ηp 2 =0.16, p=0.04), protein intake (ηp 2 =0.16, p=0.04), and insulin sensitivity index (ηp 2 =0.24, p=0.04), which were higher in HSP, and postprandial insulin, which was smaller in HSP (ηp 2 =0.27, p=0.03). Continuous analyses replicated postprandial insulin response. Compared with OSP, HSP reported numerically higher binge-eating frequency (ηp 2 =0.04), over-eating frequency (ηp 2 =0.06), and carbohydrate intake (ηp 2 =0.14), and they exhibited numerically smaller postprandial glucose AUC (ηp 2 =0.16). Sweet taste preference may have implications for glucose regulation, binge-eating frequency, and nutrient intake in BED. Copyright © 2017 Elsevier Ltd. All rights reserved.

Quantum mechanics and hidden superconformal symmetry

NASA Astrophysics Data System (ADS)

Bonezzi, R.; Corradini, O.; Latini, E.; Waldron, A.

2017-12-01

Solvability of the ubiquitous quantum harmonic oscillator relies on a spectrum generating osp (1 |2 ) superconformal symmetry. We study the problem of constructing all quantum mechanical models with a hidden osp (1 |2 ) symmetry on a given space of states. This problem stems from interacting higher spin models coupled to gravity. In one dimension, we show that the solution to this problem is the Vasiliev-Plyushchay family of quantum mechanical models with hidden superconformal symmetry obtained by viewing the harmonic oscillator as a one dimensional Dirac system, so that Grassmann parity equals wave function parity. These models—both oscillator and particlelike—realize all possible unitary irreducible representations of osp (1 |2 ).

Comparison of Oogenesis and Sex Steroid Profiles between Twice and Once Annually Spawning of Rainbow Trout Females (Oncorhynchus mykiss)

PubMed Central

Estay, Francisco; Colihueque, Nelson; Araneda, Cristian

2012-01-01

This study compares the gonadosomatic index (GSI), oocyte growth (OG), gonadal histology, and plasma level concentrations of sex hormones (estradiol-17β (E2) and vitellogenin (V)) of twice-spawning (T-SP) and once-spawning (O-SP) females of rainbow trout throughout the additional and the normal reproductive cycle, respectively. In T-SP, the GSI values rapidly increase from May to November, in contrast to O-SP, which showed low and constant GSI values (1.19 to 14.5 and 1.19 to 0.63, resp.). T-SP exhibited a marked increase of OG in the same period, reaching a maximum diameter of 4,900 ± 141.42 μm, in contrast to O-SP, which presented a slow OG. The gonadal histology of T-SP agreed with the general pattern of ovogenesis observed for O-SP (vitellogenesis, ovulation, and recrudescence); however, this process was nonsynchronous between the two breeder groups. Plasma steroid levels showed significant variation during oogenesis, which agreed with the GSI, OG, and gonadal histology patterns. The level of E2 increased to a maximum value of 26.2 ng/mL and 36.0 ng/mL in O-SP and T-SP, respectively, one or two months before the spawning event where vitellogenesis was fully active. The V concentrations followed a pattern similar to those of E2. PMID:23213308

Reductions in human Lyme disease risk due to the effects of oral vaccination on tick-to-mouse and mouse-to-tick transmission.

PubMed

Voordouw, Maarten J; Tupper, Haley; Önder, Özlem; Devevey, Godefroy; Graves, Christopher J; Kemps, Brian D; Brisson, Dustin

2013-04-01

Vaccinating wildlife is becoming an increasingly popular method to reduce human disease risks from pathogens such as Borrelia burgdorferi, the causative agent of Lyme disease. To successfully limit human disease risk, vaccines targeting the wildlife reservoirs of B. burgdorferi must be easily distributable and must effectively reduce pathogen transmission from infected animals, given that many animals in nature will be infected prior to vaccination. We assessed the efficacy of an easily distributable oral bait vaccine based on the immunogenic outer surface protein A (OspA) to protect uninfected mice from infection and to reduce transmission from previously infected white-footed mice, an important reservoir host of B. burgdorferi. Oral vaccination of white-footed mice effectively reduces transmission of B. burgdorferi at both critical stages of the Lyme disease transmission cycle. First, oral vaccination of uninfected white-footed mice elicits an immune response that protects mice from B. burgdorferi infection. Second, oral vaccination of previously infected mice significantly reduces the transmission of B. burgdorferi to feeding ticks despite a statistically nonsignificant immune response. We used the estimates of pathogen transmission to and from vaccinated and unvaccinated mice to model the efficacy of an oral vaccination campaign targeting wild white-footed mice. Projection models suggest that the effects of the vaccine on both critical stages of the transmission cycle of B. burgdorferi act synergistically in a positive feedback loop to reduce the nymphal infection prevalence, and thus human Lyme disease risk, well below what would be expected from either effect alone. This study suggests that oral immunization of wildlife with an OspA-based vaccine can be a promising long-term strategy to reduce human Lyme disease risk.

Seaglider surveys at Ocean Station Papa: Diagnosis of upper-ocean heat and salt balances using least squares with inequality constraints

NASA Astrophysics Data System (ADS)

Pelland, Noel A.; Eriksen, Charles C.; Cronin, Meghan F.

2017-06-01

Heat and salt balances in the upper 200 m are examined using data from Seaglider spatial surveys June 2008 to January 2010 surrounding a NOAA surface mooring at Ocean Station Papa (OSP; 50°N, 145°W). A least-squares approach is applied to repeat Seaglider survey and moored measurements to solve for unknown or uncertain monthly three-dimensional circulation and vertical diffusivity. Within the surface boundary layer, the estimated heat and salt balances are dominated throughout the surveys by turbulent flux, vertical advection, and for heat, radiative absorption. When vertically integrated balances are considered, an estimated upwelling of cool water balances the net surface input of heat, while the corresponding large import of salt across the halocline due to upwelling and diffusion is balanced by surface moisture input and horizontal import of fresh water. Measurement of horizontal gradients allows the estimation of unresolved vertical terms over more than one annual cycle; diffusivity in the upper-ocean transition layer decreases rapidly to the depth of the maximum near-surface stratification in all months, with weak seasonal modulation in the rate of decrease and profile amplitude. Vertical velocity is estimated to be on average upward but with important monthly variations. Results support and expand existing evidence concerning the importance of horizontal advection in the balances of heat and salt in the Gulf of Alaska, highlight time and depth variability in difficult-to-measure vertical transports in the upper ocean, and suggest avenues of further study in future observational work at OSP.

The major autoantibody epitope on factor H in atypical hemolytic uremic syndrome is structurally different from its homologous site in factor H-related protein 1, supporting a novel model for induction of autoimmunity in this disease.

PubMed

Bhattacharjee, Arnab; Reuter, Stefanie; Trojnár, Eszter; Kolodziejczyk, Robert; Seeberger, Harald; Hyvärinen, Satu; Uzonyi, Barbara; Szilágyi, Ágnes; Prohászka, Zoltán; Goldman, Adrian; Józsi, Mihály; Jokiranta, T Sakari

2015-04-10

Atypical hemolytic uremic syndrome (aHUS) is characterized by complement attack against host cells due to mutations in complement proteins or autoantibodies against complement factor H (CFH). It is unknown why nearly all patients with autoimmune aHUS lack CFHR1 (CFH-related protein-1). These patients have autoantibodies against CFH domains 19 and 20 (CFH19-20), which are nearly identical to CFHR1 domains 4 and 5 (CFHR14-5). Here, binding site mapping of autoantibodies from 17 patients using mutant CFH19-20 constructs revealed an autoantibody epitope cluster within a loop on domain 20, next to the two buried residues that are different in CFH19-20 and CFHR14-5. The crystal structure of CFHR14-5 revealed a difference in conformation of the autoantigenic loop in the C-terminal domains of CFH and CFHR1, explaining the variation in binding of autoantibodies from some aHUS patients to CFH19-20 and CFHR14-5. The autoantigenic loop on CFH seems to be generally flexible, as its conformation in previously published structures of CFH19-20 bound to the microbial protein OspE and a sialic acid glycan is somewhat altered. Cumulatively, our data suggest that association of CFHR1 deficiency with autoimmune aHUS could be due to the structural difference between CFHR1 and the autoantigenic CFH epitope, suggesting a novel explanation for CFHR1 deficiency in the pathogenesis of autoimmune aHUS. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

NASA's New Orbital Space Plane: A Bridge to the Future

NASA Technical Reports Server (NTRS)

Davis, Stephan R.; Engler, Leah M.; Fisher, Mark F.; Dumbacher, Dan L.; Boswell, Barry E.

2003-01-01

NASA is developing a new spacecraft system called the Orbital Space Plane (OSP). The OSP will be launched on an expendable launch vehicle and serve to augment the shuttle in support of the International Space Station by transporting astronauts to and from the International Space Station and by providing a crew rescue system.

Annual net community production and the biological carbon flux in the ocean

NASA Astrophysics Data System (ADS)

Emerson, Steven

2014-01-01

The flux of biologically produced organic matter from the surface ocean (the biological pump), over an annual cycle, is equal to the annual net community production (ANCP). Experimental determinations of ANCP at ocean time series sites using a variety of different metabolite mass balances have made it possible to evaluate the accuracy of sediment trap fluxes and satellite-determined ocean carbon export. ANCP values at the Hawaii Ocean Time-series (HOT), the Bermuda Atlantic Time-series Study (BATS), Ocean Station Papa (OSP) are 3 ± 1 mol C m-2 yr-1—much less variable than presently suggested by satellite remote sensing measurements and global circulation models. ANCP determined from mass balances at these locations are 3-4 times particulate organic carbon fluxes measured in sediment traps. When the roles of dissolved organic carbon (DOC) flux, zooplankton migration, and depth-dependent respiration are considered these differences are reconciled at HOT and OSP but not at BATS, where measured particulate fluxes are about 3 times lower than expected. Even in the cases where sediment trap fluxes are accurate, it is not possible to "scale up" these measurements to determine ANCP without independent determinations of geographically variable DOC flux and zooplankton migration. Estimates of ANCP from satellite remote sensing using net primary production determined by the carbon-based productivity model suggests less geographic variability than its predecessor (the vertically generalized productivity model) and brings predictions at HOT and OSP closer to measurements; however, satellite-predicted ANCP at BATS is still 3 times too low.

78 FR 19002 - Marine Mammal Protection Act; Draft Revised Stock Assessment Reports for Two Stocks of West...

Federal Register 2010, 2011, 2012, 2013, 2014

2013-03-28

... optimum sustainable population level (OSP). OSP is defined under the MMPA as '' * * * the number of animals which will result in the maximum productivity of the population or the species, keeping in mind... include: 1. A description of the stock and its geographic range; 2. A minimum population estimate, maximum...

A Molecular Epidemiologic Case-Case Study of Prostate Cancer Susceptibility.

DTIC Science & Technology

1999-09-01

Thompson, P.A., and Barbour, A. A flagella-less mutant as a live attenuated vaccine against Borrelia Burgdorferi infection in mouse model of lyme ...Immunol., 148:3385, 1992. Sadienze A., Rosa, P. A., Thompson, P. A., Hogan, D. M., and. Barbou,r A. G. Antibody-resistant mutants of Borrelia ...cells and OspA protein of Borrelia burgdorferi. New York Acad. Sci. 797:140-150, 1996. Thompson, P. A. and Berton, M. T. STAT6 is required for IL-4

Evaluation of the DC Opportunity Scholarship Program: Impacts after One Year. NCEE 2017-4022

ERIC Educational Resources Information Center

Dynarski, Mark; Rui, Ning; Webber, Ann; Gutmann, Babette

2017-01-01

The District of Columbia Opportunity Scholarship Program (OSP) was created by Congress to provide tuition vouchers to low-income parents who want their child to attend a private school. The Scholarships for Opportunity and Results (SOAR) Act of 2011 also mandated an evaluation of the OSP program. This report examines impacts one year after…

OSP Parameters and the Cognitive Component of Reaction Time to a Missing Stimulus: Linking Brain and Behavior

ERIC Educational Resources Information Center

Hernandez, Oscar H.; Vogel-Sprott, Muriel

2009-01-01

This within-subjects experiment tested the relationship between the premotor (cognitive) component of reaction time (RT) to a missing stimulus and parameters of the omitted stimulus potential (OSP) brain wave. Healthy young men (N = 28) completed trials with an auditory stimulus that recurred at 2 s intervals and ceased unpredictably. Premotor RT…

Peyton's four-step approach for teaching complex spinal manipulation techniques - a prospective randomized trial.

PubMed

Gradl-Dietsch, Gertraud; Lübke, Cavan; Horst, Klemens; Simon, Melanie; Modabber, Ali; Sönmez, Tolga T; Münker, Ralf; Nebelung, Sven; Knobe, Matthias

2016-11-03

The objectives of this prospective randomized trial were to assess the impact of Peyton's four-step approach on the acquisition of complex psychomotor skills and to examine the influence of gender on learning outcomes. We randomly assigned 95 third to fifth year medical students to an intervention group which received instructions according to Peyton (PG) or a control group, which received conventional teaching (CG). Both groups attended four sessions on the principles of manual therapy and specific manipulative and diagnostic techniques for the spine. We assessed differences in theoretical knowledge (multiple choice (MC) exam) and practical skills (Objective Structured Practical Examination (OSPE)) with respect to type of intervention and gender. Participants took a second OSPE 6 months after completion of the course. There were no differences between groups with respect to the MC exam. Students in the PG group scored significantly higher in the OSPE. Gender had no additional impact. Results of the second OSPE showed a significant decline in competency regardless of gender and type of intervention. Peyton's approach is superior to standard instruction for teaching complex spinal manipulation skills regardless of gender. Skills retention was equally low for both techniques.

"Alcohol and nicotine"--Concept and evaluation of an interdisciplinary elective course with OSPE in preclinical medical education.

PubMed

Bergelt, Corinna; Lauke, Heidrun; Petersen-Ewert, Corinna; Jücker, Manfred; Bauer, Christiane K

2014-01-01

In the last decade, increasing interest has been paid to interdisciplinary and practical courses in the medical education in Germany. This report describes the implementation and outcome of a preclinical interdisciplinary elective course with a team-teaching concept developed by lecturers in medical psychology, anatomy, physiology and biochemistry. The practical orientation of the course led to the implementation of a final interdisciplinary OSPE to ensure fair consideration of the different disciplines involved in grading. Individual OSPE results correlate well with the fact that different skills are required in medical psychology compared to those required in anatomy, physiology and biochemistry. Student course evaluation and lecturers` experience indicate the success of this elective course. Its concept can be well adapted to other interdisciplinary courses.

Optimal Design of Integrated Systems Health Management (ISHM) Systems for improving safety in NASA's Exploration Vehicles: A Two-Level Multidisciplinary Design Approach

NASA Technical Reports Server (NTRS)

Mehr, Ali Farhang; Tumer, Irem; Barszcz, Eric

2005-01-01

Integrated Vehicle Health Management (ISHM) systems are used to detect, assess, and isolate functional failures in order to improve safety of space systems such as Orbital Space Planes (OSPs). An ISHM system, as a whole, consists of several subsystems that monitor different components of an OSP including: Spacecraft, Launch Vehicle, Ground Control, and the International Space Station. In this research, therefore, we propose a new methodology to design and optimize ISHM as a distributed system with multiple disciplines (that correspond to different subsystems of OSP safety). A paramount amount of interest has been given in the literature to the multidisciplinary design optimization of problems with such architecture (as will be reviewed in the full paper).

Deformed supersymmetric quantum mechanics with spin variables

NASA Astrophysics Data System (ADS)

Fedoruk, Sergey; Ivanov, Evgeny; Sidorov, Stepan

2018-01-01

We quantize the one-particle model of the SU(2|1) supersymmetric multiparticle mechanics with the additional semi-dynamical spin degrees of freedom. We find the relevant energy spectrum and the full set of physical states as functions of the mass-dimension deformation parameter m and SU(2) spin q\\in (Z_{>0,}1/2+Z_{≥0}) . It is found that the states at the fixed energy level form irreducible multiplets of the supergroup SU(2|1). Also, the hidden superconformal symmetry OSp(4|2) of the model is revealed in the classical and quantum cases. We calculate the OSp(4|2) Casimir operators and demonstrate that the full set of the physical states belonging to different energy levels at fixed q are unified into an irreducible OSp(4|2) multiplet.

Protracted tectono-metamorphic history of the SE Superior Province : contribution of 40Ar/39Ar thermochronology in the Abitibi-Opatica contact zone, Québec, Canada

NASA Astrophysics Data System (ADS)

Daoudene, Yannick; Tremblay, Alain; Ruffet, Gilles; Leclerc, François; Goutier, Jean

2015-04-01

Archean orogens mainly consist of greenstone belts juxtaposing deeper crustal domains of TTG-type plutonic rocks. The greenstone belts show regional folds, penetrative steeply-dipping fabrics, and localised shear zones, whereas the plutonic belts predominantly display dome structures. Concurrently, rocks in Archean orogens undergone MT/HT-LP/MP metamorphic conditions that vary, from upper to lower crustal domains, between greenschist- and granulite-facies, respectively. These structural and metamorphic variations are well-documented, but modes of deformation related to such orogens is still debated. Some studies suggest that the Archean tectonic processes were comparable to present-day plate tectonics and the Archean greenstone belts were interpreted as tectonic collages commonly documented in Phanerozoic subduction/collision zones. Alternative models propose that the Archean tectonics were different from those predicted by the plate tectonics paradigm, mainly due to the existence of a hotter mantle and a mechanically weak crust. In such models, the burying and exhumation of crustal rocks are attributed to the vertical transfer of material, resulting in the development of pop-down and domes structures. As a contribution of the study of mechanisms that might have operated during the Archean, we present a structural and metamorphic study of the contact zone between the Abitibi subprovince (ASP), which contains greenstone belts, and the Opatica subprovince (OSP), which is dominated by plutonic rocks, of the Superior Province. The 40Ar/39Ar dating of amphiboles and micas is used to constrain the age and duration of regional metamorphism and associated deformations. On the basis of seismic profiling, showing a north-dipping lithospheric-scale reflector, the ASP-OSP contact has been interpreted as the surficial trace of an Archean subduction zone. However, our structural analysis suggest that the ASP overlies the OSP and that the ASP-OSP contact does not show evidences of an important sub-vertical shearing deformation as expected if it was a major upper plate-lower plate boundary. Furthermore, the contact does not present significant metamorphic break between the two domains, but a progressive increasing of metamorphism toward the OSP, from greenschist- to amphibolite-facies conditions. Based on these structural and metamorphic characteristics, we suggest that the OSP exposes the deepest rocks at outcrop of an ASP-OSP crust in the study area. Regionally, the 40Ar/39Ar ages acquired during this study indicate that the ASP-OSP contact records a protracted metamorphic history that started around 2685 Ma. The structural and isotopic age data suggest that, from ~2685 Ma to ~2632 Ma, the deepest level of the ASP and the underlying OSP reached amphibolite-facies metamorphic conditions and that regional deformation was accommodated by an overall horizontal shortening and sub-vertical transfers of crustal material. Subsequently, the cooling of these crustal rocks was accompanied by strain localisation, which led to the development of oblique strike-slip shear zones from ~2600 Ma, when the lateral flowing of crustal material became predominant. Our 40Ar/39Ar data compared with metamorphic ages documented in adjacent areas of the Superior Province suggests that the peak and duration of regional metamorphism might have been coeval over a large region. This rather favours a mode of pervasive deformation as expected in vertical tectonics.

Effect of Pneumoperitoneum and Lateral Position on Oropharyngeal Seal Pressures of Proseal LMA in Laparoscopic Urological Procedures.

PubMed

Rustagi, Preeti; Patkar, Geeta A; Ourasang, Anil Kumar; Tendolkar, Bharati A

2017-02-01

A sustained and effective oropharyngeal sealing with supraglottic airway is required to maintain the ventilation during laparoscopic surgery. Previous studies have observed the Oropharyngeal Seal Pressure (OSP) for Proseal Laryngeal Mask Airway (PLMA) after pneumoperitoneum in supine and trendelenburg position, where PLMA was found to be an effective airway device. This study was conducted with ProSeal LMA, for laparoscopic Urologic procedures done in lateral position. To measure OSP in supine and lateral position and to observe the effect of pneumoperitoneum in lateral position on OSP. Secondary objectives were to assess adequacy of ventilation and incidence of adverse events. A total number of 25 patients of American Society of Anaesthesiologists (ASA) physical status II and I were enrolled. After induction of anaesthesia using a standardized protocol, PLMA was inserted. Ryle's tube was inserted through drain tube. The position of PLMA was confirmed with ease of insertion of Ryle's tube and fibreoptic grading of vocal cords. Patients were then put in lateral position. The OSP was measured in supine position. This value was baseline comparison for OSP in lateral position and that after pneumoperitoneum. We assessed the efficacy of PLMA for ventilation, after carboperitoneum in lateral position (peak airway pressure, End Tidal Carbon dioxide (EtCO 2 ), SPO 2 ). Incidence of adverse effects (displacement of device, gastric insufflation, regurgitation, coughing, sore throat, blood on device, trauma) was also noted. The OSP was above Peak Airway Pressure (PAP) in supine (22.1±5.4 and 15.4±4.49cm of H 2 O) and lateral position (22.6±5.3 and 16.1±4.6). After pneumoperitoneum, which was in lateral position, there was statistically significant (p-value <0.05) increase in both PAP (19.96±4.015) and OSP (24.32±4.98, p-value 0.03). There was no intraoperative displacement of PLMA. There was no event of suboptimal oxygenation. EtCO 2 was always within normal limits. Gastric insufflation was present in one patient. One patient had coughing and blood was detected on device. Three patients had throat discomfort post-operatively. In this study, Oropharyngeal seal pressures with PLMA were found to increase after pneumoperitoneum in lateral position. PLMA forms an effective seal around airway and is an efficient and safe alternative for airway management in urological laparoscopic surgeries done in lateral position.

Long-Term Observations of Ocean Biogeochemistry with Nitrate and Oxygen Sensors in Apex Profiling Floats

NASA Astrophysics Data System (ADS)

Johnson, K. S.; Coletti, L.; Jannasch, H.; Martz, T.; Swift, D.; Riser, S.

2008-12-01

Long-term, autonomous observations of ocean biogeochemical cycles are now feasible with chemical sensors in profiling floats. These sensors will enable decadal-scale observations of trends in global ocean biogeochemical cycles. Here, we focus on measurements on nitrate and dissolved oxygen. The ISUS (In Situ Ultraviolet Spectrophotometer) optical nitrate sensor has been adapted to operate in a Webb Research, Apex profiling float. The Apex float is of the type used in the Argo array and is designed for multi-year, expendable deployments in the ocean. Floats park at 1000 m depth and make 60 nitrate and oxygen measurements at depth intervals ranging from 50 m below 400 m to 5 m in the upper 100 m as they profile to the surface. All data are transmitted to shore using the Iridium telemetry system and they are available on the Internet in near-real time. Floats equipped with ISUS and an Aanderaa oxygen sensor are capable of making 280 vertical profiles from 1000 m. At a 5 day cycle time, the floats should have nearly a four year endurance. Three floats have now been deployed at the Hawaii Ocean Time series station (HOT), Ocean Station Papa (OSP) in the Gulf of Alaska and at 50 South, 30 East in the Southern Ocean. Two additional floats are designated for deployment at the Bermuda Atlantic Time Series station (BATS) and in the Drake Passage. The HOT float has made 56 profiles over 260 days and should continue operating for 3 more years. Nitrate concentrations are in excellent agreement with the long-term mean observed at HOT. No significant long-term drift in sensor response has occurred. A variety of features have been observed in the HOT nitrate data that are linked to contemporaneous changes in oxygen production and mesoscale dynamics. The impacts of these features will be briefly described. The Southern Ocean float has operated for 200 days and is now observing reinjection of nitrate into surface waters as winter mixing occurs(surface nitrate > 24 micromolar). We expect that the OSP and Southern Ocean floats will provide a quantitative measurement of the timing and magnitude of the spring bloom via the drawdown of surface nitrate. We are funded through NSF and NOPP to continue float deployments at HOT, BATS, OSP and the Southern Ocean for the next 3 years and to refine the sensor so it can be offered as a commercial option for all float users. New sensors in development for float deployments include a stable ISFET pH sensor.

Seaglider surveys at Ocean Station Papa: Circulation and water mass properties in a meander of the North Pacific Current

NASA Astrophysics Data System (ADS)

Pelland, Noel A.; Eriksen, Charles C.; Cronin, Meghan F.

2016-09-01

A Seaglider autonomous underwater vehicle augmented the Ocean Station Papa (OSP; 50°N, 145°W) surface mooring, measuring spatial structure on scales relevant to the monthly evolution of the moored time series. During each of three missions from June 2008 to January 2010, a Seaglider made biweekly 50 km × 50 km surveys in a bowtie-shaped survey track. Horizontal temperature and salinity gradients measured by these surveys were an order of magnitude stronger than climatological values and sometimes of opposite sign. Geostrophically inferred circulation was corroborated by moored acoustic Doppler current profiler measurements and AVISO satellite altimetry estimates of surface currents, confirming that glider surveys accurately resolved monthly scale mesoscale spatial structure. In contrast to climatological North Pacific Current circulation, upper-ocean flow was modestly northward during the first half of the 18 month survey period, and weakly westward during its latter half, with Rossby number O>(0.01>). This change in circulation coincided with a shift from cool and fresh to warm, saline, oxygen-rich water in the upper-ocean halocline, and an increase in vertical fine structure there and in the lower pycnocline. The anomalous flow and abrupt water mass transition were due to the slow growth of an anticyclonic meander within the North Pacific Current with radius comparable to the scale of the survey pattern, originating to the southeast of OSP.

Does providing more services increase the primary hospitals' revenue? An assessment of national essential medicine policy based on 2,675 counties in China.

PubMed

Chen, Fei; Yang, Min; Li, Qian; Pan, Jay; Li, Xiaosong; Meng, Qun

2018-01-01

To understand whether the increased outpatient service provision (OSP) brings in enough additional income (excluding income from essential medicine) for primary hospitals (INCOME) to compensate for reduced costs of medicine. The two outcomes, annual OSP and INCOME for the period of 2008-2012, were collected from 34,506 primary hospitals in 2,675 counties in 31 provinces in China by the national surveillance system. The data had a four-level hierarchical structure; time points were nested within primary hospital, hospitals within county, and counties within province. We fitted bivariate five-level random effects regression models to examine correlations between OSP and INCOME in terms of their mean values and dose-response effects of the essential medicine policy (EMP). We adjusted for the effects of time period and selected hospital resources. The estimated correlation coefficients between the two outcomes' mean values were strongly positive among provinces (r = 0.910), moderately positive among counties (r = 0.380), and none among hospitals (r = 0.002) and time (r = 0.007). The correlation between their policy effects was weakly positive among provinces (r = 0.234), but none at the county and hospital levels. However, there were markedly negative correlation coefficients between the mean and policy effects at -0.328 for OSP and -0.541 for INCOME at the hospital level. There was no evidence to suggest an association between the two outcomes in terms of their mean values and dose-response effects of EMP at the hospital level. This indicated that increased OSP did not bring enough additional INCOME. Sustainable mechanisms to compensate primary hospitals are needed.

Toward a hybrid brain-computer interface based on repetitive visual stimuli with missing events.

PubMed

Wu, Yingying; Li, Man; Wang, Jing

2016-07-26

Steady-state visually evoked potentials (SSVEPs) can be elicited by repetitive stimuli and extracted in the frequency domain with satisfied performance. However, the temporal information of such stimulus is often ignored. In this study, we utilized repetitive visual stimuli with missing events to present a novel hybrid BCI paradigm based on SSVEP and omitted stimulus potential (OSP). Four discs flickering from black to white with missing flickers served as visual stimulators to simultaneously elicit subject's SSVEPs and OSPs. Key parameters in the new paradigm, including flicker frequency, optimal electrodes, missing flicker duration and intervals of missing events were qualitatively discussed with offline data. Two omitted flicker patterns including missing black/white disc were proposed and compared. Averaging times were optimized with Information Transfer Rate (ITR) in online experiments, where SSVEPs and OSPs were identified using Canonical Correlation Analysis in the frequency domain and Support Vector Machine (SVM)-Bayes fusion in the time domain, respectively. The online accuracy and ITR (mean ± standard deviation) over nine healthy subjects were 79.29 ± 18.14 % and 19.45 ± 11.99 bits/min with missing black disc pattern, and 86.82 ± 12.91 % and 24.06 ± 10.95 bits/min with missing white disc pattern, respectively. The proposed BCI paradigm, for the first time, demonstrated that SSVEPs and OSPs can be simultaneously elicited in single visual stimulus pattern and recognized in real-time with satisfied performance. Besides the frequency features such as SSVEP elicited by repetitive stimuli, we found a new feature (OSP) in the time domain to design a novel hybrid BCI paradigm by adding missing events in repetitive stimuli.

Outer Space Place: Exploring Space at the Maryland Science Center

NASA Astrophysics Data System (ADS)

Jan, M. W.; Mendez, F.

1999-05-01

The Maryland Science Center has been the state's premier vehicle for informal science education for over 20 years. Every day thousands of school children, families, and out-of-state visitors come for fun and come away with ideas, exciting experiences, and an appetite for more information about science. Opened on April 15, 1999, Outer Space Place (OSP) consolidates the Science Center's space exhibits and activities, both new and refurbished. In this paper, we describe OSP, which features SpaceLink, the Crosby Ramsey Memorial Observatory, the Davis Planetarium, Earth Orbit Gallery, and the Hubble Space Telescope National Visitor Center and provides hands-on educational experiences for kids of all ages. We illustrate how astronomers contribute to and educators benefit from OSP. We conclude with concrete suggestions for astronomers and educators who wish to enhance astronomy education in their local areas.

Two Distinct Mechanisms Govern RpoS-Mediated Repression of Tick-Phase Genes during Mammalian Host Adaptation by Borrelia burgdorferi, the Lyme Disease Spirochete.

PubMed

Grove, Arianna P; Liveris, Dionysios; Iyer, Radha; Petzke, Mary; Rudman, Joseph; Caimano, Melissa J; Radolf, Justin D; Schwartz, Ira

2017-08-22

The alternative sigma factor RpoS plays a key role modulating gene expression in Borrelia burgdorferi , the Lyme disease spirochete, by transcribing mammalian host-phase genes and repressing σ 70 -dependent genes required within the arthropod vector. To identify cis regulatory elements involved in RpoS-dependent repression, we analyzed green fluorescent protein (GFP) transcriptional reporters containing portions of the upstream regions of the prototypical tick-phase genes ospAB , the glp operon, and bba74 As RpoS-mediated repression occurs only following mammalian host adaptation, strains containing the reporters were grown in dialysis membrane chambers (DMCs) implanted into the peritoneal cavities of rats. Wild-type spirochetes harboring ospAB - and glp-gfp constructs containing only the minimal (-35/-10) σ 70 promoter elements had significantly lower expression in DMCs relative to growth in vitro at 37°C; no reduction in expression occurred in a DMC-cultivated RpoS mutant harboring these constructs. In contrast, RpoS-mediated repression of bba74 required a stretch of DNA located between -165 and -82 relative to its transcriptional start site. Electrophoretic mobility shift assays employing extracts of DMC-cultivated B. burgdorferi produced a gel shift, whereas extracts from RpoS mutant spirochetes did not. Collectively, these data demonstrate that RpoS-mediated repression of tick-phase borrelial genes occurs by at least two distinct mechanisms. One (e.g., ospAB and the glp operon) involves primarily sequence elements near the core promoter, while the other (e.g., bba74 ) involves an RpoS-induced transacting repressor. Our results provide a genetic framework for further dissection of the essential "gatekeeper" role of RpoS throughout the B. burgdorferi enzootic cycle. IMPORTANCE Borrelia burgdorferi , the Lyme disease spirochete, modulates gene expression to adapt to the distinctive environments of its mammalian host and arthropod vector during its enzootic cycle. The alternative sigma factor RpoS has been referred to as a "gatekeeper" due to its central role in regulating the reciprocal expression of mammalian host- and tick-phase genes. While RpoS-dependent transcription has been studied extensively, little is known regarding the mechanism(s) of RpoS-mediated repression. We employed a combination of green fluorescent protein transcriptional reporters along with an in vivo model to define cis regulatory sequences responsible for RpoS-mediated repression of prototypical tick-phase genes. Repression of ospAB and the glp operon requires only sequences near their core promoters, whereas modulation of bba74 expression involves a putative RpoS-dependent repressor that binds upstream of the core promoter. Thus, Lyme disease spirochetes employ at least two different RpoS-dependent mechanisms to repress tick-phase genes within the mammal. Copyright © 2017 Grove et al.

Optimal sensor placement for time-domain identification using a wavelet-based genetic algorithm

NASA Astrophysics Data System (ADS)

Mahdavi, Seyed Hossein; Razak, Hashim Abdul

2016-06-01

This paper presents a wavelet-based genetic algorithm strategy for optimal sensor placement (OSP) effective for time-domain structural identification. Initially, the GA-based fitness evaluation is significantly improved by using adaptive wavelet functions. Later, a multi-species decimal GA coding system is modified to be suitable for an efficient search around the local optima. In this regard, a local operation of mutation is introduced in addition with regeneration and reintroduction operators. It is concluded that different characteristics of applied force influence the features of structural responses, and therefore the accuracy of time-domain structural identification is directly affected. Thus, the reliable OSP strategy prior to the time-domain identification will be achieved by those methods dealing with minimizing the distance of simulated responses for the entire system and condensed system considering the force effects. The numerical and experimental verification on the effectiveness of the proposed strategy demonstrates the considerably high computational performance of the proposed OSP strategy, in terms of computational cost and the accuracy of identification. It is deduced that the robustness of the proposed OSP algorithm lies in the precise and fast fitness evaluation at larger sampling rates which result in the optimum evaluation of the GA-based exploration and exploitation phases towards the global optimum solution.

Identification and molecular survey of Borrelia burgdorferi sensu lato in sika deer (Cervus nippon) from Jilin Province, north-eastern China.

PubMed

Zhai, Bintao; Niu, Qingli; Yang, Jifei; Liu, Zhijie; Liu, Junlong; Yin, Hong; Zeng, Qiaoying

2017-02-01

Lyme disease caused by Borrelia burgdorferi sensu lato (s.l.) is a common disease of domestic animals and wildlife worldwide. Sika deer is first-grade state-protected wildlife animals in China and have economic consequences for humans. It is reported that sika deer may serve as an important reservoir host for several species of B. burgdorferi s.l. and may transmit these species to humans and animals. However, little is known about the presence of Borrelia pathogens in sika deer in China. In this study, the existence and prevalence of Borrelia sp. in sika deer from four regions of Jilin Province in China was assessed. Seventy-one blood samples of sika deer were collected and tested by nested-PCRs based on 16S ribosomal RNA (16S rRNA), outer surface protein A (OspA), flagenllin (fla), and 5S-23S rRNA intergenic spacer (5S-23S rRNA) genes of B. burgdorferi s.l. Six (8.45%) samples were positive for Borrelia sp. based on sequences of 4 genes. The positive samples were detected 18 for 16S rRNA, 10 for OspA, 16 for fla and 6 for 5S-23S, with the positive rates 25.35% (95% CI=3.8-35.6), 14.08% (95% CI=3.0-21.6), 22.54% (95% CI=4.3-36.9) and 8.45% (95% CI=1.7-22.9), respectively. Sequence analysis of the positive PCR products revealed that the partial 4 genes sequences in this study were all most similar to the sequences of B. garinii and B. burgdorferi sensu stricto (s.s.), no other Borrelia genospecies were found. This is the first report of Borrelia pathogens in sika deer in China. The findings in this study indicated that sika deer as potential natural host and may spread Lyme disease pathogen to animals, ticks, and even humans. Copyright © 2016. Published by Elsevier B.V.

An Evaluation of the Effect of D.C.'s Voucher Program on Public School Achievement and Racial Integration after One Year. Education Working Paper No. 10.

ERIC Educational Resources Information Center

Greene, Jay P.; Winters, Marcus A.

2006-01-01

This study evaluates the initial effect of Washington, D.C.'s Opportunity Scholarship Program (OSP) on the academic performance of public schools and its effects on the opportunities that District students have to attend integrated schools. The OSP is a federally sponsored school voucher program that provides vouchers worth up to $7,500 for an…

Evaluation of the DC Opportunity Scholarship Program: An Early Look at Applicants and Participating Schools under the SOAR Act. Year 1 Report. NCEE 2015-4000

ERIC Educational Resources Information Center

Feldman, Jill; Lucas-McLean, Juanita; Gutmann, Babette; Dynarski, Mark; Betts, Julian

2014-01-01

This report explores implementation of the District of Columbia Opportunity Scholarship Program (OSP) in the first two years after Congress reauthorized it with some changes under the SOAR Act of 2011. Key findings include the following: (1) Just over half of all DC private schools participated in the OSP, with current schools more likely to have…

Evaluation of the DC Opportunity Scholarship Program: An Early Look at Applicants and Participating Schools under the SOAR Act. Year 1 Report, Executive Summary. NCEE 2015-4000

ERIC Educational Resources Information Center

Feldman, Jill; Lucas-McLean, Juanita; Gutmann, Babette; Dynarski, Mark; Betts, Julian

2014-01-01

This report explores implementation of the District of Columbia Opportunity Scholarship Program (OSP) in the first two years after Congress reauthorized it with some changes under the SOAR Act of 2011. Key findings include the following: (1) Just over half of all DC private schools participated in the OSP, with current schools more likely to have…

Safety of oral sulfates in rats and dogs contrasted with phosphate-induced nephropathy in rats.

PubMed

Pelham, Russell W; Russell, Robert G; Padgett, Eric L; Reno, Frederick E; Cleveland, Mark vB

2009-01-01

An oral sulfate salt solution (OSS), under development as a bowel cleansing agent for colonoscopy in humans, is studied in rats and dogs. In rats, amaximumpractical oral OSS dose (5 g/kg/d) is compared with an oral sodium phosphate (OSP) solution, both at about 7 times the clinical dose. OSS induces the intended effects of loose stools and diarrhea. In rats, higher urine sodium and potassium accompany higher clearance rates, considered adaptive to the osmotic load of OSS. OSS for 28 days is well tolerated in rats and dogs. In contrast, OSP causes increased mortality, reduced body weight and food consumption, severe kidney tubular degeneration, and calcium phosphate deposition in rats. These are accompanied by mineralization in the stomach and aorta, along with cardiac and hepatic degeneration and necrosis. The greater safety margin of OSS over OSP at similarmultiples of the clinical dose indicates its suitability for human use.

Vaccination against Lyme disease: Are we ready for it?

PubMed

Kaaijk, Patricia; Luytjes, Willem

2016-03-03

Lyme disease is the most common tick-borne illness in the Northern hemisphere and is caused by spirochetes of the Borrelia burgdorferi sensu lato complex. A first sign of Borrelia infection is a circular skin rash, erythema migrans, but it can develop to more serious manifestations affecting skin, nervous system, joints, and/or heart. The marked increase in Lyme disease incidence over the past decades, the severity of the disease, and the associated high medical costs of, in particular, the persistent forms of Lyme disease requires adequate measures for control. Vaccination would be the most effective intervention for prevention, but at present no vaccine is available. In the 1990s, 2 vaccines against Lyme disease based on the OspA protein from the predominant Borrelia species of the US showed to be safe and effective in clinical phase III studies. However, failed public acceptance led to the demise of these monovalent OspA-based vaccines. Nowadays, public seem to be more aware of the serious health problems that Lyme disease can cause and seem more ready for the use of a broadly protective vaccine. This article discusses several aspects that should be considered to enable the development and implementation of a vaccine to prevent Lyme disease successfully.

Vaccination against Lyme disease: Are we ready for it?

PubMed Central

Kaaijk, Patricia; Luytjes, Willem

2016-01-01

Abstract Lyme disease is the most common tick-borne illness in the Northern hemisphere and is caused by spirochetes of the Borrelia burgdorferi sensu lato complex. A first sign of Borrelia infection is a circular skin rash, erythema migrans, but it can develop to more serious manifestations affecting skin, nervous system, joints, and/or heart. The marked increase in Lyme disease incidence over the past decades, the severity of the disease, and the associated high medical costs of, in particular, the persistent forms of Lyme disease requires adequate measures for control. Vaccination would be the most effective intervention for prevention, but at present no vaccine is available. In the 1990s, 2 vaccines against Lyme disease based on the OspA protein from the predominant Borrelia species of the US showed to be safe and effective in clinical phase III studies. However, failed public acceptance led to the demise of these monovalent OspA-based vaccines. Nowadays, public seem to be more aware of the serious health problems that Lyme disease can cause and seem more ready for the use of a broadly protective vaccine. This article discusses several aspects that should be considered to enable the development and implementation of a vaccine to prevent Lyme disease successfully. PMID:26337648

Repurposing Ospemifene for Potentiating an Antigen-Specific Immune Response

PubMed Central

Kao, Chiao-Jung; Wurz, Gregory T.; Lin, Yi-Chen; Vang, Daniel P.; Phong, Brian; DeGregorio, Michael W.

2016-01-01

Objective Ospemifene, an estrogen receptor agonist/antagonist approved for treatment of dyspareunia and vaginal dryness in postmenopausal women, has potential new indications as an immune modulator. The overall objective of the present series of preclinical studies was to evaluate the immunomodulatory activity of ospemifene in combination with a peptide cancer vaccine. Methods Immune regulating effects, mechanism of action and structure activity relationships of ospemifene and related compounds were evaluated by examining expression of T cell activating cytokines in vitro, and antigen-specific immune response and cytotoxic T-lymphocyte activity in vivo. The effects of ospemifene (OSP) on the immune response to a peptide cancer vaccine (PV) were evaluated following chronic [control (n=22); OSP 50 mg/kg (n=16); PV (n=6); OSP+PV (n=11)], intermittent [control (n=10); OSP 10 and 50 mg/kg (n=11); PV (n=11); combination treatment (n=11 each dose)] and pretreatment [control; OSP 100 mg/kg; PV 100 µg; combination treatment (n=8 all groups)] ospemifene oral dosing schedules in a total of 317 mixed-sex tumor-bearing and non-tumor-bearing mice. Results The results showed that ospemifene induced expression of the key TH1 cytokines interferon gamma and interleukin-2 in vitro, which may be mediated by stimulating T cells through phosphoinositide 3-kinase and calmodulin signaling pathways. In combination with an antigen-specific peptide cancer vaccine, ospemifene increased antigen-specific immune response and increased cytotoxic T-lymphocyte activity in tumor-bearing and non-tumor-bearing mice. The pretreatment, intermittent, and chronic dosing schedules of ospemifene activate naïve T cells, modulate antigen-induced tolerance and reduce tumor-associated, pro-inflammatory cytokines, respectively. Conclusions Taken together, ospemifene’s dose response and schedule-dependent immune modulating activity offers a method of tailoring and augmenting the efficacy of previously failed antigen-specific cancer vaccines for a wide range of malignancies. PMID:27922937

Occurrence of Borrelia burgdorferi s.l. in different genera of mosquitoes (Culicidae) in Central Europe.

PubMed

Melaun, Christian; Zotzmann, Sina; Santaella, Vanesa Garcia; Werblow, Antje; Zumkowski-Xylander, Helga; Kraiczy, Peter; Klimpel, Sven

2016-03-01

Lyme disease or Lyme borreliosis is a vector-borne infectious disease caused by spirochetes of the Borrelia burgdorferi sensu lato complex. Some stages of the borrelial transmission cycle in ticks (transstadial, feeding and co-feeding) can potentially occur also in insects, particularly in mosquitoes. In the present study, adult as well as larval mosquitoes were collected at 42 different geographical locations throughout Germany. This is the first study, in which German mosquitoes were analyzed for the presence of Borrelia spp. Targeting two specific borrelial genes, flaB and ospA encoding for the subunit B of flagellin and the outer surface protein A, the results show that DNA of Borrelia afzelii, Borrelia bavariensis and Borrelia garinii could be detected in ten Culicidae species comprising four distinct genera (Aedes, Culiseta, Culex, and Ochlerotatus). Positive samples also include adult specimens raised in the laboratory from wild-caught larvae indicating that transstadial and/or transovarial transmission might occur within a given mosquito population. Copyright © 2015 Elsevier GmbH. All rights reserved.

Traverse Planning Experiments for Future Planetary Surface Exploration

NASA Technical Reports Server (NTRS)

Hoffman, Stephen J.; Voels, Stephen A.; Mueller, Robert P.; Lee, Pascal C.

2012-01-01

The purpose of the investigation is to evaluate methodology and data requirements for remotely-assisted robotic traverse of extraterrestrial planetary surface to support human exploration program, assess opportunities for in-transit science operations, and validate landing site survey and selection techniques during planetary surface exploration mission analog demonstration at Haughton Crater on Devon Island, Nunavut, Canada. Additionally, 1) identify quality of remote observation data sets (i.e., surface imagery from orbit) required for effective pre-traverse route planning and determine if surface level data (i.e., onboard robotic imagery or other sensor data) is required for a successful traverse, and if additional surface level data can improve traverse efficiency or probability of success (TRPF Experiment). 2) Evaluate feasibility and techniques for conducting opportunistic science investigations during this type of traverse. (OSP Experiment). 3) Assess utility of remotely-assisted robotic vehicle for landing site validation survey. (LSV Experiment).

Fifty Years of Ocean Observations in the Pacific Northeast

NASA Astrophysics Data System (ADS)

Whitney, Frank; Tortell, Philippe

2006-12-01

Ocean Station Papa, at 50°N, 145°W in the Alaska Gyre (Figure 1), started as a weather station in the 1940s. In 1956, oceanographers began collecting a suite of standard measurements from the cool subarctic waters at Ocean Station Papa (OSP), including temperature, salinity, oxygen, and plankton. Three years later, a series of sampling stations was added along the 1400-kilometer `Line P' from the Canadian coast to OSP, to aid in understanding ocean variability.

Graphs and Tracks Revisited

NASA Astrophysics Data System (ADS)

Christian, Wolfgang; Belloni, Mario

2013-04-01

We have recently developed a Graphs and Tracks model based on an earlier program by David Trowbridge, as shown in Fig. 1. Our model can show position, velocity, acceleration, and energy graphs and can be used for motion-to-graphs exercises. Users set the heights of the track segments, and the model displays the motion of the ball on the track together with position, velocity, and acceleration graphs. This ready-to-run model is available in the ComPADRE OSP Collection at www.compadre.org/osp/items/detail.cfm?ID=12023.

One-Year Duration of Immunity Induced by Vaccination with a Canine Lyme Disease Bacterin▿

PubMed Central

LaFleur, Rhonda L.; Callister, Steven M.; Dant, Jennifer C.; Jobe, Dean A.; Lovrich, Steven D.; Warner, Thomas F.; Wasmoen, Terri L.; Schell, Ronald F.

2010-01-01

Laboratory-reared beagles were vaccinated with a placebo or a bacterin comprised of Borrelia burgdorferi S-1-10 and ospA-negative/ospB-negative B. burgdorferi 50772 and challenged after 1 year with B. burgdorferi-infected Ixodes scapularis ticks. For the placebo recipients, spirochetes were recovered from 9 (60%) skin biopsy specimens collected after 1 month, and the organisms persisted in the skin thereafter. Ten (67%) dogs also developed joint infection (3 dogs), lameness or synovitis (7 dogs), or B. burgdorferi-specific antibodies (8 dogs). For the vaccine recipients, spirochetes were recovered from 6 (40%) skin biopsy specimens collected after 1 month. However, subsequent biopsy specimens were negative, and the dogs failed to develop joint infection (P = 0.224), lameness/synovitis (P = 0.006), or Lyme disease-specific antibody responses (P = 0.002). The bacterin provided a high level of protection for 1 year after immunization, and the addition of the OspC-producing B. burgdorferi 50772 provided enhanced protection. PMID:20237200

Plastidic phosphoglucomutase and ADP-glucose pyrophosphorylase mutants impair starch synthesis in rice pollen grains and cause male sterility

PubMed Central

Lee, Sang-Kyu; Eom, Joon-Seob; Hwang, Seon-Kap; Shin, Dongjin; An, Gynheung; Okita, Thomas W.; Jeon, Jong-Seong

2016-01-01

To elucidate the starch synthesis pathway and the role of this reserve in rice pollen, we characterized mutations in the plastidic phosphoglucomutase, OspPGM, and the plastidic large subunit of ADP-glucose (ADP-Glc) pyrophosphorylase, OsAGPL4. Both genes were up-regulated in maturing pollen, a stage when starch begins to accumulate. Progeny analysis of self-pollinated heterozygous lines carrying the OspPGM mutant alleles, osppgm-1 and osppgm-2, or the OsAGPL4 mutant allele, osagpl4-1, as well as reciprocal crosses between the wild type (WT) and heterozygotes revealed that loss of OspPGM or OsAGPL4 caused male sterility, with the former condition rescued by the introduction of the WT OspPGM gene. While iodine staining and transmission electron microscopy analyses of pollen grains from homozygous osppgm-1 lines produced by anther culture confirmed the starch null phenotype, pollen from homozygous osagpl4 mutant lines, osagpl4-2 and osagpl4-3, generated by the CRISPR/Cas system, accumulated small amounts of starch which were sufficient to produce viable seed. Such osagpl4 mutant pollen, however, was unable to compete against WT pollen successfully, validating the important role of this reserve in fertilization. Our results demonstrate that starch is mainly polymerized from ADP-Glc synthesized from plastidic hexose phosphates in rice pollen and that starch is an essential requirement for successful fertilization in rice. PMID:27588462

Injuries in children with extra physical education in primary schools.

PubMed

Rexen, Christina Trifonov; Andersen, Lars Bo; Ersbøll, Annette Kjær; Jespersen, Eva; Franz, Claudia; Wedderkopp, Niels

2014-04-01

(1) Examine the influence of extra physical education (EPE) on the number of musculoskeletal injuries in public schools accounting for organized sports participation (OSP) outside school. (2) Examine the major injury subgroup: growth-related overuse (GRO) through the overuse-related injury group. A longitudinal controlled school-based study among Danish public schools. At baseline, 1216 children participated age 6.2-12.4 yr. Six schools (701 children) with EPE and four control schools (515 children) were followed up with weekly automated mobile phone text messages for information on musculoskeletal problems and OSP. Health care personnel diagnosed the children according to the World Health Organization's International Classification of Diseases, Tenth Revision. Data were analyzed using a two-part zero-inflated negative binomial (ZINB) regression model. School type had no influence on the odds of sustaining an injury but increased the probability of sustaining a higher injury count for children with injuries, with total injuries by a factor of 1.29 (95% confidence interval [CI] = 1.07-1.56), overuse by a factor of 1.29 (95% CI = 1.06-1.55), and GRO by a factor of 1.38 (95% CI = 1.02-1.80). Weekly mean OSP decreased the odds of belonging to the group of children with no injuries, by a factor of 0.29 (95% CI = 0.14-0.58), 0.26 (95% CI = 0.14-0.48), and 0.17 (95% CI = 0.06-0.52) for total, overuse, and GRO, respectively. OSP also increased the probability of sustaining a higher injury count for children with injuries by a factor of 1.11 (95% CI = 1.02-1.22), 1.10 (95% CI = 1.00-1.22), and 1.14 (95% CI = 1.00-1.30), respectively. Children enrolled in EPE schools with high OSP have the highest odds of injury and a high probability of sustaining a higher injury count compared to their peers at schools with normal PE. Special attention should be assigned to these children during compulsory PE.

Kidney injury after sodium phosphate solution beyond the acute renal failure.

PubMed

Fernández-Juárez, Gema; Parejo, Leticia; Villacorta, Javier; Tato, Ana; Cazar, Ramiro; Guerrero, Carmen; Marin, Isabel Martinez; Ocaña, Javier; Mendez-Abreu, Angel; López, Katia; Gruss, Enrique; Gallego, Eduardo

2016-01-01

Screening colonoscopy with polipectomy reduces colonorectal cancer incidence and mortality. An adequate bowel cleansing is one of the keys to achieving best results with this technique. Oral sodium phosphate solution (OSP) had a widespread use in the 90s decade. Its efficacy was similar to polyethylene glycol (PEG) solution, but with less cost and convenient administration. Series of patients with acute renal failure due to OSP use have been reported. However, large cohorts of patients found no difference in the incidence of renal damage between these two solutions. From 2006 to 2009 we identified twelve cases of phosphate nephropathy after colonoscopy prepared with OSP. All patients were followed up to six months. All patients had received just a single dose. We analyzed 12 cases with phosphate nephropathy; three patients debuted with AKI and nine patients had chronic renal injury. Four cases were confirmed with renal biopsy. One patient with AKI needed hemodialysis at diagnosis without subsequent recovery. Two patients (both with chronic damage) fully recovered their previous renal function. The remaining patients (nine) had an average loss of estimated glomerular filtration rate of 24ml/min/1.73m(2). The use of OSP can lead to both acute and chronic renal damage. However, chronic injury was the most common pattern. Both forms of presentation imply a significant and irreversible loss of renal function. Further studies analyzing renal damage secondary to bowel cleaning should consider these two different patterns of injury. Copyright © 2016 Sociedad Española de Nefrología. Published by Elsevier España, S.L.U. All rights reserved.

Vaccination with Shigella flexneri 2a conjugate induces type 2a and cross-reactive type 6 antibodies in humans but not in mice.

PubMed

Farzam, Nahid; Ramon-Saraf, Reut; Banet-Levi, Yonit; Lerner-Geva, Liat; Ashkenazi, Shai; Kubler-Kielb, Joanna; Vinogradov, Evgeny; Robbins, John B; Schneerson, Rachel

2017-09-05

Shigella flexneri (S. flexneri) 6 has emerged as an important cause of shigellosis. Our efficacy study of Shigella sonnei and S. flexneri 2a O-specific polysaccharide (O-SP) conjugates in 1-4year-olds had too few S. flexneri 2a cases for efficacy evaluation but surprisingly showed protection of 3-4year-olds, S. flexneri 2a-recipients, from S. flexneri 6 infection. To investigate this cross-protection antibodies to both Shigella types were investigated in all sera remaining from previous studies. Twenty to 30% of 3-44year-old humans injected with S. flexneri 2a conjugate responded with ≥4-fold increases of IgG anti type 6, p<0.00001. The specificity of these antibodies was shown by inhibition studies. S. flexneri 6 infection of 2 children induced besides S. flexneri 6, also S. flexneri 2a antibodies, at levels of S. flexneri 2a vaccinees. S. flexneri 2a antibodies induced by S. flexneri 6 conjugates could not be studied since no such conjugate was assessed in humans and mice responded almost exclusively to the O-SP of the injected conjugate, with no cross-reactive antibodies. Our results indicate induction of cross-reactive protective antibodies. The O-acetylated disaccharide shared by S. flexneri 6 and 2a O-SPs, is the likely basis for their cross-reactivity. S. flexneri 6 O-SP conjugates, alone and in combination with S. flexneri 2a, merit further investigation for broad S. flexneri protection. Published by Elsevier Ltd.

The Relationship between Parameters of Long-Latency Evoked Potentials in a Multisensory Design.

PubMed

Hernández, Oscar H; García-Martínez, Rolando; Monteón, Victor

2016-10-01

In previous papers, we have shown that parameters of the omitted stimulus potential (OSP), which occurs at the end of a train of sensory stimuli, strongly depend on the modality. A train of stimuli also produces long-latency evoked potentials (LLEP) at the beginning of the train. This study is an extension of the OSP research, and it tested the relationship between parameters (ie, rate of rise, amplitude, and peak latency) of the P2 waves when trains of auditory, visual, or somatosensory stimuli were applied. The dynamics of the first 3 potentials in the train, related to habituation, were also studied. Twenty healthy young college volunteers participated in the study. As in the OSP, the P2 was faster and higher for auditory than for visual or somatosensory stimuli. The first P2 was swifter and higher than the second and the third potentials. The strength of habituation depends on the sensory modality and the parameter used. All these findings support the view that many long-latency brain potentials could share neural mechanisms related to wave generation. © EEG and Clinical Neuroscience Society (ECNS) 2015.

Hyperphosphatemia after sodium phosphate laxatives in low risk patients: Prospective study

PubMed Central

Casais, Marcela Noemi; Rosa-Diez, Guillermo; Pérez, Susana; Mansilla, Elina Noemi; Bravo, Susana; Bonofiglio, Francisco Carlos

2009-01-01

AIM: To establish the frequency of hyperphosphatemia following the administration of sodium phosphate laxatives in low-risk patients. METHODS: One hundred consecutive ASA I-II individuals aged 35-74 years, who were undergoing colonic cleansing with oral sodium phosphate (OSP) before colonoscopy were recruited for this prospective study. Exclusion criteria: congestive heart failure, chronic kidney disease, diabetes, liver cirrhosis, intestinal obstruction, decreased bowel motility, increased bowel permeability, and hyperparathyroidism. The day before colonoscopy, all the participants entered a 24-h period of diet that consisted of 4 L of clear fluids with sugar or honey and 90 mL (60 g) of OSP in two 45-mL doses, 5 h apart. Serum phosphate was measured before and after the administration of the laxative. RESULTS: The main demographic data (mean ± SD) were: age, 58.9 ± 8.4 years; height, 163.8 ± 8.6 cm; weight, 71 ± 13 kg; body mass index, 26 ± 4; women, 66%. Serum phosphate increased from 3.74 ± 0.56 to 5.58 ± 1.1 mg/dL, which surpassed the normal value (2.5-4.5 mg/dL) in 87% of the patients. The highest serum phosphate was 9.6 mg/dL. Urea and creatinine remained within normal limits. Post-treatment OSP serum phosphate concentration correlated inversely with glomerular filtration rate (P < 0.007, R2 = 0.0755), total body water (P < 0.001, R2 = 0.156) and weight (P < 0.013, R2 = 0.0635). CONCLUSION: In low-risk, well-hydrated patients, the standard dose of OSP-laxative-induced hyperphosphatemia is related to body weight. PMID:20014460

Outcomes of minimally invasive simple prostatectomy for benign prostatic hyperplasia: a systematic review and meta-analysis.

PubMed

Lucca, Ilaria; Shariat, Shahrokh F; Hofbauer, Sebastian L; Klatte, Tobias

2015-04-01

(1) To assess the outcomes of minimally invasive simple prostatectomy (MISP) for the treatment of symptomatic benign prostatic hyperplasia in men with large prostates and (2) to compare them with open simple prostatectomy (OSP). A systematic review of outcomes of MISP for benign prostatic hyperplasia with meta-analysis was conducted. The article selection process was conducted according to the PRISMA guidelines. Twenty-seven observational studies with 764 patients were analyzed. The mean prostate volume was 113.5 ml (95 % CI 106-121). The mean increase in Qmax was 14.3 ml/s (95 % CI 13.1-15.6), and the mean improvement in IPSS was 17.2 (95 % CI 15.2-19.2). Mean duration of operation was 141 min (95 % CI 124-159), and the mean intraoperative blood loss was 284 ml (95 % CI 243-325). One hundred and four patients (13.6 %) developed a surgical complication. In comparative studies, length of hospital stay (WMD -1.6 days, p = 0.02), length of catheter use (WMD -1.3 days, p = 0.04) and estimated blood loss (WMD -187 ml, p = 0.015) were significantly lower in the MISP group, while the duration of operation was longer than in OSP (WMD 37.8 min, p < 0.0001). There were no differences in improvements in Qmax, IPSS and perioperative complications between both procedures. The small study sizes, publication bias, lack of systematic complication reporting and short follow-up are limitations. MISP seems an effective and safe treatment option. It provides similar improvements in Qmax and IPSS as OSP. Despite taking longer, it results in less blood loss and shorter hospital stay. Prospective randomized studies comparing OSP, MISP and laser enucleation are needed to define the standard surgical treatment for large prostates.

Specialty-specific multi-source feedback: assuring validity, informing training.

PubMed

Davies, Helena; Archer, Julian; Bateman, Adrian; Dewar, Sandra; Crossley, Jim; Grant, Janet; Southgate, Lesley

2008-10-01

The white paper 'Trust, Assurance and Safety: the Regulation of Health Professionals in the 21st Century' proposes a single, generic multi-source feedback (MSF) instrument in the UK. Multi-source feedback was proposed as part of the assessment programme for Year 1 specialty training in histopathology. An existing instrument was modified following blueprinting against the histopathology curriculum to establish content validity. Trainees were also assessed using an objective structured practical examination (OSPE). Factor analysis and correlation between trainees' OSPE performance and the MSF were used to explore validity. All 92 trainees participated and the assessor response rate was 93%. Reliability was acceptable with eight assessors (95% confidence interval 0.38). Factor analysis revealed two factors: 'generic' and 'histopathology'. Pearson correlation of MSF scores with OSPE performances was 0.48 (P = 0.001) and the histopathology factor correlated more highly (histopathology r = 0.54, generic r = 0.42; t = - 2.76, d.f. = 89, P < 0.01). Trainees scored least highly in relation to ability to use histopathology to solve clinical problems (mean = 4.39) and provision of good reports (mean = 4.39). Three of six doctors whose means were < 4.0 received free text comments about report writing. There were 83 forms with aggregate scores of < 4. Of these, 19.2% included comments about report writing. Specialty-specific MSF is feasible and achieves satisfactory reliability. The higher correlation of the 'histopathology' factor with the OSPE supports validity. This paper highlights the importance of validating an MSF instrument within the specialty-specific context as, in addition to assuring content validity, the PATH-SPRAT (Histopathology-Sheffield Peer Review Assessment Tool) also demonstrates the potential to inform training as part of a quality improvement model.

Why is There Still no Human Vaccine Against Lyme Borreliosis?

PubMed

Skotarczak, Bogumiła

2015-01-01

Lyme disease, transmitted by ticks, is a complex illness that can be difficult to diagnose but easy to treat in most early cases, yet difficult in its latest stage. Every year, infections with Borrelia burgdorferi sensu lato spirochetes cause thousands of new cases of illness around the world, including people with a normal immunological reaction. Prevention in the form of vaccines is difficult due to e.g. very high variability of Borrelia antigen proteins, which precludes the construction of an effective vaccine. After the withdrawal of the OspA vaccine (LYMErix) in the USA, despite promising results, no vaccine protecting humans against all pathogenic species from the B. burgdorferi s.l. group is available. Recent data indicate that an effective vaccine may require a combination of several antigens or multiple epitopes based on vector-borne proteins and several outer membrane proteins of Borrelia. With the discontinuance of Lyme vaccines, personal protective behavior and the avoidance of exposure in high-risk areas remain necessary resources of prevention.

Ligand-free palladium-mediated site-specific protein labeling inside gram-negative bacterial pathogens.

PubMed

Li, Jie; Lin, Shixian; Wang, Jie; Jia, Shang; Yang, Maiyun; Hao, Ziyang; Zhang, Xiaoyu; Chen, Peng R

2013-05-15

Palladium, a key transition metal in advancing modern organic synthesis, mediates diverse chemical conversions including many carbon-carbon bond formation reactions between organic compounds. However, expanding palladium chemistry for conjugation of biomolecules such as proteins, particularly within their native cellular context, is still in its infancy. Here we report the site-specific protein labeling inside pathogenic Gram-negative bacterial cells via a ligand-free palladium-mediated cross-coupling reaction. Two rationally designed pyrrolysine analogues bearing an aliphatic alkyne or an iodophenyl handle were first encoded in different enteric bacteria, which offered two facial handles for palladium-mediated Sonogashira coupling reaction on proteins within these pathogens. A GFP-based bioorthogonal reaction screening system was then developed, allowing evaluation of both the efficiency and the biocompatibilty of various palladium reagents in promoting protein-small molecule conjugation. The identified simple compound-Pd(NO3)2 exhibited high efficiency and biocompatibility for site-specific labeling of proteins in vitro and inside living E. coli cells. This Pd-mediated protein coupling method was further utilized to label and visualize a Type-III Secretion (T3S) toxin-OspF in Shigella cells. Our strategy may be generally applicable for imaging and tracking various virulence proteins within Gram-negative bacterial pathogens.

Orbital Space Plane (OSP) Program at Lockheed Martin

NASA Technical Reports Server (NTRS)

Ford, Robert

2003-01-01

Lockheed Martin has been an active participant in NASA's Space Launch Initiative (SLI) programs over the past several years. SLI, part of NASA's Integrated Space Transportation Plan (ISTP), was restructured in November 2002 to focus the overall theme of safer, more affordable space transportation along two paths the Orbital Space Plane (OSP) and the Next Generation Launch Technology programs. The Orbital Space Plane program has the goal of providing rescue capability from the International Space Station by 2008 or earlier and transfer capability for crew (and contingency cargo) by 2012. The Next Generation Launch Technology program is combining research and development efforts from the 2d Generation Reusable Launch Vehicle (2GRLV) program with cutting-edge, advanced space transportation programs (previously designated 31d Generation) into one program aimed at enabling safe, reliable, cost-effective reusable launch systems by the middle of the next decade. Lockheed Martin is one of three prime contractors working to bring Orbital Space Plane system concepts to a system design level of maturity by December 2003. This paper and presentation will update the aerospace community on the progress of the OSP program, from an industry perspective, and provide insights into Lockheed Martin's role in enabling the vision of a safer, more affordable means of taking people to and from space.

A formulation of a matrix sparsity approach for the quantum ordered search algorithm

NASA Astrophysics Data System (ADS)

Parmar, Jupinder; Rahman, Saarim; Thiara, Jaskaran

One specific subset of quantum algorithms is Grovers Ordered Search Problem (OSP), the quantum counterpart of the classical binary search algorithm, which utilizes oracle functions to produce a specified value within an ordered database. Classically, the optimal algorithm is known to have a log2N complexity; however, Grovers algorithm has been found to have an optimal complexity between the lower bound of ((lnN-1)/π≈0.221log2N) and the upper bound of 0.433log2N. We sought to lower the known upper bound of the OSP. With Farhi et al. MITCTP 2815 (1999), arXiv:quant-ph/9901059], we see that the OSP can be resolved into a translational invariant algorithm to create quantum query algorithm restraints. With these restraints, one can find Laurent polynomials for various k — queries — and N — database sizes — thus finding larger recursive sets to solve the OSP and effectively reducing the upper bound. These polynomials are found to be convex functions, allowing one to make use of convex optimization to find an improvement on the known bounds. According to Childs et al. [Phys. Rev. A 75 (2007) 032335], semidefinite programming, a subset of convex optimization, can solve the particular problem represented by the constraints. We were able to implement a program abiding to their formulation of a semidefinite program (SDP), leading us to find that it takes an immense amount of storage and time to compute. To combat this setback, we then formulated an approach to improve results of the SDP using matrix sparsity. Through the development of this approach, along with an implementation of a rudimentary solver, we demonstrate how matrix sparsity reduces the amount of time and storage required to compute the SDP — overall ensuring further improvements will likely be made to reach the theorized lower bound.

Studies of short-range tungsten migration in DIII-D divertor

NASA Astrophysics Data System (ADS)

Rudakov, D. L.; Stangeby, P. C.; Elder, J. D.; Ding, R.; Abrams, T.; Unterberg, E. A.; Briesemeister, A.; Donovan, D.; McLean, A. G.; Guo, H. Y.; Thomas, D. M.; Hinson, E.; Wampler, W. R.; Watkins, J. G.

2016-10-01

Two toroidal rings of 5 cm wide W-coated TZM inserts were installed in the lower divertor of DIII-D. Migration of W on the graphite tile surfaces 1-6 cm radially outwards from the outermost ring was studied in a series of 23 reproducible lower single null L-mode discharges with the Outer Strike Point (OSP) placed on the ring. The discharges used 3.2 MW of NBI heating power; plasma density and electron temperature at the OSP were about 1x1020m-3 and 30 eV. W gross erosion rates were measured via monitoring 400.9 nm WI line and applying S/XB coefficient. W deposition was measured on a graphite DiMES sample used as a divertor collector probe. The sample featured two 1 mm wide radial inserts; one was exposed for the whole experiment, the other was exchanged every 4-8 plasma discharges. Measurements of the areal density of W on the inserts by post-mortem RBS analysis show that W deposition is largest in the area of net carbon deposition, possibly due to W re-erosion suppression by C deposits. Measured W coverage in the area of net C erosion is comparable to ERO modeling predictions. Supported by US DOE under DE-FG02-07ER54917, DE-AC04-94AL85000, DE-AC05-00OR22725, DE-AC52-07NA27344, DE-FC02-04ER54698.

Reflection K-matrices for a nineteen vertex model with Uq [ osp (2 | 2) (2) ] symmetry

NASA Astrophysics Data System (ADS)

Vieira, R. S.; Lima Santos, A.

2017-09-01

We derive the solutions of the boundary Yang-Baxter equation associated with a supersymmetric nineteen vertex model constructed from the three-dimensional representation of the twisted quantum affine Lie superalgebra Uq [ osp (2 | 2) (2) ]. We found three classes of solutions. The type I solution is characterized by three boundary free-parameters and all elements of the corresponding reflection K-matrix are different from zero. In the type II solution, the reflection K-matrix is even (every element of the K-matrix with an odd parity is null) and it has only one boundary free-parameter. Finally, the type III solution corresponds to a diagonal reflection K-matrix with two boundary free-parameters.

An admissible level \\widehat{osp} ( 1 \\big \\vert 2 ) -model: modular transformations and the Verlinde formula

NASA Astrophysics Data System (ADS)

Snadden, John; Ridout, David; Wood, Simon

2018-05-01

The modular properties of the simple vertex operator superalgebra associated with the affine Kac-Moody superalgebra \\widehat{{osp}} (1|2) at level -5/4 are investigated. After classifying the relaxed highest-weight modules over this vertex operator superalgebra, the characters and supercharacters of the simple weight modules are computed and their modular transforms are determined. This leads to a complete list of the Grothendieck fusion rules by way of a continuous superalgebraic analog of the Verlinde formula. All Grothendieck fusion coefficients are observed to be non-negative integers. These results indicate that the extension to general admissible levels will follow using the same methodology once the classification of relaxed highest-weight modules is completed.

Characterizing Tungsten Sourcing and SOL Transport during the Metal Rings Campaign

NASA Astrophysics Data System (ADS)

Thomas, D. M.; Abrams, T.; Unterberg, E. A.; Donovan, D.; Elder, J. D.; Wampler, W. R.; DIII-D Team

2017-10-01

The Metal Rings Campaign on DIII-D utilized two isotopically and poloidally distinct toroidal arrays of tungsten coated inserts in the lower divertor to study W divertor erosion near the outer strike point (OSP) and divertor entrance and subsequent migration in a mixed-material (C-W) environment. In AT hybrid discharges (PAUX = 14 MW, H98 = 1.6, βN = 3.7) with rapid ELMs (fELM 200 Hz, δW/W 0.7%) W impurities are seen to reach the midplane predominantly from the OSP region rather than the divertor entrance (far-SOL). Conversely, in scenarios with less frequent larger ELMs (fELM 60 Hz, δW/W 3.6%), the W impurities are found to transport equally from the OSP and entrance region. ELM-resolved spectroscopic measurements of W sourcing indicate that large ELMs can source W at many times the inter ELM rate. The peak W erosion rate can shift radially outwards consistent with the ELM energy flux, thereby shifting the balance between strikepoint and far-SOL sources. Changes in the peak erosion locations between forward and reversed Bt discharges are consistent with ExB ion drift effects. Evidence for a near-SOL impurity buildup between the divertors driven by the parallel grad-Ti force is also seen. Work supported under USDOE Cooperative Agreement DE-FC02-04ER54698.

Human Granulocytic Ehrlichiosis Agent Infection in a Pony Vaccinated with a Borrelia burgdorferi Recombinant OspA Vaccine and Challenged by Exposure to Naturally Infected Ticks

PubMed Central

Chang, Yung-Fu; McDonough, Sean P.; Chang, Chao-Fu; Shin, Kwang-Soon; Yen, William; Divers, Thomas

2000-01-01

A pony was vaccinated with recombinant OspA vaccine (rOspA) and then exposed 3 months later to Borrelia burgdorferi-infected ticks (Ixodes scapularis) collected in Westchester County, N.Y. At 2 weeks after tick exposure, the pony developed a high fever (105°F). Buffy coat smears showed that 20% of neutrophils contained ehrlichial inclusion bodies (morulae). Flunixin Meglumine (1 g daily) was given for 2 days, and the body temperature returned to normal. PCR for ehrlichial DNA was performed on blood samples for 10 consecutive days beginning when the pony was first febrile. This pony was monitored for another 3.5 months but developed no further clinical signs. The 44-kDa immunodominant human granulocytic ehrlichiosis antigen gene was amplified by PCR and cloned into a pCR2.1 vector. DNA sequence analysis of this gene showed it was only 8 bp different (99% identity) from the results reported by others (J.W. Ijdo et al., Infect. Immun. 66:3264–3269, 1998). Western blot analysis, growth inhibition assays, and repeated attempts to isolate B. burgdorferi all demonstrated the pony was protected against B. burgdorferi infection. These results highlight the potential for ticks to harbor and transmit several pathogens simultaneously, which further complicates the diagnosis and vaccination of these emerging tick-borne diseases. PMID:10618280

Impact of movie-based simulation training, with or without conventional verbal demonstration on observed OSPE scores in medical undergraduates: a double control study.

PubMed

Malik, Samina; Zaheer, Reema; Bilal, Muhammad

2013-01-01

Movie-based simulation training may be useful in delivering the preclinical observed OSPE curriculum, minimising the need of subjects/patients: however, a double-control trial needs to be performed and optimal timing and duration of training is yet to be defined. Likewise, gender-based response and students' feedback has to be assessed. The objective of this study was to compare the movie-based and traditional verbal demonstration teaching methodologies. Second-year medical undergraduates (n = 90) of Avicenna Medical College were randomised to movie-based simulation training (group B, n = 30), traditional verbal training alone (group C, n=30), and a combination (group A, n = 30). The scores were marked by observers using a standardised key and were compared for performance at 2 observed OSPE stations. Group B and A performed significantly better than group C on station 1 and 2. Gender factor did not seem to influence the score. A total of 99% students reported that combination of the 2 teaching modes is the best option. They believed it offers more clear understanding with interest (61%), long term memory (21%), use of both senses; seeing & hearing (10%) & better focus of attention (3%). Even half an hour of movie-based simulation training with traditional instructor-based training may improve student performance significantly, and the students prefer a combination of the both.

Molecular and Pathogenic Characterization of Borrelia burgdorferi Sensu Lato Isolates from Spain

PubMed Central

Escudero, Raquel; Barral, Marta; Pérez, Azucena; Vitutia, M. Mar; García-Pérez, Ana L.; Jiménez, Santos; Sellek, Ricela E.; Anda, Pedro

2000-01-01

Fifteen Borrelia burgdorferi sensu lato isolates from questing ticks and skin biopsy specimens from erythema migrans patients in three different areas of Spain were characterized. Four different genospecies were found (nine Borrelia garinii, including the two human isolates, three B. burgdorferi sensu stricto, two B. valaisiana, and one B. lusitaniae), showing a diverse spectrum of B. burgdorferi sensu lato species. B. garinii isolates were highly variable in terms of pulsed-field gel electrophoresis pattern and OspA serotype, with four of the seven serotypes described. One of the human isolates was OspA serotype 5, the same found in four of seven tick isolates. The second human isolate was OspA serotype 3, which was not present in ticks from the same area. Seven B. garinii isolates were able to disseminate through the skin of C3H/HeN mice and to cause severe inflammation of joints. One of the two B. valaisiana isolates also caused disease in mice. Only one B. burgdorferi sensu stricto isolate was recovered from the urinary bladder. One isolate each of B. valaisiana and B. lusitaniae were not able to disseminate through the skin of mice or to infect internal organs. In summary, there is substantial diversity in the species and in the pathogenicity of B. burgdorferi sensu lato in areas in northern Spain where Lyme disease is endemic. PMID:11060064

C6 Peptide-Based Multiplex Phosphorescence Analysis (PHOSPHAN) for Serologic Confirmation of Lyme Borreliosis.

PubMed

Pomelova, Vera G; Korenberg, Edward I; Kuznetsova, Tatiana I; Bychenkova, Tatiana A; Bekman, Natalya I; Osin, Nikolay S

2015-01-01

A single-tier immunoassay using the C6 peptide of VlsE (C6) from Borrelia burgdorferi sensu stricto (Bb) has been proposed as a potential alternative to conventional two-tier testing for the serologic diagnosis of Lyme disease in the United States and Europe. To evaluate the performance of C6 peptide based multiplex Phosphorescence Analysis (PHOSPHAN) for the serologic confirmation of Lyme borreliosis (LB) in Russian patients. Serum samples (n = 351) were collected from 146 patients with erythema migrans (EM); samples from 131 of these patients were taken several times prior to treatment and at different stages of recovery. The control group consisted of 197 healthy blood donors and 31 patients with other diseases, all from the same highly endemic region of Russia. All samples were analyzed by PHOSPHAN for IgM and IgG to Bb C6, recombinant OspC and VlsE proteins, and C6 peptides from B. garinii and B. afzelii. IgM and IgG to Bb C6 were identified in 43 and 95 out of 131 patients (32.8 and 72.5%, respectively); seroconversion of IgM antibodies was observed in about half of the patients (51.2%), and of IgG antibodies, in almost all of them (88.4%). Additional detection of OspC-IgM and VlsE-IgM or IgG to C6 from B. garinii or B. afzelii did not contribute significantly to the overall sensitivity of the multiplex immunoassay. The multiplex phosphorescence immunoassay is a promising method for simultaneously revealing the spectrum of antibodies to several Borrelia antigens. Detection of IgM and IgG to Bb C6 in the sera of EM patients provides effective serologic confirmation of LB and, with high probability, indicates an active infection process.

The application of similar image retrieval in electronic commerce.

PubMed

Hu, YuPing; Yin, Hua; Han, Dezhi; Yu, Fei

2014-01-01

Traditional online shopping platform (OSP), which searches product information by keywords, faces three problems: indirect search mode, large search space, and inaccuracy in search results. For solving these problems, we discuss and research the application of similar image retrieval in electronic commerce. Aiming at improving the network customers' experience and providing merchants with the accuracy of advertising, we design a reasonable and extensive electronic commerce application system, which includes three subsystems: image search display subsystem, image search subsystem, and product information collecting subsystem. This system can provide seamless connection between information platform and OSP, on which consumers can automatically and directly search similar images according to the pictures from information platform. At the same time, it can be used to provide accuracy of internet marketing for enterprises. The experiment shows the efficiency of constructing the system.

The Application of Similar Image Retrieval in Electronic Commerce

PubMed Central

Hu, YuPing; Yin, Hua; Han, Dezhi; Yu, Fei

2014-01-01

Traditional online shopping platform (OSP), which searches product information by keywords, faces three problems: indirect search mode, large search space, and inaccuracy in search results. For solving these problems, we discuss and research the application of similar image retrieval in electronic commerce. Aiming at improving the network customers' experience and providing merchants with the accuracy of advertising, we design a reasonable and extensive electronic commerce application system, which includes three subsystems: image search display subsystem, image search subsystem, and product information collecting subsystem. This system can provide seamless connection between information platform and OSP, on which consumers can automatically and directly search similar images according to the pictures from information platform. At the same time, it can be used to provide accuracy of internet marketing for enterprises. The experiment shows the efficiency of constructing the system. PMID:24883411

The Genetics of a Small Autosomal Region of DROSOPHILA MELANOGASTER Containing the Structural Gene for Alcohol Dehydrogenase. I. Characterization of Deficiencies and Mapping of ADH and Visible Mutations

PubMed Central

Woodruff, R. C.; Ashburner, M.

1979-01-01

The position of the structural gene coding for alcohol dehydrogenase (ADH) in Drosophila melanogaster has been shown to be within polytene chromosome bands 35B1 and 35B3, most probably within 35B2. The genetic and cytological properties of twelve deficiencies in polytene chromosome region 34–35 have been characterized, eleven of which include Adh. Also mapped cytogenetically are seven other recessive visible mutant loci. Flies heterozygous for overlapping deficiencies that include both the Adh locus and that for the outspread mutant (osp: a recessive wing phenotype) are homozygous viable and show a complete ADH negative phenotype and strong osp phenotype. These deficiencies probably include two polytene chromosome bands, 35B2 and 35B3. PMID:115743

A triaxial accelerometer monkey algorithm for optimal sensor placement in structural health monitoring

NASA Astrophysics Data System (ADS)

Jia, Jingqing; Feng, Shuo; Liu, Wei

2015-06-01

Optimal sensor placement (OSP) technique is a vital part of the field of structural health monitoring (SHM). Triaxial accelerometers have been widely used in the SHM of large-scale structures in recent years. Triaxial accelerometers must be placed in such a way that all of the important dynamic information is obtained. At the same time, the sensor configuration must be optimal, so that the test resources are conserved. The recommended practice is to select proper degrees of freedom (DOF) based upon several criteria and the triaxial accelerometers are placed at the nodes corresponding to these DOFs. This results in non-optimal placement of many accelerometers. A ‘triaxial accelerometer monkey algorithm’ (TAMA) is presented in this paper to solve OSP problems of triaxial accelerometers. The EFI3 measurement theory is modified and involved in the objective function to make it more adaptable in the OSP technique of triaxial accelerometers. A method of calculating the threshold value based on probability theory is proposed to improve the healthy rate of monkeys in a troop generation process. Meanwhile, the processes of harmony ladder climb and scanning watch jump are proposed and given in detail. Finally, Xinghai NO.1 Bridge in Dalian is implemented to demonstrate the effectiveness of TAMA. The final results obtained by TAMA are compared with those of the original monkey algorithm and EFI3 measurement, which show that TAMA can improve computational efficiency and get a better sensor configuration.

Detecting the Lyme Disease Spirochete, Borrelia Burgdorferi, in Ticks Using Nested PCR.

PubMed

Wills, Melanie K B; Kirby, Andrea M; Lloyd, Vett K

2018-02-04

Lyme disease is a serious vector-borne infection that is caused by the Borrelia burgdorferi sensu lato family of spirochetes, which are transmitted to humans through the bite of infected Ixodes ticks. The primary etiological agent in North America is Borrelia burgdorferi sensu stricto. As geographic risk regions expand, it is prudent to support robust surveillance programs that can measure tick infection rates, and communicate findings to clinicians, veterinarians, and the general public. The molecular technique of nested polymerase chain reaction (nPCR) has long been used for this purpose, and it remains a central, inexpensive, and robust approach in the detection of Borrelia in both ticks and wildlife. This article demonstrates the application of nPCR to tick DNA extracts to identify infected specimens. Two independent B. burgdorferi targets, genes encoding Flagellin B (FlaB) and Outer surface protein A (OspA), have been used extensively with this technique. The protocol involves tick collection, DNA extraction, and then an initial round of PCR to detect each of the two Borrelia-specific loci. Subsequent polymerase chain reaction (PCR) uses the product of the first reaction as a new template to generate smaller, internal amplification fragments. The nested approach improves upon both the specificity and sensitivity of conventional PCR. A tick is considered positive for the pathogen when inner amplicons from both Borrelia genes can be detected by agarose gel electrophoresis.

REGIONAL RESEARCH, METHODS, AND SUPPORT

EPA Science Inventory

The Human Exposure and Atmospheric Sciences Division (HEASD) has several collaborations with regional partners through the Regional Science Program (RSP) managed by ORD's Office of Science Policy (OSP). These projects resulted from common interests outlined in the Regional Appli...

Effects of temperature and operation parameters on the galvanic corrosion of Cu coupled to Au in organic solderability preservatives process

NASA Astrophysics Data System (ADS)

Oh, SeKwon; Kim, YoungJun; Jung, KiMin; Kim, JongSoo; Shon, MinYoung; Kwon, HyukSang

2017-03-01

In this work, we quantitatively examined the effects of temperature and operation parameters such as anode (Cu) to cathode (Au) area ratio, stirring speed, and Cu ion concentration on the galvanic corrosion kinetics of Cu coupled to Au (icouple ( Cu-Au)) on print circuit board in organic solderability preservative (OSP) soft etching solution. With the increase of temperature, galvanic corrosion rate (icouple ( Cu-Au) was increased; however, the degree of galvanic corrosion rate (icouple ( Cu-Au) - icorr (Cu)) was decreased owing to the lower activation energy of Cu coupled to Au, than that of Cu alone. With the increase of area ratio (cathode/anode), stirring speed of the system, icouple ( Cu-Au) was increased by the increase of cathodic reaction kinetics. And icouple ( Cu-Au) was decreased by the increase of the Cu-ion concentration in the OSP soft etching solution.

Lockheed Martin Response to the OSP Challenge

NASA Technical Reports Server (NTRS)

Sullivan, Robert T.; Munkres, Randy; Megna, Thomas D.; Beckham, Joanne

2003-01-01

The Lockheed Martin Orbital Space Plane System provides crew transfer and rescue for the International Space Station more safely and affordably than current human space transportation systems. Through planned upgrades and spiral development, it is also capable of satisfying the Nation's evolving space transportation requirements and enabling the national vision for human space flight. The OSP System, formulated through rigorous requirements definition and decomposition, consists of spacecraft and launch vehicle flight elements, ground processing facilities and existing transportation, launch complex, range, mission control, weather, navigation, communication and tracking infrastructure. The concept of operations, including procurement, mission planning, launch preparation, launch and mission operations and vehicle maintenance, repair and turnaround, is structured to maximize flexibility and mission availability and minimize program life cycle cost. The approach to human rating and crew safety utilizes simplicity, performance margin, redundancy, abort modes and escape modes to mitigate credible hazards that cannot be designed out of the system.

Effects of PCB Pad Metal Finishes on the Cu-Pillar/Sn-Ag Micro Bump Joint Reliability of Chip-on-Board (COB) Assembly

NASA Astrophysics Data System (ADS)

Kim, Youngsoon; Lee, Seyong; Shin, Ji-won; Paik, Kyung-Wook

2016-06-01

While solder bumps have been used as the bump structure to form the interconnection during the last few decades, the continuing scaling down of devices has led to a change in the bump structure to Cu-pillar/Sn-Ag micro-bumps. Cu-pillar/Sn-Ag micro-bump interconnections differ from conventional solder bump interconnections in terms of their assembly processing and reliability. A thermo-compression bonding method with pre-applied b-stage non-conductive films has been adopted to form solder joints between Cu pillar/Sn-Ag micro bumps and printed circuit board vehicles, using various pad metal finishes. As a result, various interfacial inter-metallic compounds (IMCs) reactions and stress concentrations occur at the Cu pillar/Sn-Ag micro bumps joints. Therefore, it is necessary to investigate the influence of pad metal finishes on the structural reliability of fine pitch Cu pillar/Sn-Ag micro bumps flip chip packaging. In this study, four different pad surface finishes (Thin Ni ENEPIG, OSP, ENEPIG, ENIG) were evaluated in terms of their interconnection reliability by thermal cycle (T/C) test up to 2000 cycles at temperatures ranging from -55°C to 125°C and high-temperature storage test up to 1000 h at 150°C. The contact resistances of the Cu pillar/Sn-Ag micro bump showed significant differences after the T/C reliability test in the following order: thin Ni ENEPIG > OSP > ENEPIG where the thin Ni ENEPIG pad metal finish provided the best Cu pillar/Sn-Ag micro bump interconnection in terms of bump joint reliability. Various IMCs formed between the bump joint areas can account for the main failure mechanism.

Nanoindentation on SnAgCu lead-free solder joints and analysis

NASA Astrophysics Data System (ADS)

Xu, Luhua; Pang, John H. L.

2006-12-01

The lead-free SnAgCu (SAC) solder joint on copper pad with organic solderability preservative (Cu-OSP) and electroless nickel and immersion gold (ENIG) subjected to thermal testing leads to intermetallic growth. It causes corresponding reliability concerns at the interface. Nanoindentation characterization on SnAgCu solder alloy, intermetallic compounds (IMCs), and the substrates subjected to thermal aging is reported. The modulus and hardness of thin IMC layers were measured by nanoindentation continuous stiffness measurement (CSM) from planar IMC surface. When SAC/Ni(Au) solder joints were subject to thermal aging, the Young’s modulus of the NiCuSn IMC at the SAC/ENIG specimen changed from 207 GPa to 146 GPa with different aging times up to 500 h. The hardness decreased from 10.0 GPa to 7.3 GPa. For the SAC/Cu-OSP reaction couple, the Young’s modulus of Cu6Sn5 stayed constant at 97.0 GPa and hardness about 5.7 GPa. Electron-probe microanalysis (EPMA) was used to thermal aging. The creep effect on the measured result was analyzed when measuring SnAgCu solder; it was found that the indentation penetration, and thus the hardness, is loading rate dependent. With the proposed constant P/P experiment, a constant indentation strain rate h/h and hardness could be achieved. The log-log plot of indentation strain rate versus hardness for the data from the constant P/P experiments yields a slope of 7.52. With the optimized test method and CSM Technique, the Modulus of SAC387 solder alloy and all the layers in a solder joint were investigated.

Antibody Secreting Cell Responses following Vaccination with Bivalent Oral Cholera Vaccine among Haitian Adults.

PubMed

Matias, Wilfredo R; Falkard, Brie; Charles, Richelle C; Mayo-Smith, Leslie M; Teng, Jessica E; Xu, Peng; Kováč, Pavol; Ryan, Edward T; Qadri, Firdausi; Franke, Molly F; Ivers, Louise C; Harris, Jason B

2016-06-01

The bivalent whole-cell (BivWC) oral cholera vaccine (Shanchol) is effective in preventing cholera. However, evaluations of immune responses following vaccination with BivWC have been limited. To determine whether BivWC induces significant mucosal immune responses, we measured V. cholerae O1 antigen-specific antibody secreting cell (ASC) responses following vaccination. We enrolled 24 Haitian adults in this study, and administered doses of oral BivWC vaccine 14 days apart (day 0 and day 14). We drew blood at baseline, and 7 days following each vaccine dose (day 7 and 21). Peripheral blood mononuclear cells (PBMCs) were isolated, and ASCs were enumerated using an ELISPOT assay. Significant increases in Ogawa (6.9 cells per million PBMCs) and Inaba (9.5 cells per million PBMCs) OSP-specific IgA ASCs were detected 7 days following the first dose (P < 0.001), but not the second dose. The magnitude of V. cholerae-specific ASC responses did not appear to be associated with recent exposure to cholera. ASC responses measured against the whole lipolysaccharide (LPS) antigen and the OSP moiety of LPS were equivalent, suggesting that all or nearly all of the LPS response targets the OSP moiety. Immunization with the BivWC oral cholera vaccine induced ASC responses among a cohort of healthy adults in Haiti after a single dose. The second dose of vaccine resulted in minimal ASC responses over baseline, suggesting that the current dosing schedule may not be optimal for boosting mucosal immune responses to V. cholerae antigens for adults in a cholera-endemic area.

Understanding Preclerkship Medical Students' Poor Performance in Prescription Writing.

PubMed

James, Henry; Al Khaja, Khalid A J; Tayem, Yasin I; Veeramuthu, Sindhan; Sequeira, Reginald P

2016-05-01

This study aimed to explore reasons for poor performance in prescription writing stations of the objective structured practical examination (OSPE) and absenteeism in prescription writing sessions among preclerkship medical students at the Arabian Gulf University (AGU) in Manama, Bahrain. This descriptive study was carried out between September 2014 and June 2015 among 157 preclerkship medical students at AGU. Data were collected using focus group discussions and a questionnaire with closed- and open-ended items. All 157 students participated in the study (response rate: 100.0%). The most frequently cited reasons for poor performance in OSPE stations were an inability to select the correct drugs (79.6%), treatment duration (69.4%), drug quantity (69.4%) and drug formulation (68.2%). Additionally, students reported inadequate time for completing the stations (68.8%). During focus group discussions, students reported other reasons for poor performance, including examination stress and the difficulty of the stations. Absenteeism was attributed to the length of each session (55.4%), lack of interest (50.3%), reliance on peers for information (48.4%) and optional attendance policies (47.1%). Repetitive material, large group sessions, unmet student expectations and the proximity of the sessions to summative examinations were also indicated to contribute to absenteeism according to open-ended responses or focus group discussions. This study suggests that AGU medical students perform poorly in prescription writing OSPE stations because of inadequate clinical pharmacology knowledge. Participation in prescription writing sessions needs to be enhanced by addressing the concerns identified in this study. Strategies to improve attendance and performance should take into account the learner-teacher relationship.

Trends in cervical cancer incidence and mortality in Poland: is there an impact of the introduction of the organised screening?

PubMed

Nowakowski, Andrzej; Wojciechowska, Urszula; Wieszczy, Paulina; Cybulski, Marek; Kamiński, Michał F; Didkowska, Joanna

2017-06-01

Aside from existing opportunistic screening, an organised screening programme (OSP) for cervical cancer (CC) was implemented in 2006/2007 in Poland. We applied joinpoint regression and age-period-cohort model to look for the impact of the OSP on CC incidence/mortality trends. Decline of age-standardised incidence rates (ASIRs) in the screening-age group (25-59 years) accelerated from -2.2% (95% CI -2.7 to -1.7%) between 1993 and 2008 to -6.1% (95% CI -7.7 to -4.4%) annually after 2008. In women aged 60+ years, ASIRs declined from 1986 until 2005 [annual percent change (APC) = -2.6%, 95% CI -2.9 to -2.4%] and stabilised thereafter. Decline of age-standardised mortality rates (ASMRs) in the screening-age group accelerated from -1.3% (95% CI -1.5 to -1.1%) between 1980 and 2005 to -4.7% (95% CI -5.6 to -3.8%) annually after 2005. In women aged 60+ ASMR declined between 1991 and 2004 (APC = -2.9%, 95% CI -3.5 to -2.3%) and stabilised thereafter. Relative risks of CC diagnosis and death were 0.63 (95% CI 0.62-0.65) and 0.61 (95% CI 0.59-0.63), respectively, for the most recent period compared to the reference around 1982. Implementation of the OSP possibly accelerated downward trends in the burden of CC in Polish women under the age of 60, but recent stabilisation of trends in older women requires actions.

PLMA vs. I-gel: A Comparative Evaluation of Respiratory Mechanics in Laparoscopic Cholecystectomy

PubMed Central

Sharma, Bimla; Sehgal, Raminder; Sahai, Chand; Sood, Jayashree

2010-01-01

Background: Supraglottic airway devices (SADs), such as ProSealTM laryngeal mask airway (PLMA), which produce high oropharyngeal seal pressure (OSP) and have the facility for gastric decompression have been used in laparoscopic procedures. i-gel is a new SAD which shares these features with the PLMA. This study was designed to compare the respiratory mechanics of these two devices during positive pressure ventilation in anaesthetised adult patients undergoing laparoscopic cholecystectomy. Patients & Methods: The study included 60 ASA I-II adult patients scheduled for laparoscopic cholecystectomy. The patients were randomized to two groups of 30 each, with either PLMA or i-gel as their airway device. Anaesthesia and premedication were standardized for both the groups. In addition to routine monitoring, neuromuscular monitoring with TOF ratio, OSP and respiratory mechanics monitoring (dynamic compliance, resistance, work of breathing, measured minute ventilation and peak airway pressures) were employed. Fibreoptic evaluation of positioning of the devices and adverse events related to them were also compared. Results: The OSP (cm H2O) were higher for PLMA (38.9 vs. 35.6, P=0.007). The respiratory mechanics parameters using the two devices were comparable apart from the dynamic compliance, which was significantly higher with i-gel (P < 0.05). Malrotation was higher with i-gel than with PLMA (15 vs. 5, P = 0.006). Conclusion: The PLMA formed a better seal while the dynamic compliance was higher with the i-gel. Both devices provided optimal ventilation and oxygenation and the adverse events were also comparable. PMID:21547168

THE OPEN SOURCING OF EPANET

EPA Science Inventory

A proposal was made at the 2009 EWRI Congress in Kansas City, MO to establish an Open Source Project (OSP) for the widely used EPANET pipe network analysis program. This would be an ongoing collaborative effort among a group of geographically dispersed advisors and developers, wo...

Autonomous Space Shuttle

NASA Technical Reports Server (NTRS)

Siders, Jeffrey A.; Smith, Robert H.

2004-01-01

The continued assembly and operation of the International Space Station (ISS) is the cornerstone within NASA's overall Strategic P an. As indicated in NASA's Integrated Space Transportation Plan (ISTP), the International Space Station requires Shuttle to fly through at least the middle of the next decade to complete assembly of the Station, provide crew transport, and to provide heavy lift up and down mass capability. The ISTP reflects a tight coupling among the Station, Shuttle, and OSP programs to support our Nation's space goal . While the Shuttle is a critical component of this ISTP, there is a new emphasis for the need to achieve greater efficiency and safety in transporting crews to and from the Space Station. This need is being addressed through the Orbital Space Plane (OSP) Program. However, the OSP is being designed to "complement" the Shuttle as the primary means for crew transfer, and will not replace all the Shuttle's capabilities. The unique heavy lift capabilities of the Space Shuttle is essential for both ISS, as well as other potential missions extending beyond low Earth orbit. One concept under discussion to better fulfill this role of a heavy lift carrier, is the transformation of the Shuttle to an "un-piloted" autonomous system. This concept would eliminate the loss of crew risk, while providing a substantial increase in payload to orbit capability. Using the guidelines reflected in the NASA ISTP, the autonomous Shuttle a simplified concept of operations can be described as; "a re-supply of cargo to the ISS through the use of an un-piloted Shuttle vehicle from launch through landing". Although this is the primary mission profile, the other major consideration in developing an autonomous Shuttle is maintaining a crew transportation capability to ISS as an assured human access to space capability.

76 FR 50482 - Proposed Information Collection Activity; Comment Request

Federal Register 2010, 2011, 2012, 2013, 2014

2011-08-15

... Information Collection Activity; Comment Request Title: Objective Work Plan (OSP), Objective Progress Report... previous OPR. Project Sustainability: Content is the same. Questions 12 and 13 were originally questions 18..., Office of Information Services, 370 L'Enfant Promenade, SW., Washington, DC 20447, Attn: ACF Reports...

Reliability and paste process optimization of eutectic and lead-free for mixed packaging

NASA Technical Reports Server (NTRS)

Ramkumar, S. M.; Ganeshan, V.; Thenalur, K.; Ghaffarian, R.

2002-01-01

This paper reports the results of an experiment that utilized the JPL's area array consortium test vehicle design, containing a myriad of mixed technology components with an OSP finish. The details of the reliability study are presented in this paper.

Study of the impact of resonant magnetic perturbation fields on gross tungsten erosion using DiMES samples in DIII-D

NASA Astrophysics Data System (ADS)

Hinson, E. T.; Schmitz, O.; Frerichs, H.; Abrams, T.; Briesemeister, A.; Rudakov, D. L.; Unterberg, E. A.; Wampler, W. R.; Watkins, J. G.; Wang, H. Q.

2017-12-01

An experiment was conducted in DIII-D to compare gross tungsten (W) erosion on samples exposed to outer strike point (OSP) sweeps in L-mode plasmas for three conditions. These included two phases of resonant magnetic perturbations (RMPs), and a set with no perturbations. Upon RMP application, lobe structures indicative of strike point splitting of the OSP were evident in divertor camera data and on Langmuir probes. Gross W erosion flux, {{{Γ }}}{{W}}, inferred spectroscopically using the S/XB method applied to the 400.9 nm W-I line, was generally in the range {{{Γ }}}{{W}}/{{{Γ }}}{{D}+,\\perp }=2× {10}-4 referenced to incident deuterium ion flux {{{Γ }}}{{D}+,\\perp }, and was increased in the RMP cases by no more than 30% of the level observed in unperturbed discharges. A large reduction in gross erosion (50%) was observed in the private flux region at the W sample for one specific toroidal phase of the RMP field.

The need for a single cleaning standard for OEM and OSP fiber optic connections

NASA Astrophysics Data System (ADS)

Forrest, Edward J., Jr.; Blair, Paul

2009-01-01

Development of fiber optic communications networks has continued in virtually all geographies of the world. Some calculate that expansion in the USA alone will continue to 2025. After that time there will maintenance, upgrade, and replacement. While some nations are more advanced than others and some enterprises more sophisticated than others, the undeniable reality is that previous networks will, in some fashion or another, include fiber optics. The future need is for speed and reliable bandwidth: lots of it. Decisions will be made based on new concepts that will enable far more data at far faster rates than once considered feasible or necessary. As the consumer enters the market with newly engineered home entertainment systems, additional burdens will be placed on both OEM and OSP. These are not negative challenges, but rather indicators of a new generation of consumers with awareness and demand far more advanced any may have imagined just a few years ago.

Relativistic effects in iron-, ruthenium-, and osmium porphyrins

NASA Astrophysics Data System (ADS)

Liao, Meng-Sheng; Scheiner, Steve

2002-12-01

Nonrelativistic and relativistic DFT calculations are performed on four-coordinate metal porphyrins MP and their six-coordinate adducts MP(py) 2 and MP(py)(CO) (py=pyridine) with M=Fe, Ru, and Os. The electronic structures of the MPs are investigated by considering all possible low-lying states with different configurations of nd-electrons. FeP and OsP have a 3A2 g ground state, while this state is nearly degenerate with 3Eg for RuP. Without relativistic corrections, the ground states of both RuP and OsP would be 3Eg. For the six-coordinate adducts with py and CO, the strong-field axial ligands raise the energy of the M d z2-orbital, thereby making the M II ion diamagnetic. The calculated redox properties of MP(py) 2 and MP(py)(CO) are in agreement with experiment. The difference between RuP(py)(CO) and OsP(py)(CO), in terms of site of oxidation, is due to relativistic effects.

Er Effect of Low Molecular Liquid Crystal on One-Sided Patterned Electrodes

NASA Astrophysics Data System (ADS)

Kikuchi, Takehito; Inoue, Akio; Furusho, Junji; Kawamuki, Ryohei

Several kinds of ER fluids (ERF) have been developed and have been applied to some mechatronics devices and processing technologies. In many conventional applications of ERFs, these devices consist of bilateral electrodes to apply electric field in ERF. However, the electric field of several kV/mm may be necessary to generate an ER effect sufficiently for practical purposes. The gap between a pair of electrodes should be, therefore, maintained narrowly and exactly for fears of short-circuit. At the same time, this electrode system also requires an interconnection on driving parts. To improve these disadvantages, we proposed "one-sided patterned electrode" (OSPE) systems in previous works. In this report, we confirmed the flow characteristics of low molecular liquid crystal (LMLC) on OSPE. Next, we also confirmed the different characteristics depending on the pattern type. Depending on results of electro-static analysis, we conclude that such a difference may results from the directors of LC molecules derived by electric field.

Broad diversity of host responses of the white-footed mouse Peromyscus leucopus to Borrelia infection and antigens.

PubMed

Cook, Vanessa; Barbour, Alan G

2015-07-01

Peromyscus leucopus, the white-footed mouse, is one of the more abundant mammals of North America and is a major reservoir host for at least five tickborne diseases of humans, including Lyme disease and a newly-recognized form of relapsing fever. In comparison to Mus musculus, which is not a natural reservoir for any of these infections, there has been little research on experimental infections in P. leucopus. With the aim of further characterizing the diversity of phenotypes of host responses, we studied a selection of quantitative traits in colony-bred and -reared outbred P. leucopus adults that were uninfected, infected with the relapsing fever agent Borrelia hermsii alone, or infected after immunization with Lyme disease vaccine antigen OspA and keyhole limpet hemocyanin (KLH). The methods included measurements of organ weights, hematocrits, and bleeding times, quantitative PCR for bacterial burdens, and enzyme immunoassays for serum antibodies against both the immunization proteins and cellular antigens of the infecting organism. The results included the following: (i) uninfected animals displayed wide variation in relative sizes of their spleens and in their bleeding times. (ii) In an experiment with matched littermates, no differences were observed between females and males at 7 days of infection in bacterial burdens in blood and spleen, relative spleen size, or antibody responses to the B. hermsii specific-antigen, FbpC. (iii) In studies of larger groups of males or females, the wide variations between bacterial burdens and in relative spleen sizes between individuals was confirmed. (iv) In these separate groups of males and females, all animals showed moderate-to-high levels of antibodies to KLH but wide variation in antibody levels to OspA and to FbpC. The study demonstrated the diversity of host responses to infection and immunization in this species and identified quantitative traits that may be suitable for forward genetics approaches to reservoir-pathogen interactions. Copyright © 2015 Elsevier GmbH. All rights reserved.

Quantum supergroups and solutions of the Yang-Baxter equation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bracken, A.J.; Gould, M.D.; Zhang, R.B.

1990-05-10

A method is developed for systematically constructing trigonometric and rational solutions of the Yang-Baxter equation using the representation theory of quantum supergroups. New quantum R-matrices are obtained by applying the method to the vector representations of quantum osp(1/2) and gl(m/n).

Unlicensed and unshackled : a joint OSP-OET white paper on unlicensed devices and their regulatory issues

DOT National Transportation Integrated Search

2003-05-01

In this paper, the authors present a survey of the origins of unlicensed wireless devices, their governing regulation, the current technological state of the art, an overview of the market with information from publicly available sources, and an anal...

50 CFR 403.02 - Definitions.

Code of Federal Regulations, 2012 CFR

2012-10-01

.... Maximum net productivity is the greatest net annual increment in population numbers or biomass resulting... term species includes any population stock. (b) Optimum Sustainable Population or OSP means a population size which falls within a range from the population level of a given species or stock which is the...

50 CFR 403.02 - Definitions.

Code of Federal Regulations, 2014 CFR

2014-10-01

.... Maximum net productivity is the greatest net annual increment in population numbers or biomass resulting... term species includes any population stock. (b) Optimum Sustainable Population or OSP means a population size which falls within a range from the population level of a given species or stock which is the...

50 CFR 403.02 - Definitions.

Code of Federal Regulations, 2013 CFR

2013-10-01

.... Maximum net productivity is the greatest net annual increment in population numbers or biomass resulting... term species includes any population stock. (b) Optimum Sustainable Population or OSP means a population size which falls within a range from the population level of a given species or stock which is the...

50 CFR 403.02 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-10-01

.... Maximum net productivity is the greatest net annual increment in population numbers or biomass resulting... term species includes any population stock. (b) Optimum Sustainable Population or OSP means a population size which falls within a range from the population level of a given species or stock which is the...

48 CFR 1201.301-70 - Amendment of (TAR) 48 CFR chapter 12.

Code of Federal Regulations, 2011 CFR

2011-10-01

... from internal DOT personnel, other Government agencies, or the public. Changes shall be submitted in the following format to the Office of the Senior Procurement Executive (OSPE), 400 7th Street, SW... formulate Departmental acquisition policies and procedures. (1) Transportation Acquisition Circular (TAC...

Seasonal and spatial patterns of heterotrophic bacterial production, respiration, and biomass in the subarctic NE Pacific

NASA Astrophysics Data System (ADS)

Sherry, Nelson D.; Boyd, Philip W.; Sugimoto, Kugako; Harrison, Paul J.

1999-11-01

Heterotrophic bacterial biomass, production, and respiration rates were measured during winter, spring, and summer in the subarctic NE Pacific from September 1995 to June 1997. Sampling took place on six cruises at five hydrographic stations along the east/west line-P transect from slope waters at P4 (1200 m depth) to the open-ocean waters at Ocean Station Papa (OSP) (4250 m depth). Interannual variability was small relative to seasonal and spatial variability. Biomass, derived from cell counts (assuming 20 fg C cell -1), was ca. 12 μg C l -1 in the winter and increased to 20-35 μg C l -1 in the spring and summer all along line-P. Bacterial production from [ 3H]-thymidine and [ 14C]-leucine incorporation rates was lowest in the winter (ca. 0.5 μg C l -1 d -1) with little spatial variability. Production increased 10-fold in spring at P4 (to ca. 4.5 μg C l -1 d -1). In contrast, only a 2-fold increase in bacterial production was observed over this period at the more oceanic stations. Rates of production in late summer were highest over the annual cycle at all stations ranging from ca. 6 at P4 to ca. 2 μg C l -1 d -1 at OSP. Bacterial (<1 μm size fraction) respiration, measured from dark-bottle O 2 consumption over 24 or 48 h, was <10 μg C l -1 d -1 during the winter and spring. Respiration rates increased >10-fold to ca. 100 μg C l -1 d -1 at P4 in the summer, but, interestingly, did not increase from spring to summer at the more oceanic stations. Thus bacterial growth efficiency, defined as production/(production+respiration), decreased in the spring westwards from the slope waters (P4) to the open-ocean (OSP), but increased westwards in the summer. Bacterial production was highly correlated with temperature at OSP ( r2=0.88) and less so at P4 ( r2=0.50). The observed temporal and spatial trends presented in this study suggest that seasonal changes in bacterial biomass were greatly affected by changes in loss processes, that bacterial biomass is regulated by different processes than bacterial production, and that bacterial production alone, without respiration measurements, is not a robust proxy for bacterial activity in the subarctic NE Pacific.

ORD/REGIONAL/OAR AIR TOXICS EXPOSURE ASSESSMENT WORKSHOP (SAN FRANCISCO,JUNE 25-27, 2002)

EPA Science Inventory

Approximately, one hundred scientists from ORD, OAR, Regional offices, and invited external presentors met for a three day conference in Region 9 offices in San Francisco, CA. Dr. Winona Victery, Science Advisor in Region 9 and Dr. David Klauder, ORD/OSP program manager worked ...

Making the Invisible Visible: The Oklahoma Science Project.

ERIC Educational Resources Information Center

McCarty, Robbie; Pedersen, Jon E.

2002-01-01

Reports that teachers in preservice education programs still view the teaching of science much in the same traditional ways as our predecessors. "The Oklahoma Science Project (OSP) Model for Professional Development: Practicing Science Across Contexts" will build discourses and relationships that can be extended across contexts to establish…

75 FR 51272 - Proposed Collection; Comment Request; STAR METRICS-Science and Technology in America's...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-08-19

... data collection projects, the Office of Science Policy Analysis (OSPA), the National Institutes of..., Health Science Policy Analyst, Office of Science and Technology Policy, OSP, OD; NIH, Building 1, Room... this publication. Dated: August 12, 2010. Lynn D. Hudson, Director, Office of Science Policy Analysis...

Vaccination against Lyme disease: past, present, and future.

PubMed

Embers, Monica E; Narasimhan, Sukanya

2013-01-01

Lyme borreliosis is a zoonotic disease caused by Borrelia burgdorferi sensu lato bacteria transmitted to humans and domestic animals by the bite of an Ixodes spp. tick (deer tick). Despite improvements in diagnostic tests and public awareness of Lyme disease, the reported cases have increased over the past decade to approximately 30,000 per year. Limitations and failed public acceptance of a human vaccine, comprised of the outer surface A (OspA) lipoprotein of B. burgdorferi, led to its demise, yet current research has opened doors to new strategies for protection against Lyme disease. In this review we discuss the enzootic cycle of B. burgdorferi, and the unique opportunities it poses to block infection or transmission at different levels. We present the correlates of protection for this infectious disease, the pros and cons of past vaccination strategies, and new paradigms for future vaccine design that would include elements of both the vector and the pathogen.

Vaccination against Lyme disease: past, present, and future

PubMed Central

Embers, Monica E.; Narasimhan, Sukanya

2013-01-01

Lyme borreliosis is a zoonotic disease caused by Borrelia burgdorferi sensu lato bacteria transmitted to humans and domestic animals by the bite of an Ixodes spp. tick (deer tick). Despite improvements in diagnostic tests and public awareness of Lyme disease, the reported cases have increased over the past decade to approximately 30,000 per year. Limitations and failed public acceptance of a human vaccine, comprised of the outer surface A (OspA) lipoprotein of B. burgdorferi, led to its demise, yet current research has opened doors to new strategies for protection against Lyme disease. In this review we discuss the enzootic cycle of B. burgdorferi, and the unique opportunities it poses to block infection or transmission at different levels. We present the correlates of protection for this infectious disease, the pros and cons of past vaccination strategies, and new paradigms for future vaccine design that would include elements of both the vector and the pathogen. PMID:23407755

Family Reflections on the District of Columbia Opportunity Scholarship Program: Final Summary Report

ERIC Educational Resources Information Center

Stewart, Thomas; Wolf, Patrick; Cornman, Stephen Q.; McKenzie-Thompson, Kenann; Butcher, Jonathan

2009-01-01

During the spring of 2004, the first federally funded voucher program--the District of Columbia Opportunity Scholarship Program (OSP)--was established. The School Choice Demonstration Project (SCDP) recognized that publicly-funded school vouchers represent a relatively new and unstudied approach to school choice and education reform. To address …

Open Source Projects in Software Engineering Education: A Mapping Study

ERIC Educational Resources Information Center

Nascimento, Debora M. C.; Almeida Bittencourt, Roberto; Chavez, Christina

2015-01-01

Context: It is common practice in academia to have students work with "toy" projects in software engineering (SE) courses. One way to make such courses more realistic and reduce the gap between academic courses and industry needs is getting students involved in open source projects (OSP) with faculty supervision. Objective: This study…

A Trial of the Objective Structured Practical Examination in Physiology at Melaka Manipal Medical College, India

ERIC Educational Resources Information Center

Abraham, Reem Rachel; Raghavendra, Rao; Surekha, Kamath; Asha, Kamath

2009-01-01

A single examination does not fulfill all the functions of assessment. The present study was undertaken to determine the reliability and student satisfaction regarding the objective structured practical examination (OSPE) as a method of assessment of laboratory exercises in physiology before implementing it in the forthcoming university…

Optimal sensors placement and spillover suppression

NASA Astrophysics Data System (ADS)

Hanis, Tomas; Hromcik, Martin

2012-04-01

A new approach to optimal placement of sensors (OSP) in mechanical structures is presented. In contrast to existing methods, the presented procedure enables a designer to seek for a trade-off between the presence of desirable modes in captured measurements and the elimination of influence of those mode shapes that are not of interest in a given situation. An efficient numerical algorithm is presented, developed from an existing routine based on the Fischer information matrix analysis. We consider two requirements in the optimal sensor placement procedure. On top of the classical EFI approach, the sensors configuration should also minimize spillover of unwanted higher modes. We use the information approach to OSP, based on the effective independent method (EFI), and modify the underlying criterion to meet both of our requirements—to maximize useful signals and minimize spillover of unwanted modes at the same time. Performance of our approach is demonstrated by means of examples, and a flexible Blended Wing Body (BWB) aircraft case study related to a running European-level FP7 research project 'ACFA 2020—Active Control for Flexible Aircraft'.

GENETIC AND IMMUNOLOGICAL EVIDENCES OF BORRELIA BURGDORFERI IN DOG IN THAILAND.

PubMed

Sthitmatee, Nattawooti; Jinawan, Wanna; Jaisan, Nawaporn; Tangjitjaroen, Weerapongse; Chailangkarn, Sasisophin; Sodarat, Chollada; Ekgatat, Monaya; Padungtod, Pawin

2016-01-01

Lyme disease is a tick-borne zoonotic disease caused by spirochete Borrelia burgdorferi. It is transmitted from animals to humans by the bite of infected ticks of the genus Ixodes. Although Lyme disease has been reported in China and Japan, the disease has never been reported in Thailand. Blood samples and ticks were collected from 402 dogs from 7 and 3 animal clinics in Chiang Mai and Phuket Provinces, Thailand, respectively. Blood samples were tested for antibodies against B. burgdorferi, Anaplasma spp, Ehrlichia spp and Dirofilaria immitis using a commercial kit, and positive blood samples were subjected to nested PCR assay for B. burgdorferi fla, ospA and ospC, amplicons of which also were sequenced. Only one dog (from Chiang Mai) was positive for B. burgdorferi, with 97% to 100% genetic identity, depending on the sequences used for comparison, with strains from United State of America. All 376 ticks collected were Rhipicephalus sanguineus, but no tick was found on the infected dog. Further investigations of the infection source and vector are needed to understand potential risks of Lyme disease to dogs and humans in Thailand.

The effects of mindfulness-based stress reduction on objective and subjective sleep parameters in women with breast cancer: a randomized controlled trial.

PubMed

Lengacher, Cecile A; Reich, Richard R; Paterson, Carly L; Jim, Heather S; Ramesar, Sophia; Alinat, Carissa B; Budhrani, Pinky H; Farias, Jerrica R; Shelton, Melissa M; Moscoso, Manolete S; Park, Jong Y; Kip, Kevin E

2015-04-01

The purpose of this study was to investigate the effects of mindfulness-based stress reduction for breast cancer survivors (MBSR(BC)) on multiple measures of objective and subjective sleep parameters among breast cancer survivors (BCS). Data were collected using a two-armed randomized controlled design among BCS enrolled in either a 6-week MBSR(BC) program or a usual care (UC) group with a 12-week follow-up. The present analysis is a subset of the larger parent trial (ClinicalTrials.gov Identifier: NCT01177124). Seventy-nine BCS participants (mean age 57 years), stages 0-III, were randomly assigned to either the formal (in-class) 6-week MBSR(BC) program or UC. Subjective sleep parameters (SSP) (i.e., sleep diaries and the Pittsburgh Sleep Quality Index (PSQI)) and objective sleep parameters (OSP) (i.e., actigraphy) were measured at baseline, 6 weeks, and 12 weeks after completing the MBSR(BC) or UC program. Results showed indications of a positive effect of MBSR(BC) on OSP at 12 weeks on sleep efficiency (78.2% MBSR(BC) group versus 74.6% UC group, p = 0.04), percent of sleep time (81.0% MBSR(BC) group versus 77.4% UC group, p = 0.02), and less number waking bouts (93.5 in MBSR(BC) group versus 118.6 in the UC group, p < 0.01). Small nonsignificant improvements were found in SSP in the MBSR(BC) group from baseline to 6 weeks (PSQI total score, p = 0.09). No significant relationship was observed between minutes of MBSR(BC) practice and SSP or OSP. These data suggest that MBSR(BC) may be an efficacious treatment to improve objective and subjective sleep parameters in BCS. Copyright © 2014 John Wiley & Sons, Ltd.

Study and treatment of post Lyme disease (STOP-LD): a randomized double masked clinical trial.

PubMed

Krupp, L B; Hyman, L G; Grimson, R; Coyle, P K; Melville, P; Ahnn, S; Dattwyler, R; Chandler, B

2003-06-24

To determine whether post Lyme syndrome (PLS) is antibiotic responsive. The authors conducted a single-center randomized double-masked placebo-controlled trial on 55 patients with Lyme disease with persistent severe fatigue at least 6 or more months after antibiotic therapy. Patients were randomly assigned to receive 28 days of IV ceftriaxone or placebo. The primary clinical outcomes were improvement in fatigue, defined by a change of 0.7 points or more on an 11-item fatigue questionnaire, and improvement in cognitive function (mental speed), defined by a change of 25% or more on a test of reaction time. The primary laboratory outcome was an experimental measure of CSF infection, outer surface protein A (OspA). Outcome data were collected at the 6-month visit. Patients assigned to ceftriaxone showed improvement in disabling fatigue compared to the placebo group (rate ratio, 3.5; 95% CI, 1.50 to 8.03; p = 0.001). No beneficial treatment effect was observed for cognitive function or the laboratory measure of persistent infection. Four patients, three of whom were on placebo, had adverse events associated with treatment, which required hospitalization. Ceftriaxone therapy in patients with PLS with severe fatigue was associated with an improvement in fatigue but not with cognitive function or an experimental laboratory measure of infection in this study. Because fatigue (a nonspecific symptom) was the only outcome that improved and because treatment was associated with adverse events, this study does not support the use of additional antibiotic therapy with parenteral ceftriaxone in post-treatment, persistently fatigued patients with PLS.

Impact of Isothermal Aging on Long-Term Reliability of Fine-Pitch Ball Grid Array Packages with Sn-Ag-Cu Solder Interconnects: Surface Finish Effects

NASA Astrophysics Data System (ADS)

Lee, Tae-Kyu; Ma, Hongtao; Liu, Kuo-Chuan; Xue, Jie

2010-12-01

The interaction between isothermal aging and the long-term reliability of fine-pitch ball grid array (BGA) packages with Sn-3.0Ag-0.5Cu (wt.%) solder ball interconnects was investigated. In this study, 0.4-mm fine-pitch packages with 300- μm-diameter Sn-Ag-Cu solder balls were used. Two different package substrate surface finishes were selected to compare their effects on the final solder composition, especially the effect of Ni, during thermal cycling. To study the impact on thermal performance and long-term reliability, samples were isothermally aged and thermally cycled from 0°C to 100°C with 10 min dwell time. Based on Weibull plots for each aging condition, package lifetime was reduced by approximately 44% by aging at 150°C. Aging at 100°C showed a smaller impact but similar trend. The microstructure evolution was observed during thermal aging and thermal cycling with different phase microstructure transformations between electrolytic Ni/Au and organic solderability preservative (OSP) surface finishes, focusing on the microstructure evolution near the package-side interface. Different mechanisms after aging at various conditions were observed, and their impacts on the fatigue lifetime of solder joints are discussed.

MBBS Student Perceptions about Physiology Subject Teaching and Objective Structured Practical Examination Based Formative Assessment for Improving Competencies

ERIC Educational Resources Information Center

Lakshmipathy, K.

2015-01-01

The objectives of the present study were to 1) assess student attitudes to physiology, 2) evaluate student opinions about the influence of an objective structured practical examination (OSPE) on competence, and 3) assess the validity and reliability of an indigenously designed feedback questionnaire. A structured questionnaire containing 16 item…

Signaling via the CD2 receptor enhances HTLV-1 replication in T lymphocytes.

PubMed

Guyot, D J; Newbound, G C; Lairmore, M D

1997-07-21

Human T lymphotropic virus type 1 (HTLV-1) is considered the etiologic agent of adult T cell leukemia/lymphoma and several chronic progressive immune-mediated diseases. Approximately 1-4% of infected individuals develop disease, generally decades following infection. Increased proviral transcription, mediated by the viral 40-kDa trans-activating protein, Tax, has been implicated in the pathogenesis of HTLV-1-associated diseases. Since the HTLV-1 promoter contains sequences responsive to cyclic AMP and protein kinase C, we hypothesized that lymphocyte activation signals initiated through the TCR/CD3 complex or CD2 receptor promote viral replication in HTLV-1-infected lymphocytes. We demonstrate that mAbs directed against the CD2, but not the CD3 receptor increase viral p24 capsid protein 1.5- to 5.7-fold in CD2/CD3+ HTLV-1-infected cell culture supernatants. Northern blot analysis demonstrated a 2.5- to 4-fold increase in all species of viral mRNA following CD2 cross-linking of OSP2/4 cells, an immortalized HTLV-1 cell line. Consistent with transcriptional regulation, reporter gene activity increased approximately 11-fold in CD2-stimulated Jurkat T cells cotransfected with a Tax-expressing plasmid and a CAT reporter gene construct under control of the HTLV-1 promoter. These data suggest a possible physiologic mechanism, whereby CD2-mediated cell adhesion and lymphocyte activation may promote viral transcription in infected lymphocytes.

School Vouchers in Washington, DC: Achievement Impacts and Their Implications for Social Justice

ERIC Educational Resources Information Center

Wolf, Patrick J.

2010-01-01

The District of Columbia Opportunity Scholarship Program (OSP) is a school voucher initiative targeted to disadvantaged students in the US Capital. Vouchers worth up to $7,500 annually are awarded by lottery to students with family incomes near or below the federal poverty line. Students can then use their voucher at any of 60 participating…

Office of Strategic Programs FY 2017 Budget At-A-Glance

DOE Office of Scientific and Technical Information (OSTI.GOV)

None

2016-03-01

The Office of Strategic Programs (OSP) increases the overall effectiveness and impact of all EERE activities through key cross-cutting initiatives and strategic analysis, communications, and technology-to-market activities. OSP’s work directly contributes to EERE’s mission, facilitates and amplifies the successes of EERE technology offices, and soundly and consistently informs the Assistant Secretary’s decisions.

New Failure Mode of Flip-Chip Solder Joints Related to the Metallization of an Organic Substrate

NASA Astrophysics Data System (ADS)

Jang, J. W.; Yoo, S. J.; Hwang, H. I.; Yuk, S. Y.; Kim, C. K.; Kim, S. J.; Han, J. S.; An, S. H.

2015-10-01

We report a new failure phenomenon during flip-chip die attach. After reflow, flip-chip bumps were separated between the Al and Ti layers on the Si die side. This was mainly observed at the Si die corner. Transmission electron microscopy images revealed corrosion of the Al layer at the edge of the solder bump metallization. The corrosion at the metallization edge exhibited a notch shape with high stress concentration factor. The organic substrate had Cu metallization with an organic solderable preservative (OSP) coating layer, where a small amount of Cl ions were detected. A solder bump separation mechanism is suggested based on the reaction between Al and Cl, related to the flow of soldering flux. During reflow, the flux will dissolve the Cl-containing OSP layer and flow up to the Al layer on the Si die side. Then, the Cl-dissolved flux will actively react with Al, forming AlCl3. During cooling, solder bumps at the Si die corner will separate through the location of Al corrosion. This demonstrated that the chemistry of the substrate metallization can affect the thermomechanical reliability of flip-chip solder joints.

Established Population of Blacklegged Ticks with High Infection Prevalence for the Lyme Disease Bacterium, Borrelia burgdorferi Sensu Lato, on Corkscrew Island, Kenora District, Ontario

PubMed Central

Scott, John D.; Foley, Janet E.; Clark, Kerry L.; Anderson, John F.; Durden, Lance A.; Manord, Jodi M.; Smith, Morgan L.

2016-01-01

We document an established population of blacklegged ticks, Ixodes scapularis, on Corkscrew Island, Kenora District, Ontario, Canada. Primers of the outer surface protein A (OspA) gene, the flagellin (fla) gene, and the flagellin B (flaB) gene were used in the PCR assays to detect Borrelia burgdorferi sensu lato (s.l.), the Lyme disease bacterium. In all, 60 (73%) of 82 adult I. scapularis, were infected with B. burgdorferi s.l. As well, 6 (43%) of 14 unfed I. scapularis nymphs were positive for B. burgdorferi s.l. An I. scapularis larva was also collected from a deer mouse, and several unfed larvae were gathered by flagging leaf litter. Based on DNA sequencing of randomly selected Borrelia amplicons from six nymphal and adult I. scapularis ticks, primers for the flagellin (fla) and flagellin B (flaB) genes reveal the presence of B. burgdorferi sensu stricto (s.s.), a genospecies pathogenic to humans and certain domestic animals. We collected all 3 host-feeding life stages of I. scapularis in a single year, and report the northernmost established population of I. scapularis in Ontario. Corkscrew Island is hyperendemic for Lyme disease and has the highest prevalence of B. burgdorferi s.l. for any established population in Canada. Because of this very high infection prevalence, this population of I. scapularis has likely been established for decades. Of epidemiological significance, cottage owners, island visitors, outdoors enthusiasts, and medical professionals must be vigilant that B. burgdorferi s.l.-infected I. scapularis on Corkscrew Island pose a serious public health risk. PMID:27877080

Established Population of Blacklegged Ticks with High Infection Prevalence for the Lyme Disease Bacterium, Borrelia burgdorferi Sensu Lato, on Corkscrew Island, Kenora District, Ontario.

PubMed

Scott, John D; Foley, Janet E; Clark, Kerry L; Anderson, John F; Durden, Lance A; Manord, Jodi M; Smith, Morgan L

2016-01-01

We document an established population of blacklegged ticks, Ixodes scapularis , on Corkscrew Island, Kenora District, Ontario, Canada. Primers of the outer surface protein A ( OspA ) gene, the flagellin ( fla ) gene, and the flagellin B ( flaB ) gene were used in the PCR assays to detect Borrelia burgdorferi sensu lato (s.l.), the Lyme disease bacterium. In all, 60 (73%) of 82 adult I. scapularis , were infected with B. burgdorferi s.l. As well, 6 (43%) of 14 unfed I. scapularis nymphs were positive for B. burgdorferi s.l. An I. scapularis larva was also collected from a deer mouse, and several unfed larvae were gathered by flagging leaf litter. Based on DNA sequencing of randomly selected Borrelia amplicons from six nymphal and adult I. scapularis ticks, primers for the flagellin ( fla ) and flagellin B ( flaB ) genes reveal the presence of B. burgdorferi sensu stricto (s.s.), a genospecies pathogenic to humans and certain domestic animals. We collected all 3 host-feeding life stages of I. scapularis in a single year, and report the northernmost established population of I. scapularis in Ontario. Corkscrew Island is hyperendemic for Lyme disease and has the highest prevalence of B. burgdorferi s.l. for any established population in Canada. Because of this very high infection prevalence, this population of I. scapularis has likely been established for decades. Of epidemiological significance, cottage owners, island visitors, outdoors enthusiasts, and medical professionals must be vigilant that B. burgdorferi s.l.-infected I. scapularis on Corkscrew Island pose a serious public health risk.

Occurrence of Borrelia burgdorferi Sensu Lato in Ixodes ricinus Ticks with First Identification of Borrelia miyamotoi in Vojvodina, Serbia.

PubMed

Potkonjak, Aleksandar; Kleinerman, Gabriela; Gutiérrez, Ricardo; Savić, Sara; Vračar, Vuk; Nachum-Biala, Yaarit; Jurišić, Aleksandar; Rojas, Alicia; Petrović, Aleksandra; Ivanović, Ivana; Harrus, Shimon; Baneth, Gad

2016-10-01

Lyme borreliosis is the most common tick-borne infectious disease in Eurasia. Borrelia miyamotoi is the only known relapsing fever Borrelia group spirochete transmitted by Ixodes species. The aim of this study was to investigate the presence of Lyme Borrelia spp. and relapsing fever Borrelia spp. in Ixodes ricinus ticks collected from dogs and the vegetation from different parts of Vojvodina, Serbia. A total of 71 Ixodes ricinus ticks were collected and screened for the presence of Lyme Borrelia spp. group and relapsing fever Borrelia spp. by real-time PCR for the Borrelia flagellin B (flaB) gene followed by DNA sequencing of PCR products. Species identification was verified by PCR of the outer surface protein A (ospA) gene for Lyme Disease Borrelia spp. and by PCR of the glycerophosphodiester phosphodiesterase (glpQ) gene for relapsing fever Borrelia spp. Lyme Borrelia spp. were found in 15/71 (21.13%) of the ticks evaluated and included B. luisitaniae (11.3%), B. afzelii (7%), B. valaisiana (1.4%), and B. garinii (1.4%). Borrelia miyamotoi, from the relapsing fever Borrelia complex, was found, for the first time in Serbia, in one (1.4%) nymph collected from the environment. Co-infections between Borrelia species in ticks were not detected. These results suggest that the dominance of species within B. burgdorferi s.l. complex in I. ricinus ticks may vary over time and in different geographic regions. Further systematic studies of Borrelia species in vectors and reservoir hosts are needed to understand eco-epidemiology of these zoonotic infections and how to prevent human infection in the best way.

Marine/Continental History of Aerosols at San Nicolas Island During CEWCOM-78 and OSP III. Summary.

DTIC Science & Technology

1979-04-01

important because of their scat- tering and aborption effects on energy. To determine the repre- sentativeneF.- of aerosol conditions encountered at SNI to...Systems (o m inand Headquarters SIh:lhU;i rIcrs Washington, I)C 20360 Washintrton, I’ 2 .3* Attn: FLEX 035 (R. Golding ) 1 Attn NA\\SI..A-(3 iS. Marcusi

Divertor Heat Flux Reduction and Detachment in the National Spherical Torus eXperiment.

NASA Astrophysics Data System (ADS)

Soukhanovskii, Vsevolod

2007-11-01

Steady-state handling of the heat flux is a critical divertor issue for both the International Thermonuclear Experimental Reactor and spherical torus (ST) devices. Because of an inherently compact divertor, it was thought that ST-based devices might not be able to fully utilize radiative and dissipative divertor techniques based on induced power and momentum loss. However, initial experiments conducted in the National Spherical Torus Experiment in an open geometry horizontal carbon plate divertor using 0.8 MA 2-6 MW NBI-heated lower single null H-mode plasmas at the lower end of elongations κ=1.8-2.4 and triangularities δ=0.45-0.75 demonstrated that high divertor peak heat fluxes, up to 6-10 MW/ m^2, could be reduced by 50-75% using a high-recycling radiative divertor regime with D2 injection. Furthermore, similar reduction was obtained with a partially detached divertor (PDD) at high D2 injection rates, however, it was accompanied by an X-point MARFE that quickly led to confinement degradation. Another approach takes advantage of the ST relation between strong shaping and high performance, and utilizes the poloidal magnetic flux expansion in the divertor region. Up to 60 % reduction in divertor peak heat flux was achieved at similar levels of scrape-off layer power by varying plasma shaping and thereby increasing the outer strike point (OSP) poloidal flux expansion from 4-6 to 18-22. In recent experiments conducted in highly-shaped 1.0-1.2 MA 6 MW NBI heated H-mode plasmas with divertor D2 injection at rates up to 10^22 s-1, a PDD regime with OSP peak heat flux 0.5-1.5 MW/m^2 was obtained without noticeable confinement degradation. Calculations based on a two point scrape-off layer model with parameterized power and momentum losses show that the short parallel connection length at the OSP sets the upper limit on the radiative exhaust channel, and both the impurity radiation and large momentum sink achievable only at high divertor neutral pressures are required for detachment.

Evidence and modeling of 3D divertor footprint induced by lower hybrid waves on EAST with tungsten divertor operations

NASA Astrophysics Data System (ADS)

Feng, W.; Wang, L.; Rack, M.; Liang, Y.; Guo, H. Y.; Xu, G. S.; Xu, J. C.; Liu, J. B.; Sun, Y. W.; Jia, M. N.; Yang, Q. Q.; Zhang, B.; Zou, X. L.; Liu, H.; Zhang, T.; Ding, F.; Chen, J. B.; Duan, Y. M.; Zheng, X. W.; Dai, S. Y.; Deng, G. Z.; Chen, R.; Hu, G. H.; Yan, N.; Si, H.; Liu, S. C.; Xu, S.; Wang, M.; Li, M. H.; Ding, B. J.; Wingen, A.; Huang, J.; Gao, X.; Luo, G. N.; Gong, X. Z.; Garofalo, A. M.; Li, J.; Wan, B. N.; the EAST Team

2017-12-01

Three dimensional (3D) divertor particle flux footprints induced by the lower hybrid wave (LHW) have been systematically investigated in the EAST superconducting tokamak during the recent experimental campaign. We find that the striated particle flux (SPF) peaks away from the strike point (SP) closely fit the pitch of the edge magnetic field line for different safety factors q 95, as predicted by a field line tracing code taking into account the helical current filaments (HCFs) in the scrape-off-layer (SOL). As LHW power increases, it requires the fuelling to be increased e.g. by super molecular beam injection (SMBI), to maintain a similar plasma density, which may be attributed to the pump-out effect due to LHW, and may thus be beneficial for EAST steady state operations. The 3D SPF structure is observed with a LHW power threshold (P LHW ~ 0.9 MW). The ratio of the particle fluxes between SPF and outer strike point (OSP), i.e. {{Γ }ion,SPF}/{{Γ }ion,OSP} , increases with the LHW power. Upon transition to divertor detachment, the particle flux at the main OSP decreases, as expected, however, the particle flux at SPF continues increasing, in contrast to the RMP-induced striations that vanish with increasing divertor density. In addition, we also find that the in-out asymmetry of the 3D particle flux footprint pattern exhibits a clear dependence on the toroidal field direction (B  ×    ∇   B  ↓  and B  ×    ∇   B↑). Experiments using neon impurity seeding show a promising capability in 3D particle and heat flux control on EAST. LHW-induced particle and heat flux striations are also present in the H-mode plasmas, reducing the peak heat flux and erosion at the main strike point, thus facilitating long-pulse operation with a new steady-state H-mode over 60 s being recently achieved in EAST.

Effects of PCB Substrate Surface Finish, Flux, and Phosphorus Content on Ionic Contamination

NASA Astrophysics Data System (ADS)

Bacior, M.; Sobczak, N.; Siewiorek, A.; Kudyba, A.; Homa, M.; Nowak, R.; Dziula, M.; Masłoń, S.

2015-02-01

The ionic contamination on printed circuit boards (PCB) having different surface finishes was examined using ionograph. The study was performed at the RT on three types of PCBs covered with: (i) hot air solder leveling (HASL LF), (ii) electroless nickel immersion gold (ENIG), and (iii) organic surface protectant (OSP), all on Cu substrates, as well as two types of fluxes, namely EF2202 and RF800. In the group of boards without soldered components, the lowest average value of contamination was for the ENIG 18 µm surface (0.01 μg NaCl/cm2). Boards with soldered components were more contaminated (from 0.29 μg NaCl/cm2 for the HASL LF 18 µm surface). After spraying boards with fluxing agents, the values of contaminants were the highest. The influence of phosphorus content in Ni-P layer of ENIG finish on ionic contamination was examined. In the group of PCBs with Au coating, the smallest amount of surface contaminants (0.32 μg NaCl/cm2) was for Ni-2-5%P layer. PCBs with Ni-11%P layer were higher contaminated (0.47 μg NaCl/cm2), and another with Ni-8%P layer had 0.81 μg NaCl/cm2. PCBs without Au coating, had the lowest contamination (0.48 μg NaCl/cm2) at phosphorous content equal 11%P. Higher contamination (0.67 μg NaCl/cm2) was at 2-5%P, up to 1.98 μg NaCl/cm2 for 8% of P. Boards with Au finish have lower value of contamination than identical boards without Au layer thus contributing to better reliability of electronic assemblies, since its failures due to current leakage and corrosion can be caused by contaminants.

District of Columbia Opportunity Scholarship Program: Additional Policies and Procedures Would Improve Internal Controls and Program Operations. Report to Congressional Requesters. GAO-08-9

ERIC Educational Resources Information Center

Ashby, Cornelia M.; Franzel, Jeanette M.

2007-01-01

The D.C. School Choice Incentive Act created the first private kindergarten-through-grade-12 school-choice program supported by federal funds. The program was named the D.C. Opportunity Scholarship Program (OSP). The United States Government Accountability Office (GAO) was asked to assess the (1) accountability mechanisms governing the use of…

Blocking pathogen transmission at the source: reservoir targeted OspA-based vaccines against Borrelia burgdorferi.

PubMed

Gomes-Solecki, Maria

2014-01-01

Control strategies are especially challenging for microbial diseases caused by pathogens that persist in wildlife reservoirs and use arthropod vectors to cycle amongst those species. One of the most relevant illnesses that pose a direct human health risk is Lyme disease; in the US, the Centers for Disease Control and Prevention recently revised the probable number of cases by 10-fold, to 300,000 cases per year. Caused by Borrelia burgdorferi, Lyme disease can affect the nervous system, joints and heart. No human vaccine is approved by the Food and Drug Administration. In addition to novel human vaccines, new strategies for prevention of Lyme disease consist of pest management interventions, vector-targeted vaccines and reservoir-targeted vaccines. However, even human vaccines can not prevent Lyme disease expansion into other geographical areas. The other strategies aim at reducing tick density and at disrupting the transmission of B. burgdorferi by targeting one or more key elements that maintain the enzootic cycle: the reservoir host and/or the tick vector. Here, I provide a brief overview of the application of an OspA-based wildlife reservoir targeted vaccine aimed at reducing transmission of B. burgdorferi and present it as a strategy for reducing Lyme disease risk to humans.

Orbital Space Plane (OSP) Program

NASA Technical Reports Server (NTRS)

McKenzie, Patrick M.

2003-01-01

Lockheed Martin has been an active participant in NASA's Space Launch Initiative (SLI) programs over the past several years. SLI, part of NASA's Integrated Space Transportation Plan (ISTP), was restructured in November of 2002 to focus the overall theme of safer, more afford-able space transportation along two paths - the Orbital Space Plane Program and the Next Generation Launch Technology programs. The Orbital Space Plane Program has the goal of providing rescue capability from the International Space Station by 2008 and transfer capability for crew (and limited cargo) by 2012. The Next Generation Launch Technology program is combining research and development efforts from the 2nd Generation Reusable Launch Vehicle (2GRLV) program with cutting-edge, advanced space transportation programs (previously designated 3rd Generation) into one program aimed at enabling safe, reliable, cost-effective reusable launch systems by the middle of the next decade. Lockheed Martin is one of three prime contractors working to bring Orbital Space Plane system concepts to a system definition level of maturity by December of 2003. This paper and presentation will update the international community on the progress of the' OSP program, from an industry perspective, and provide insights into Lockheed Martin's role in enabling the vision of a safer, more affordable means of taking people to and from space.

Blocking pathogen transmission at the source: reservoir targeted OspA-based vaccines against Borrelia burgdorferi

PubMed Central

Gomes-Solecki, Maria

2014-01-01

Control strategies are especially challenging for microbial diseases caused by pathogens that persist in wildlife reservoirs and use arthropod vectors to cycle amongst those species. One of the most relevant illnesses that pose a direct human health risk is Lyme disease; in the US, the Centers for Disease Control and Prevention recently revised the probable number of cases by 10-fold, to 300,000 cases per year. Caused by Borrelia burgdorferi, Lyme disease can affect the nervous system, joints and heart. No human vaccine is approved by the Food and Drug Administration. In addition to novel human vaccines, new strategies for prevention of Lyme disease consist of pest management interventions, vector-targeted vaccines and reservoir-targeted vaccines. However, even human vaccines can not prevent Lyme disease expansion into other geographical areas. The other strategies aim at reducing tick density and at disrupting the transmission of B. burgdorferi by targeting one or more key elements that maintain the enzootic cycle: the reservoir host and/or the tick vector. Here, I provide a brief overview of the application of an OspA-based wildlife reservoir targeted vaccine aimed at reducing transmission of B. burgdorferi and present it as a strategy for reducing Lyme disease risk to humans. PMID:25309883

Site Safety Plan for Lawrence Livermore National Laboratory CERCLA investigations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bainer, R.; Duarte, J.

1993-07-01

The safety policy of LLNL is to take every reasonable precaution in the performance of work to protect the environment and the health and safety of employees and the public, and to prevent property damage. With respect to hazardous agents, this protection is provided by limiting human exposures, releases to the environment, and contamination of property to levels that are as low as reasonably achievable (ALARA). It is the intent of this Plan to supply the broad outline for completing environmental investigations within ALARA guidelines. It may not be possible to determine actual working conditions in advance of the work;more » therefore, planning must allow the opportunity to provide a range of protection based upon actual working conditions. Requirements will be the least restrictive possible for a given set of circumstances, such that work can be completed in an efficient and timely fashion. Due to the relatively large size of the LLNL Site and the different types of activities underway, site-specific Operational Safety Procedures (OSPs) will be prepared to supplement activities not covered by this Plan. These site-specific OSPs provide the detailed information for each specific activity and act as an addendum to this Plan, which provides the general plan for LLNL Main Site operation.« less

Minimal unitary representation of SO∗(8)=SO(6,2) and its SU(2) deformations as massless 6D conformal fields and their supersymmetric extensions

NASA Astrophysics Data System (ADS)

Fernando, Sudarshan; Günaydin, Murat

2010-12-01

We study the minimal unitary representation (minrep) of SO(6,2) over an Hilbert space of functions of five variables, obtained by quantizing its quasiconformal realization. The minrep of SO(6,2), which coincides with the minrep of SO(8) similarly constructed, corresponds to a massless conformal scalar field in six spacetime dimensions. There exists a family of "deformations" of the minrep of SO(8) labeled by the spin t of an SU(2 subgroup of the little group SO(4) of lightlike vectors. These deformations labeled by t are positive energy unitary irreducible representations of SO(8) that describe massless conformal fields in six dimensions. The SU(2 spin t is the six-dimensional counterpart of U(1) deformations of the minrep of 4D conformal group SU(2,2) labeled by helicity. We also construct the supersymmetric extensions of the minimal unitary representation of SO(8) to minimal unitary representations of OSp(8|2N) that describe massless six-dimensional conformal supermultiplets. The minimal unitary supermultiplet of OSp(8|4) is the massless supermultiplet of (2,0) conformal field theory that is believed to be dual to M-theory on AdS×S.

Effect of Industrial By-Products on Unconfined Compressive Strength of Solidified Organic Marine Clayey Soils

PubMed Central

Park, Chan-Gi; Yun, Sung-Wook; Baveye, Phillippe C.; Yu, Chan

2015-01-01

The use of industrial by-products as admixture to ASTM Type I cement (ordinary Portland cement (OPC)) was investigated with the objective of improving the solidification of organic marine clayey soils. The industrial by-products considered in this paper were oyster-shell powder (OSP), steelmaking slag dust (SMS) and fuel-gas-desulfurized (FGD) gypsum. The industrial by-products were added to OPC at a ratio of 5% based on dry weight to produce a mixture used to solidify organic marine clayey soils. The dosage ratios of mixtures to organic marine clayey soils were 5, 10 and 15% on a dry weight basis. Unconfined compressive strength (UCS) test after 28 days revealed that the highest strength was obtained with the OPC + SMS 15% mixing ratio. The UCS of specimens treated with this mixture was >500 kPa, compared with 300 kPa for specimens treated with a 15% OPC + OSP mixture and 200 kPa when 15% of OPC was used alone. These results were attributed to the more active hydration and pozzolanic reaction of the OPC + SMS mixture. This hypothesis was verified through X-ray diffraction (XRD) and scanning electron microscopy (SEM) analyses, and was confirmed by variations in the calcium carbonate (CaCO3) content of the materials during curing. PMID:28793493

Understanding tungsten divertor sourcing and SOL transport using multiple poloidally-localized sources in DIII-D ELM-y H-mode discharges

NASA Astrophysics Data System (ADS)

Unterberg, Ea; Donovan, D.; Barton, J.; Wampler, Wr; Abrams, T.; Thomas, Dm; Petrie, T.; Guo, Hy; Stangeby, Pg; Elder, Jd; Rudakov, D.; Grierson, B.; Victor, B.

2017-10-01

Experiments using metal inserts with novel isotopically-enriched tungsten coatings at the outer divertor strike point (OSP) have provided unique insight into the ELM-induced sourcing, main-SOL transport, and core accumulation control mechanisms of W for a range of operating conditions. This experimental approach has used a multi-head, dual-facing collector probe (CP) at the outboard midplane, as well as W-I and core W spectroscopy. Using the CP system, the total amount of W deposited relative to source measurements shows a clear dependence on ELM size, ELM frequency, and strike point location, with large ELMs depositing significantly more W on the CP from the far-SOL source. Additionally, high spatial ( 1mm) and ELM resolved spectroscopic measurements of W sourcing indicate shifts in the peak erosion rate. Furthermore, high performance discharges with rapid ELMs show core W concentrations of few 10-5, and the CP deposition profile indicates W is predominantly transported to the midplane from the OSP rather than from the far-SOL region. The low central W concentration is shown to be due to flattening of the main plasma density profile, presumably by on-axis electron cyclotron heating. Work supported under USDOE Cooperative Agreement DE-FC02-04ER54698.

Prevalence of Borrelia miyamotoi and Borrelia burgdorferi sensu lato in questing ticks from a recreational coniferous forest of East Saxony, Germany.

PubMed

Szekeres, Sándor; Lügner, Jenny; Fingerle, Volker; Margos, Gabriele; Földvári, Gábor

2017-10-01

The hard tick Ixodes ricinus is the most important vector of tick-transmitted pathogens in Europe, frequently occurring in urban parks and greenbelts utilized for recreational activities. This species is the most common vector of the causative agents of Lyme borreliosis in Europe. Similarly, the species spreads Borrelia miyamotoi, causing a relapsing-fever like illness. A total of 1774 Ixodes ricinus (50 females, 68 males, 840 nymphs and 818 larvae) were collected with flagging between March and September 2014 in a coniferous forest patch in Niederkaina near the town of Bautzen in Saxony, Germany. To measure questing tick density a time-based density estimating method was utilized. From each month, a total of 100 adults and nymphal ticks and all larvae (pools of 10 individuals per tube/month) were selected for the molecular analyses. For simultaneous detection of B. burgdorferi s.l. and B. miyamotoi a duplex real-time PCR targeting the flaB locus was performed. Prevalence of B. burgdorferi s.l. was 9.4% (female: 6%, male: 2.9%, nymph: 12.2%, larva: 0%) and minimum prevalence of B. miyamotoi was 1.2% (female: 0%, male: 4.3%, nymph: 2.8%, larva: 0.1%) in the 714 samples with real-time polymerase chain reaction. A real-time PCR reaction was utilized first to target the histone-like protein gene (hbb) of B. burgdorferi s.l., a hemi-nested outer surface protein (ospA) gene conventional PCR was then performed followed by a restriction enzyme analysis to distinguish B. burgdorferi s.l. genospecies. Seven B. afzelii, one B. burgdorferi s.s., one B. bavariensis and four B. miyamotoi infections were confirmed. Prevalence of Lyme borreliosis spirochetes was significantly higher in nymphs than in adults (p<0.01, Fisher exact test) probably due to the diluting effect of the local roe deer population. Our data highlight the potential risk of human infection with the emerging pathogen B. miyamotoi within the study area. Copyright © 2017 Elsevier GmbH. All rights reserved.

Reclassification of Borrelia spp. isolated in South Korea using Multilocus Sequence Typing.

PubMed

Park, Kyung-Hee; Choi, Yeon-Joo; Kim, Jeoungyeon; Park, Hye-Jin; Song, Dayoung; Jang, Won-Jong

2018-05-31

Using Borrelia isolated from South Korea, we evaluated by MLST and three intergenic genes (16S rRNA, ospA, and 5S-23S IGS) typing to analyze the relationship between host and vector and molecular background. Using the MLST analysis, we identified B. afzelii, B. yangtzensis, B. garinii, and B. bavariensis. This study was first report of the identification of B. yangtzensis using the MLST in South Korea.

Evaluation of Selected Borrelia burgdorferi lp54 Plasmid-Encoded Gene Products Expressed during Mammalian Infection as Antigens To Improve Serodiagnostic Testing for Early Lyme Disease

PubMed Central

Weiner, Zachary P.; Crew, Rebecca M.; Brandt, Kevin S.; Ullmann, Amy J.; Schriefer, Martin E.; Molins, Claudia R.

2015-01-01

Laboratory testing for the diagnosis of Lyme disease is performed primarily by serologic assays and is accurate for detection beyond the acute stage of the infection. Serodiagnostic assays to detect the early stages of infection, however, are limited in their sensitivity, and improvement is warranted. We analyzed a series of Borrelia burgdorferi proteins known to be induced within feeding ticks and/or during mammalian infection for their utility as serodiagnostic markers against a comprehensive panel of Lyme disease patient serum samples. The antigens were assayed for IgM and IgG reactivity in line immunoblots and separately by enzyme-linked immunosorbent assay (ELISA), with a focus on reactivity against early Lyme disease with erythema migrans (EM), early disseminated Lyme neuroborreliosis, and early Lyme carditis patient serum samples. By IgM immunoblotting, we found that recombinant proteins BBA65, BBA70, and BBA73 reacted with early Lyme EM samples at levels comparable to those of the OspC antigen used in the current IgM blotting criteria. Additionally, these proteins reacted with serum samples from patients with early neuroborreliosis and early carditis, suggesting value in detecting early stages of this disease progression. We also found serological reactivity against recombinant proteins BBA69 and BBA73 with early-Lyme-disease samples using IgG immunoblotting and ELISA. Significantly, some samples that had been scored negative by the Centers for Disease Control and Prevention-recommended 2-tiered testing algorithm demonstrated positive reactivity to one or more of the antigens by IgM/IgG immunoblot and ELISA. These results suggest that incorporating additional in vivo-expressed antigens into the current IgM/IgG immunoblotting tier in a recombinant protein platform assay may improve the performance of early-Lyme-disease serologic testing. PMID:26376927

Evaluation of Selected Borrelia burgdorferi lp54 Plasmid-Encoded Gene Products Expressed during Mammalian Infection as Antigens To Improve Serodiagnostic Testing for Early Lyme Disease.

PubMed

Weiner, Zachary P; Crew, Rebecca M; Brandt, Kevin S; Ullmann, Amy J; Schriefer, Martin E; Molins, Claudia R; Gilmore, Robert D

2015-11-01

Laboratory testing for the diagnosis of Lyme disease is performed primarily by serologic assays and is accurate for detection beyond the acute stage of the infection. Serodiagnostic assays to detect the early stages of infection, however, are limited in their sensitivity, and improvement is warranted. We analyzed a series of Borrelia burgdorferi proteins known to be induced within feeding ticks and/or during mammalian infection for their utility as serodiagnostic markers against a comprehensive panel of Lyme disease patient serum samples. The antigens were assayed for IgM and IgG reactivity in line immunoblots and separately by enzyme-linked immunosorbent assay (ELISA), with a focus on reactivity against early Lyme disease with erythema migrans (EM), early disseminated Lyme neuroborreliosis, and early Lyme carditis patient serum samples. By IgM immunoblotting, we found that recombinant proteins BBA65, BBA70, and BBA73 reacted with early Lyme EM samples at levels comparable to those of the OspC antigen used in the current IgM blotting criteria. Additionally, these proteins reacted with serum samples from patients with early neuroborreliosis and early carditis, suggesting value in detecting early stages of this disease progression. We also found serological reactivity against recombinant proteins BBA69 and BBA73 with early-Lyme-disease samples using IgG immunoblotting and ELISA. Significantly, some samples that had been scored negative by the Centers for Disease Control and Prevention-recommended 2-tiered testing algorithm demonstrated positive reactivity to one or more of the antigens by IgM/IgG immunoblot and ELISA. These results suggest that incorporating additional in vivo-expressed antigens into the current IgM/IgG immunoblotting tier in a recombinant protein platform assay may improve the performance of early-Lyme-disease serologic testing. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Applying to the DC Opportunity Scholarship Program: How Do Parents Rate Their Children's Current Schools at Time of Application and What Do They Want in New Schools? NCEE Evaluation Brief. NCEE 2016-4003

ERIC Educational Resources Information Center

Dynarski, Mark; Betts, Julian; Feldman, Jill

2016-01-01

The DC Opportunity Scholarship Program (OSP), established in 2004, is the only federally-funded private school voucher program for low-income parents in the United States. This evaluation brief describes findings using data from more than 2,000 applicants' parents, who applied to the program from spring 2011 to spring 2013 following…

Measurements and modeling of intra-ELM tungsten sourcing and transport in DIII-D

NASA Astrophysics Data System (ADS)

Abrams, T.; Leonard, A. W.; Thomas, D. M.; McLean, A. G.; Makowski, M. A.; Wang, H. Q.; Unterberg, E. A.; Briesemeister, A. R.; Rudakov, D. L.; Bykov, I.; Donovan, D.

2017-10-01

Intra-ELM tungsten erosion profiles in the DIII-D divertor, acquired via W I spectroscopy with high temporal and spatial resolution, are consistent with SDTrim.SP sputtering modeling using measured ion saturation currents and impact energies during ELMs as input and an ad-hoc 2% C2+ impurity flux. The W sputtering profile peaks close to the OSP both during and between ELMs in the favorable BT direction. In reverse BT the W source peaks close to the OSP between ELMs but strongly broadens and shifts outboard during ELMs, heuristically consistent with radially outward ion transport via ExB drifts. Ion impact energies during ELMs (inferred taking the ratio of divertor heat flux to the ion saturation current) are found to be approximately equal to Te,ped, lower than the 4*Te,ped value predicted by the Fundamenski/Moulton free streaming model. These impact energies imply both D main ions and C impurities contribute strongly to W sputtering during ELMs on DIII-D. This work represents progress towards a predictive model to link upstream conditions (i.e., pedestal height) and SOL impurity levels to the ELM-induced W impurity source at both the strike-point and far-target regions in the ITER divertor. Correlations between ELM size/frequency and SOL W fluxes measured via a midplane deposition probe will also be presented. Work supported by US DOE under DE-FC02-04ER54698.

RMP effects on the W and C erosion/deposition balance on W test samples in DIII-D

NASA Astrophysics Data System (ADS)

Hinson, E. T.; Frerichs, H.; Schmitz, O.; Evans, T. E.; Guo, H. Y.; Thomas, D. M.; Rudakov, D. L.; Abrams, T.; Unterberg, E. A.; Briesemeister, A.; Lasnier, C. J.; McLean, A. G.; Makowski, M.; Wampler, W. R.; Watkins, J. G.; Wang, H. Q.

2016-10-01

Clear evidence for alteration of the W and C erosion by resonant magnetic perturbation (RMP) fields has been obtained in an experiment exposing W-coated DiMES samples in the DIII-D divertor to outer strike point (OSP) sweeps in comparable series of discharges with and without the application of RMP. Gross erosion measurements of W and C during these sweeps using the S/XB method show that the 3-D boundary induced by the RMP significantly alters the erosion rate from DiMES. In particular, application of RMP smooths radial W erosion anisotropy seen for the axisymmetric case, where the W erosion rate for the OSP sweep in the outward direction significantly exceeds the erosion rate observed for the subsequent inward radial sweep over the sample. This finding is likely related to a change in the W/C erosion and redeposition balance in the C-dominated wall environment at DIII-D. Moreover, non-axisymmetric plasma structure on the W sample has to be considered. This challenge will be further examined by comparison of experimental results to EMC3-EIRENE modeling. Work supported by US DOE DE-SC0013911, DE-FC02-04ER54698, DE-FH02-07ER54917, DE-AC05-06OR23100, DE-AC05-00OR22725, DE-AC52-07NA27344, and DE-AC04-94AL85000.

The suitability of EIT to estimate EELV in a clinical trial compared to oxygen wash-in/wash-out technique.

PubMed

Karsten, Jan; Meier, Torsten; Iblher, Peter; Schindler, Angela; Paarmann, Hauke; Heinze, Hermann

2014-02-01

Open endotracheal suctioning procedure (OSP) and recruitment manoeuvre (RM) are known to induce severe alterations of end-expiratory lung volume (EELV). We hypothesised that EIT lung volumes lack clinical validity. We studied the suitability of EIT to estimate EELV compared to oxygen wash-in/wash-out technique. Fifty-four postoperative cardiac surgery patients were enrolled and received standardized ventilation and OSP. Patients were randomized into two groups receiving either RM after suctioning (group RM) or no RM (group NRM). Measurements were conducted at the following time points: Baseline (T1), after suctioning (T2), after RM or NRM (T3), and 15 and 30 min after T3 (T4 and T5). We measured EELV using the oxygen wash-in/wash-out technique (EELVO2) and computed EELV from EIT (EELVEIT) by the following formula: EELVEITTx,y…=EELVO2+ΔEELI×VT/ΔZ. EELVEIT values were compared with EELVO2 using Bland-Altman analysis and Pearson correlation. Limits of agreement ranged from -0.83 to 1.31 l. Pearson correlation revealed significant results. There was no significant impact of RM or NRM on EELVO2-EELVEIT relationship (p=0.21; p=0.23). During typical routine respiratory manoeuvres like endotracheal suctioning or alveolar recruitment, EELV cannot be estimated by EIT with reasonable accuracy.

Relationships between spray parameters, microstructures and ultrasonic cavitation erosion behavior of HVOF sprayed Fe-based amorphous/nanocrystalline coatings.

PubMed

Qiao, Lei; Wu, Yuping; Hong, Sheng; Zhang, Jianfeng; Shi, Wei; Zheng, Yugui

2017-11-01

Fe-based amorphous/nanocrystalline coatings were prepared on the AISI 321 steel substrate by the high-velocity oxygen-fuel (HVOF) thermal spraying technology. The effect of selected parameters (oxygen flow, kerosene flow and spray distance) on the cavitation erosion resistance (denoted as Rc) of the coating were investigated by using the Taguchi method. Statistical tools such as design of experiments (DOE), signal-to-noise (S/N) ratio and analysis of variance (ANOVA) were used to meet the expected objective. It was concluded that the kerosene flow had greater influence on the Rc of the coating and followed by the spray distance and the oxygen flow, respectively. The optimum spray parameters (OSP) were 963L/min for the oxygen flow, 28L/h for the kerosene flow, and 330mm for the spray distance. The Rc of the coating increased with the increase of hardness or the decrease of porosity, and the hardness had a greater influence on Rc than the porosity. The Fe-based coating deposited under the OSP exhibited the best cavitation erosion resistance in distilled water. The cracks initiated at the edge of the pores and the interfaces between the un-melted or half-melted particles, and finally leaded to the delamination of the coating. Copyright © 2017 Elsevier B.V. All rights reserved.

Measuring the Effects of an Ever-Changing Environment on Malaria Control

DTIC Science & Technology

2004-04-01

immunity on improved protection has been described in relationship to vaccine trials against Lyme disease (5). Herd immunity has been shown to play a...rural northern Ghana. Am. J. Trop. Med. Hyg. 59:80–85. 5. de Silva, A. M., S. R. Telford III, L. R. Brunet, S. W. Barthold, and E. Fikrig. 1996. Borrelia ...burgdorferi OspA is an arthropod-specific transmission-block- ing Lyme disease vaccine. Exp. Med. 183:271–275. 6. Diallo, D. A., A. Habluetzel, N

The Solution Construction of Heterotic Super-Liouville Model

NASA Astrophysics Data System (ADS)

Yang, Zhan-Ying; Zhen, Yi

2001-12-01

We investigate the heterotic super-Liouville model on the base of the basic Lie super-algebra Osp(1|2).Using the super extension of Leznov-Saveliev analysis and Drinfeld-Sokolov linear system, we construct the explicit solution of the heterotic super-Liouville system in component form. We also show that the solutions are local and periodic by calculating the exchange relation of the solution. Finally starting from the action of heterotic super-Liouville model, we obtain the conserved current and conserved charge which possessed the BRST properties.

Basic echocardiography for undergraduate students: a comparison of different peer-teaching approaches.

PubMed

Gradl-Dietsch, G; Menon, A K; Gürsel, A; Götzenich, A; Hatam, N; Aljalloud, A; Schrading, S; Hölzl, F; Knobe, M

2018-02-01

The aim of this study was to assess the impact of different teaching interventions in a peer-teaching environment on basic echocardiography skills and to examine the influence of gender on learning outcomes. We randomly assigned 79 s year medical students (55 women, 24 men) to one of four groups: peer teaching (PT), peer teaching using Peyton's four-step approach (PPT), team based learning (TBL) and video-based learning (VBL). All groups received theoretical and practical hands-on training according to the different approaches. Using a pre-post-design we assessed differences in theoretical knowledge [multiple choice (MC) exam], practical skills (Objective Structured Practical Examination, OSPE) and evaluation results with respect to gender. There was a significant gain in theoretical knowledge for all students. There were no relevant differences between the four groups regarding the MC exam and OSPE results. The majority of students achieved good or very good results. Acceptance of the peer-teaching concept was moderate and all students preferred medical experts to peer tutors even though the overall rating of the instructors was fairly good. Students in the Video group would have preferred a different training method. There was no significant effect of gender on evaluation results. Using different peer-teaching concepts proved to be effective in teaching basic echocardiography. Gender does not seem to have an impact on effectiveness of the instructional approach. Qualitative analysis revealed limited acceptance of peer teaching and especially of video-based instruction.

More on the hidden symmetries of 11D supergravity

NASA Astrophysics Data System (ADS)

Andrianopoli, Laura; D'Auria, Riccardo; Ravera, Lucrezia

2017-09-01

In this paper we clarify the relations occurring among the osp (1 | 32) algebra, the M-algebra and the hidden superalgebra underlying the Free Differential Algebra of D=11 supergravity (to which we will refer as DF-algebra) that was introduced in the literature by D'Auria and Frè in 1981 and is actually a (Lorentz valued) central extension of the M-algebra including a nilpotent spinor generator, Q‧. We focus in particular on the 4-form cohomology in 11D superspace of the supergravity theory, strictly related to the presence in the theory of a 3-form A (3). Once formulated in terms of its hidden superalgebra of 1-forms, we find that A (3) can be decomposed into the sum of two parts having different group-theoretical meaning: One of them allows to reproduce the FDA of the 11D Supergravity due to non-trivial contributions to the 4-form cohomology in superspace, while the second one does not contribute to the 4-form cohomology, being a closed 3-form in the vacuum, defining however a one parameter family of trilinear forms invariant under a symmetry algebra related to osp (1 | 32) by redefining the spin connection and adding a new Maurer-Cartan equation. We further discuss about the crucial role played by the 1-form spinor η (dual to the nilpotent generator Q‧) for the 4-form cohomology of the eleven dimensional theory on superspace.

What controls the variability of oxygen in the subpolar North Pacific?

NASA Astrophysics Data System (ADS)

Takano, Yohei

Dissolved oxygen is a widely observed chemical quantity in the oceans along with temperature and salinity. Changes in the dissolved oxygen have been observed over the world oceans. Observed oxygen in the Ocean Station Papa (OSP, 50°N, 145°W) in the Gulf of Alaska exhibits strong variability over interannual and decadal timescales, however, the mechanisms driving the observed variability are not yet fully understood. Furthermore, irregular sampling frequency and relatively short record length make it difficult to detect a low-frequency variability. Motivated by these observations, we investigate the mechanisms driving the low-frequency variability of oxygen in the subpolar North Pacific. The specific purposes of this study are (1) to evaluate the robustness of the observed low-frequency variability of dissolved oxygen and (2) to determine the mechanisms driving the observed variability using statistical data analysis and numerical simulations. To evaluate the robustness of the low-frequency variability, we conducted spectral analyses on the observed oxygen at OSP. To address the irregular sampling frequency we randomly sub-sampled the raw data to form 500 ensemble members with a regular time interval, and then performed spectral analyses. The resulting power spectrum of oxygen exhibits a robust low-frequency variability and a statistically significant spectral peak is identified at a timescale of 15--20 years. The wintertime oceanic barotropic streamfunction is significantly correlated with the observed oxygen anomaly at OSP with a north-south dipole structure over the North Pacific. We hypothesize that the observed low-frequency variability is primarily driven by the variability of large-scale ocean circulation in the North Pacific. To test this hypothesis, we simulate the three-dimensional distribution of oxygen anomaly between 1952 to 2001 using data-constrained circulation fields. The simulated oxygen anomaly shows an outstanding variability in the Gulf of Alaska, showing that this region is a hotspot of oxygen fluctuation. Anomalous advection acting on the climatological mean oxygen gradient is the source of oxygen variability in this simulation. Empirical Orthogonal Function (EOF) analyses of the simulated oxygen show that the two dominant modes of the oxygen anomaly explains more than 50% of oxygen variance over the North Pacific, that are closely related to the dominant modes of climate variability in the North Pacific (Pacific Decadal Oscillation and North Pacific Oscillation). Our results imply the important link between large-scale climate fluctuations, ocean circulation and biogeochemical tracers in the North Pacific.

Multistrain Infections with Lyme Borreliosis Pathogens in the Tick Vector.

PubMed

Durand, Jonas; Herrmann, Coralie; Genné, Dolores; Sarr, Anouk; Gern, Lise; Voordouw, Maarten J

2017-02-01

Mixed or multiple-strain infections are common in vector-borne diseases and have important implications for the epidemiology of these pathogens. Previous studies have mainly focused on interactions between pathogen strains in the vertebrate host, but little is known about what happens in the arthropod vector. Borrelia afzelii and Borrelia garinii are two species of spirochete bacteria that cause Lyme borreliosis in Europe and that share a tick vector, Ixodes ricinus Each of these two tick-borne pathogens consists of multiple strains that are often differentiated using the highly polymorphic ospC gene. For each Borrelia species, we studied the frequencies and abundances of the ospC strains in a wild population of I. ricinus ticks that had been sampled from the same field site over a period of 3 years. We used quantitative PCR (qPCR) and 454 sequencing to estimate the spirochete load and the strain diversity within each tick. For B. afzelii, there was a negative relationship between the two most common ospC strains, suggesting the presence of competitive interactions in the vertebrate host and possibly the tick vector. The flat relationship between total spirochete abundance and strain richness in the nymphal tick indicates that the mean abundance per strain decreases as the number of strains in the tick increases. Strains with the highest spirochete load in the nymphal tick were the most common strains in the tick population. The spirochete abundance in the nymphal tick appears to be an important life history trait that explains why some strains are more common than others in nature. Lyme borreliosis is the most common vector-borne disease in the Northern Hemisphere and is caused by spirochete bacteria that belong to the Borrelia burgdorferi sensu lato species complex. These tick-borne pathogens are transmitted among vertebrate hosts by hard ticks of the genus Ixodes Each Borrelia species can be further subdivided into genetically distinct strains. Multiple-strain infections are common in both the vertebrate host and the tick vector and can result in competitive interactions. To date, few studies on multiple-strain vector-borne pathogens have investigated patterns of cooccurrence and abundance in the arthropod vector. We demonstrate that the abundance of a given strain in the tick vector is negatively affected by the presence of coinfecting strains. In addition, our study suggests that the spirochete abundance in the tick is an important life history trait that can explain why some strains are more common in nature than others. Copyright © 2017 American Society for Microbiology.

Induction of Apg-1, a member of the heat shock protein 110 family, following transient forebrain ischemia in the rat brain.

PubMed

Xue, J H; Fukuyama, H; Nonoguchi, K; Kaneko, Y; Kido, T; Fukumoto, M; Fujibayashi, Y; Itoh, K; Fujita, J

1998-06-29

Apg-1 (Osp94) and apg-2 belong to the heat shock protein (hsp) 110 family. In mouse somatic cells the apg-1 and hsp105/110 transcripts are inducible by a 32 degrees C to 39 degrees C heat shock, while apg-2 is not heat-inducible. Since ischemia is known to induce expression of hsp70, its effect on expression of apg-1 was assessed by using the 20-min forebrain ischemia model of the rat. In the cerebral cortex, Northern blot analysis and in situ hybridization histochemistry demonstrated an increased expression in neuronal cells of apg-1 transcripts 3 h after the onset of reperfusion, with a peak at 12 h, followed by a decline. In the hippocampus, the level was increased at 3 h, remained constant until 24 h, and then declined. Transcript levels of apg-2 as well as hsp 105 were also increased under the present conditions, indicating that the expression of apg-2 was differentially regulated in response to heat and ischemic stresses. The induction kinetics of hsp 105, but neither apg-2 nor hsp 70, were identical to those of apg-1. These results demonstrated that brain ischemia/reperfusion induced expression of each member of the hsp 110 family, although the regulatory mechanisms may not be the same. They also suggest a significant role of apg-1 in both the ischemic- and heat-stress responses and in the normal functioning of the non-stressed neuronal cells.

Small Structures and Superlattices for Future High-Speed Devices

DTIC Science & Technology

1986-10-01

1985. I-5oA -undoped) [101 M. Ogawa. K. Olsata. T. Fu-usuka. and N. Kawamura . IEEE M-GoAt 1ND) _ -OSP~m Mricrowave Thie.ory Tech., vol. MTT-24, p. 3013...thicknesses range between (T.E.S.) from GTE. We also wish to thank Masako Okamoto 250 and 600 A. Forward I-V measurements that we have and Professor W. M...orientation mixed in among one of us (T.E.S.) from GTE. We also wish to thank Masako the type-B structure. This possibility is discounted by our Okamoto

IPS guidestar selection for stellar mode (ASTRO)

NASA Technical Reports Server (NTRS)

Mullins, Larry; Wooten, Lewis

1988-01-01

This report describes how guide stars are selected for the Optical Sensor Package (OSP) for the Instrument Pointing System (IPS) when it is operating in the stellar mode on the ASTRO missions. It also describes how the objective loads are written and how the various roll angles are related; i.e., the celestial roll or position angle, the objective load roll angles, and the IPS gimbal angles. There is a brief description of how the IPS operates and its various modes of operation; i.e., IDOP, IDIN, and OSPCAL.

One session treatment for specific phobias in children: Comorbid anxiety disorders and treatment outcome.

PubMed

Ryan, Sarah M; Strege, Marlene V; Oar, Ella L; Ollendick, Thomas H

2017-03-01

One-Session Treatment (OST) for specific phobias has been shown to be effective in reducing phobia severity; however, the effect of different types of co-occurring anxiety disorders on OST outcomes is unknown. The present study examined (1) the effects of co-occurring generalized anxiety disorder (GAD), social anxiety disorder (SAD), or another non-targeted specific phobia (OSP) on the efficacy of OST for specific phobias, and (2) the effects of OST on these co-occurring disorders following treatment. Three groups of 18 youth (7-15 years) with a specific phobia and comorbid GAD, SAD, or OSP were matched on age, gender, and phobia type. Outcome measures included diagnostic status and severity, and clinician rated improvement. All groups demonstrated an improvement in their specific phobia following treatment. Treatment was equally effective regardless of co-occurring anxiety disorder. In addition, comorbid anxiety disorders improved following OST; however, this effect was not equal across groups. The SAD group showed poorer improvement in their comorbid disorder than the GAD group post-treatment. However, the SAD group continued to improve and this differential effect was not evident six-months following treatment. The current study sample was small, with insufficient power to detect small and medium effect sizes. Further, the sample only included a portion of individuals with primary GAD or SAD, which may have attenuated the findings. The current study demonstrated that co-occurring anxiety disorders did not interfere with phobia treatment. OST, despite targeting a single specific phobia type, significantly reduced comorbid symptomatology across multiple anxiety disorders. Copyright © 2016 Elsevier Ltd. All rights reserved.

Temperature and slant path effects in Dobson and Brewer total ozone measurements

NASA Astrophysics Data System (ADS)

Scarnato, B.; Staehelin, J.; Peter, T.; GröBner, J.; Stübi, R.

2009-12-01

There is a worldwide tendency to replace Dobson spectrophotometers in ground-based total ozone (TOZ) measurements by more advanced Brewer spectrophotometers. Ensuring the homogeneity of these data sets is of utmost importance if changes in TOZ of a few percent over long time periods are to be diagnosed accurately. Previous studies have identified a seasonal bias of a few percent between midlatitude Brewer and Dobson measurements. At Arosa (Switzerland), two Dobson and three Brewer instruments have been colocated since 1998, providing a unique data set of quasi-simultaneous observations, invaluable to study systematic differences between these measurements. The differences are partially attributed to the seasonal variability in atmospheric temperatures and ozone slant paths (OSP). The sensitivity to the temperature dependence of the ozone absorption cross section is calculated for each operational Brewer spectrophotometers at Arosa by using different high- and low-resolution reference spectra appropriately weighted with the instruments' slit functions, whereas the information on the primary standard instruments is used for all the Dobson instruments. The Brewer retrieval algorithm reveals a higher sensitivity to the reference spectra applied than the Dobson. When adopting the Bass and Paur (1985) or Malicet et al. (1995) ozone absorption spectra with their specific temperature dependence, and correcting for the OSP effect, the seasonal bias between Dobson and Brewer TOZ measurements is reduced to 0.6%. Conversely, these differences increase when using the spectral data of Burrows et al. (1999). This finding illustrates that the accuracy of ground-based spectrophotometric TOZ measurements is limited by the uncertainty in the ozone cross sections measured by different internationally leading laboratories.

Temperature and Slant Path Effects in Dobson and Brewer Total Ozone Measurements

NASA Astrophysics Data System (ADS)

Scarnato, B.; Staehelin, J.; Peter, T.; Groebner, J.; Stuebi, R.

2009-12-01

There is a worldwide tendency to replace Dobson spectrophotometers in ground-based total ozone (TOZ) measurements by more advanced Brewer spectrophotometers. Ensuring the homogeneity of these datasets is of utmost importance if changes in TOZ of few percent over long time periods are to be diagnosed accurately. Previous studies have identified a seasonal bias of few percent between mid-latitude Brewer and Dobson measurements. At Arosa (Switzerland), two Dobson and three Brewer instruments are co-located since 1998, providing a unique dataset of quasi-simultaneous observations, invaluable to study systematic differences between these measurements. The differences are partially attributed to the seasonal variability in atmospheric temperatures and ozone slant paths (OSP). The sensitivity to the temperature dependence of the ozone absorption cross-section are calculated for each operational Brewer spectrophotometers at Arosa by using different high and low resolution reference spectra appropriately weighted with the instruments' slit functions, whereas the information of the primary standard instruments are used for all the Dobson instruments. The Brewer retrieval algorithm reveals a higher sensitivity to the reference spectra applied than the Dobson. When adopting the Bass and Paur (1985) or Malicet et al. (1995) ozone absorption spectra with their specific temperature dependence, and correcting for the OSP effect, the seasonal bias between Dobson and Brewer TOZ measurements is reduced to 0.6%. Conversely these differences increase using the spectral data of Burrows (1999). This finding illustrates, that the accuracy of ground-based spectrophotometric TOZ measurements is limited by the uncertainty in the ozone cross-sections measured by different internationally leading laboratories.

Competence of medical students in communicating drug therapy: Value of role-play demonstrations.

PubMed

Tayem, Yasin I; Altabtabaei, Abdulaziz S; Mohamed, Mohamed W; Arrfedi, Mansour M; Aljawder, Hasan S; Aldebous, Fahad A; James, Henry; Al Khaja, Khalid A J; Sequeira, Reginald P

2016-01-01

This study used role-play demonstrations to train medical students to communicate drug therapy and evaluated the perceptions on this instructional approach. The second-year medical students who attended a prescription writing session (n = 133), participated in this study. Prescription communication was introduced by using role-play demonstrations. Participant's perceptions were explored by a self-administered questionnaire and focus group discussion. The academic achievement of attendees and nonattendees was compared with an objective structured performance evaluation (OSPE) station that tested students' competence in this skill. Most attendees responded to the questionnaire (81.2%). Almost all respondents expressed their desire to have similar demonstrations in other units. A large proportion of participants reported that role-play demonstrations helped them develop their communication skills, in general, confidence to communicate drug-related information in a prescription, and the ability to explain the aim of drug therapy to patients. Most trainees thought also that they developed skills to communicate instructions on drug use including drug dose, frequency of administration, duration of therapy, adverse drug reactions, and warnings. During the focus group interviews, students thought that role-play was useful but would be more beneficial if conducted frequently in small group as part of the curriculum implementation. The majority of students also reported improved competence in writing a complete prescription. Analysis of attendees and nonattendees grades in the OSPE showed that the former scored higher than the latter group (P = 0.016). Role-play demonstrations were well accepted by medical students and led to the development of their competence in communicating drug therapy to patients.

Generalized scalar particle quantization in 1+1 dimensions and D(2,1;α)

NASA Astrophysics Data System (ADS)

Corney, S. P.; Jarvis, P. D.; Tsohantjis, I.; McAnally, D. S.

2001-05-01

The exceptional superalgebra D(2,1;α) has been classified as a candidate conformal supersymmetry algebra in two dimensions. We propose an alternative interpretation of it as an extended BFV-BRST quantization superalgebra in 2D (D(2,1;1)≃osp(2,2|2)). A superfield realization is presented wherein the standard extended phase space coordinates can be identified. The physical states are studied via the cohomology of the BRST operator. Finally we reverse engineer a classical action corresponding to the algebraic model we have constructed, and identify the Lagrangian equations of motion.

The influence of experimental inflammation and axotomy on leucine enkephalin (leuENK) distribution in intramural nervous structures of the porcine descending colon.

PubMed

Gonkowski, Slawomir; Makowska, Krystyna; Calka, Jaroslaw

2018-05-24

The enteric nervous system (ENS), located in the intestinal wall and characterized by considerable independence from the central nervous system, consists of millions of cells. Enteric neurons control the majority of functions of the gastrointestinal tract using a wide range of substances, which are neuromediators and/or neuromodulators. One of them is leucine-enkephalin (leuENK), which belongs to the endogenous opioid family. It is known that opioids in the gastrointestinal tract have various functions, including visceral pain conduction, intestinal motility and secretion and immune processes, but many aspects of distribution and function of leuENK in the ENS, especially during pathological states, remain unknown. During this experiment, the distribution of leuENK - like immunoreactive (leuENK-LI) nervous structures using the immunofluorescence technique were studied in the porcine colon in physiological conditions, during chemically-induced inflammation and after axotomy. The study included the circular muscle layer, myenteric (MP), outer submucous (OSP) and inner submucous plexus (ISP) and the mucosal layer. In control animals, the number of leuENK-LI neurons amounted to 4.86 ± 0.17%, 2.86 ± 0.28% and 1.07 ± 0.08% in the MP, OSP and ISP, respectively. Generally, both pathological stimuli caused an increase in the number of detected leuENK-LI cells, but the intensity of the observed changes depended on the factor studied and part of the ENS. The percentage of leuENK-LI perikarya amounted to 11.48 ± 0.96%, 8.71 ± 0.13% and 9.40 ± 0.76% during colitis, and 6.90 ± 0.52% 8.46 ± 12% and 4.48 ± 0.44% after axotomy in MP, OSP and ISP, respectively. Both processes also resulted in an increase in the number of leuENK-LI nerves in the circular muscle layer, whereas changes were less visible in the mucosa during inflammation and axotomy did not change the number of leuENK-LI mucosal fibers. LeuENK in the ENS takes part in intestinal regulatory processes not only in physiological conditions, but also under pathological factors. The observed changes are probably connected with the participation of leuENK in sensory and motor innervation and the neuroprotective effects of this substance. Differences in the number of leuENK-LI neurons during inflammation and after axotomy may suggest that the exact functions of leuENK probably depend on the type of pathological factor acting on the intestine.

Conducting polymer nanocomposites loaded with nanotubes and fibers for electrical and thermal applications

NASA Astrophysics Data System (ADS)

Chiguma, Jasper

The design, fabrication and measurement of electrical and thermal properties of polymers loaded with nanotubes and fibers are the foci of the work presented in this dissertation. The resulting products of blending polymers with nanomaterials are called nanocomposites and are already finding applications in many areas of human endeavour. Among some of the most recent envisioned applications of nanocomposites is in electronic devices as thermal interface materials (TIMs). This potential application as TIMs, has been made more real by the realization that carbon nanotubes, could potentially transfer their high electrical, thermal and mechanical properties to polymers in the nanocomposites. In Chapter 1, the events leading to the discovery of carbon nanotubes are reviewed followed by an elaborate discussion of their structure and properties. The discussion of the structure and properties of carbon nanotubes help in understanding the envisaged applications. Chapter 2 focuses on the fabrication of insulating polymer nanocomposites, their electrical and mechanical properties. Poly (methyl methacrylate) (PMMA) and a polyimide formed by reacting pyromellitic dianhydride (PMDA) and 4, 4'-oxydianiline (ODA) (PMDA-ODA) nanocomposites with carbon nanotubes were prepared by in-situ polymerization. Poly (1-methyl-4-pentene) (TPX), Polycarbonate (PC), Poly (vinyl chloride) (PVC), Poly (acrylonitrile-butadiene-styrene) (ABS), the alloys ABS-PC, ABS-PVC, and ABS-PC-PVC nanocomposites were prepared from the respective polymers and carbon nanotubes and their mechanical and electrical properties measured. Chapter 3 covers the nanocomposites that were prepared by the in-situ polymerization of the conducting polymers Polyaniline (PANi), Polypyrrole (PPy) and Poly (3, 4-ethylenedioxythiophene) (PEDOT) by in-situ polymerization. These are evaluated for electrical conductivity. The use of surfactants in facilitating carbon nanotube dispersion is discussed and applied in the preparation of conducting polymer nanocomposites. In Chapter 4 epoxy nanocomposites are prepared. MWCNTs, Graphite Fibers and Boron Nitride are used as filler materials. There thermal conductivity is determined by using the Flash Technique as well as Differential Scanning Calorimetry (DSC). The thermal conductivity of graphite and BN loaded epoxy was found to be much higher than for the MWCNTs filled. Chapter 5 covers the synthesis and electrical conductivity of PANi nanotubes and nanorods without the use of templates. Also covered in this Chapter is the template free synthesis of Cu (II) hydroxide and Copper nanorods. In Chapter 6, Organic Solderability Preservatives (OSPs) are evaluated for integrity after thermal stress. The two types of OSPs that are evaluated in this chapter are a benzimidazole derivative known as WPF207 and an imidazole derivative called F2LX. The OSP WPF was found to be more robust. In Chapter 7, two encapsulants are evaluated after thermal stress. The encapsulants are Sumitomo type 6730B and type 6730B-LX. No significant differences were found after analysis.

Role of NleH, a type III secreted effector from attaching and effacing pathogens, in colonization of the bovine, ovine, and murine gut.

PubMed

Hemrajani, Cordula; Marches, Olivier; Wiles, Siouxsie; Girard, Francis; Dennis, Alison; Dziva, Francis; Best, Angus; Phillips, Alan D; Berger, Cedric N; Mousnier, Aurelie; Crepin, Valerie F; Kruidenier, Laurens; Woodward, Martin J; Stevens, Mark P; La Ragione, Roberto M; MacDonald, Thomas T; Frankel, Gad

2008-11-01

The human pathogen enterohemorrhagic Escherichia coli (EHEC) O157:H7 colonizes human and animal gut via formation of attaching and effacing lesions. EHEC strains use a type III secretion system to translocate a battery of effector proteins into the mammalian host cell, which subvert diverse signal transduction pathways implicated in actin dynamics, phagocytosis, and innate immunity. The genomes of sequenced EHEC O157:H7 strains contain two copies of the effector protein gene nleH, which share 49% sequence similarity with the gene for the Shigella effector OspG, recently implicated in inhibition of migration of the transcriptional regulator NF-kappaB to the nucleus. In this study we investigated the role of NleH during EHEC O157:H7 infection of calves and lambs. We found that while EHEC DeltanleH colonized the bovine gut more efficiently than the wild-type strain, in lambs the wild-type strain exhibited a competitive advantage over the mutant during mixed infection. Using the mouse pathogen Citrobacter rodentium, which shares many virulence factors with EHEC O157:H7, including NleH, we observed that the wild-type strain exhibited a competitive advantage over the mutant during mixed infection. We found no measurable differences in T-cell infiltration or hyperplasia in colons of mice inoculated with the wild-type or the nleH mutant strain. Using NF-kappaB reporter mice carrying a transgene containing a luciferase reporter driven by three NF-kappaB response elements, we found that NleH causes an increase in NF-kappaB activity in the colonic mucosa. Consistent with this, we found that the nleH mutant triggered a significantly lower tumor necrosis factor alpha response than the wild-type strain.

Multimodality vaccination against clade C SHIV: partial protection against mucosal challenges with a heterologous tier 2 virus.

PubMed

Lakhashe, Samir K; Byrareddy, Siddappa N; Zhou, Mingkui; Bachler, Barbara C; Hemashettar, Girish; Hu, Shiu-Lok; Villinger, Francois; Else, James G; Stock, Shannon; Lee, Sandra J; Vargas-Inchaustegui, Diego A; Cofano, Egidio Brocca; Robert-Guroff, Marjorie; Johnson, Welkin E; Polonis, Victoria R; Forthal, Donald N; Loret, Erwann P; Rasmussen, Robert A; Ruprecht, Ruth M

2014-11-12

We sought to test whether vaccine-induced immune responses could protect rhesus macaques (RMs) against upfront heterologous challenges with an R5 simian-human immunodeficiency virus, SHIV-2873Nip. This SHIV strain exhibits many properties of transmitted HIV-1, such as tier 2 phenotype (relatively difficult to neutralize), exclusive CCR5 tropism, and gradual disease progression in infected RMs. Since no human AIDS vaccine recipient is likely to encounter an HIV-1 strain that exactly matches the immunogens, we immunized the RMs with recombinant Env proteins heterologous to the challenge virus. For induction of immune responses against Gag, Tat, and Nef, we explored a strategy of immunization with overlapping synthetic peptides (OSP). The immune responses against Gag and Tat were finally boosted with recombinant proteins. The vaccinees and a group of ten control animals were given five low-dose intrarectal (i.r.) challenges with SHIV-2873Nip. All controls and seven out of eight vaccinees became systemically infected; there was no significant difference in viremia levels of vaccinees vs. controls. Prevention of viremia was observed in one vaccinee which showed strong boosting of virus-specific cellular immunity during virus exposures. The protected animal showed no challenge virus-specific neutralizing antibodies in the TZM-bl or A3R5 cell-based assays and had low-level ADCC activity after the virus exposures. Microarray data strongly supported a role for cellular immunity in the protected animal. Our study represents a case of protection against heterologous tier 2 SHIV-C by vaccine-induced, virus-specific cellular immune responses. Copyright © 2014 Elsevier Ltd. All rights reserved.

Oxygen sensitive polymeric nanocapsules for optical dissolved oxygen sensors

NASA Astrophysics Data System (ADS)

Sun, Zhijuan; Cai, Chenxin; Guo, Fei; Ye, Changhuai; Luo, Yingwu; Ye, Shuming; Luo, Jianchao; Zhu, Fan; Jiang, Chunyue

2018-04-01

Immobilization of the oxygen-sensitive probes (OSPs) in the host matrix greatly impacts the performance and long-term usage of the optical dissolved oxygen (DO) sensors. In this work, fluorescent dyes, as the OSPs, were encapsulated with a crosslinked fluorinated polymer shell by interfacial confined reversible addition fragmentation chain transfer miniemulsion polymerization to fabricate oxygen sensitive polymeric nanocapsules (NCs). The location of fluorescent dyes and the fluorescent properties of the NCs were fully characterized by fourier transform infrared spectrometer, x-ray photoelectron spectrometer and fluorescent spectrum. Dye-encapsulated capacity can be precisely tuned from 0 to 1.3 wt% without self-quenching of the fluorescent dye. The crosslinked fluorinated polymer shell is not only extremely high gas permeability, but also prevents the fluorescent dyes from leakage in aqueous as well as in various organic solvents, such as ethanol, acetone and tetrahydrofuran (THF). An optical DO sensor based on the oxygen sensitive NCs was fabricated, showing high sensitivity, short response time, full reversibility, and long-term operational stability of online monitoring DO. The sensitivity of the optical DO sensor is 7.02 (the ratio of the response value in fully deoxygenated and saturated oxygenated water) in the range 0.96-14.16 mg l-1 and the response time is about 14.3 s. The sensor’s work curve was fit well using the modified Stern-Volmer equation by two-site model, and its response values are hardly affected by pH ranging from 2 to 12 and keep constant during continuous measurement for 3 months. It is believed that the oxygen sensitive polymeric NCs-based optical DO sensor could be particularly useful in long-term online DO monitoring in both aqueous and organic solvent systems.

Ready for the OR? – Clinical anatomy and basic surgical skills for students in their preclinical education

PubMed Central

Böckers, Anja; Lippold, Dominique; Fassnacht, Ulrich; Schelzig, Hubert; Böckers, Tobias M.

2011-01-01

Medical students’ first experience in the operating theatre often takes place during their electives and is therefore separated from the university’s medical curriculum. In the winter term 2009/10, the Institute of Anatomy and Cell Biology at the University of Ulm implemented an elective called “Ready for the OR” for 2nd year medical students participating in the dissection course. We attempted to improve learning motivation and examination results by transferring anatomical knowledge into a surgical setting and teaching basic surgical skills in preparation of the students’ first participation in the OR. Out of 69 online applicants, 50 students were randomly assigned to the Intervention Group (FOP) or the Control Group. In 5 teaching session students learned skills like scrubbing, stitching or the identification of frequently used surgical instruments. Furthermore, students visited five surgical interventions which were demonstrated by surgical colleagues on donated bodies that have been embalmed using the Thiel technique. The teaching sessions took place in the institute’s newly built “Theatrum Anatomicum” for an ideal simulation of a surgical setting. The learning outcomes were verified by OSPE. In a pilot study, an intervention group and a control group were compared concerning their examination results in the dissection course and their learning motivation through standardized SELLMO-test for students. Participants gained OSPE results between 60.5 and 92% of the maximum score. “Ready for the OR” was successfully implemented and judged an excellent add-on to anatomy teaching by the participants. However, we could not prove a significant difference in learning motivation or examination results. Future studies should focus on the learning orientation, the course’s long-term learning effects and the participants’ behavior in a real surgery setting. PMID:21866247

Genetic variation in transmission success of the Lyme borreliosis pathogen Borrelia afzelii.

PubMed

Tonetti, Nicolas; Voordouw, Maarten J; Durand, Jonas; Monnier, Séverine; Gern, Lise

2015-04-01

The vector-to-host and host-to-vector transmission steps are the two critical events that define the life cycle of any vector-borne pathogen. We expect negative genetic correlations between these two transmission phenotypes, if parasite genotypes specialized at invading the vector are less effective at infecting the vertebrate host and vice versa. We used the tick-borne bacterium Borrelia afzelii, a causative agent of Lyme borreliosis in Europe, to test whether genetic trade-offs exist between tick-to-host, systemic (host-to-tick), and a third mode of co-feeding (tick-to-tick) transmission. We worked with six strains of B. afzelii that were differentiated according to their ospC gene. We compared the three components of transmission among the B. afzelii strains using laboratory rodents as the vertebrate host and a laboratory colony of Ixodes ricinus as the tick vector. We used next generation matrix models to combine these transmission components into a single estimate of the reproductive number (R0) for each B. afzelii strain. We also tested whether these strain-specific estimates of R0 were correlated with the strain-specific frequencies in the field. We found significant genetic variation in the three transmission components among the B. afzelii strains. This is the first study to document genetic variation in co-feeding transmission for any tick-borne pathogen. We found no evidence of trade-offs as the three pairwise correlations of the transmission rates were all positive. The R0 values from our laboratory study explained 45% of the variation in the frequencies of the B. afzelii ospC strains in the field. Our study suggests that laboratory estimates of pathogen fitness can predict the distribution of pathogen strains in nature. Copyright © 2015 Elsevier GmbH. All rights reserved.

Seasonal and Spatial Variation of Particulate Cadmium and Phosphorous along Line P in the Northeast Pacific

NASA Astrophysics Data System (ADS)

Bourne, H.; Bishop, J. K. B.; Wood, T.

2016-02-01

Line P is a transect along 50°N in the subarctic North Pacific that stretches from Sanich Inlet out to Ocean Station PAPA (OSP). Between February 1996 and March the following year, size fraction suspended particulate matter was collected (<1 μm, 1-51 μm and >51 μm) using the Multiple Unit Large Volume Filtration System (MULVFS) technique. Over the course of 4 cruises, 17 MULFS casts were taken. Using this large dataset, we examine the seasonal and spatial variability of cadmium content of particles. Throughout this one year time span, the particulate Cd uptake relative to P varied by a factor of two at individual sites during different seasons and by a factor of three at different sites during the same season. Typically, the Cd:P content of phytoplankton increases going from Sanich Inlet out toward station PAPA. While Sanich Inlet is an iron and zinc replete coastal environment, OSP is located in an HNLC region with low dissolved Fe and Zn concentrations. We compared the Cd:P profiles along the Line P transect to that of other divalent metals, specifically Zn, Mn, Co, Ba and Fe. While Fe:P, Mn:P and Co:P are highest in the coastal environment, Cd:P is higher in the open ocean environment. Cd:P peaks in the open ocean station below the mixed layer. We also compare the relationship between oxygen and Cd content in particles. The reason for Cd uptake by marine organisms is still not fully understood; by examining particles in a well-studied region with seasonal, nutrient, environmental and other divalent metal concentrations in particles, we can better understand the causes for variability.

Atmospheric iron supply and enhanced vertical carbon flux in the NE subarctic Pacific: Is there a connection?

NASA Astrophysics Data System (ADS)

Boyd, P. W.; Wong, C. S.; Merrill, J.; Whitney, F.; Snow, J.; Harrison, P. J.; Gower, J.

1998-09-01

Recent studies have confirmed the relationship between iron supply and phytoplankton growth rates in all three high-nitrate low-chlorophyll (HNLC) oceanic provinces. However, there is little evidence, so far, of the role of iron in altering the efficiency of the biological pump via increased downward export of particulate organic carbon (POC). The NE subarctic Pacific is unique among HNLC regions in that long time series pelagic observations and deep-moored sediment trap records exist which may provide the best opportunity thus far to test aspects of the iron hypothesis. Episodic elevated levels of chlorophyll a (> 2.0 μg L-1) were observed 6 times between 1964 and 1976 at the former site of Ocean Station Papa (OSP). In addition, between 1984 and 1990 on at least three occasions, concurrent pulses of POC and biogenic silica were recorded in deep-moored traps at OSP. Possible explanations for these events, such as lateral advection of more productive waters, iron-mediated blooms, or grazing by salp swarms are discussed and tested using an existing downward POC flux model. Owing to the episodic nature of such events, no available data are sufficiently comprehensive to unequivocally rule out any of these explanations. Nevertheless, from the data available, the occurrence of pelagic or deep water pulses, approximately once every 3 years, are most consistent with iron-mediated diatom blooms, and of the sinking of POC and biogenic silica (from such a bloom) to depth, respectively. A comparison of the timing of these iron-mediated pulses with that of the transport probabilities of atmospheric dust supply from Asia and Alaska provides an opportunity to assess the likelihood of a coupling between the atmosphere and the ocean.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoilova, N. I.

Generalized quantum statistics, such as paraboson and parafermion statistics, are characterized by triple relations which are related to Lie (super)algebras of type B. The correspondence of the Fock spaces of parabosons, parafermions as well as the Fock space of a system of parafermions and parabosons to irreducible representations of (super)algebras of type B will be pointed out. Example of generalized quantum statistics connected to the basic classical Lie superalgebra B(1|1) ≡ osp(3|2) with interesting physical properties, such as noncommutative coordinates, will be given. Therefore the article focuses on the question, addressed already in 1950 by Wigner: do the equation ofmore » motion determine the quantum mechanical commutation relation?.« less

76 FR 41497 - Privacy Act System of Records

Federal Register 2010, 2011, 2012, 2013, 2014

2011-07-14

...Pursuant to subsection (e)(4) of the Privacy Act of 1974, as amended (Privacy Act), 5 U.S.C. 552a, the FCC proposes to alter one system of records, FCC/OSP-1, ``Broadband Dead Zone Report and Consumer Broadband Test.'' The altered system of records incorporates more details about the voluntary fixed and mobile consumer broadband test. The FCC will also alter the categories of individuals; categories of records; the purposes for which the information is maintained; the retrievability procedures; Routine Use (5); and delete Routine Use (2); and make other edits and revisions as necessary to update the information and to comply with the requirements of the Privacy Act.

Real-time single-molecule observations of proteins at the solid-liquid interface

NASA Astrophysics Data System (ADS)

Langdon, Blake Brianna

Non-specific protein adsorption to solid surfaces is pervasive and observed across a broad spectrum of applications including biomaterials, separations, pharmaceuticals, and biosensing. Despite great interest in and considerable literature dedicated to the phenomena, a mechanistic understanding of this complex phenomena is lacking and remains controversial, partially due to the limits of ensemble-averaging techniques used to study it. Single-molecule tracking (SMT) methods allow us to study distinct protein dynamics (e.g. adsorption, desorption, diffusion, and intermolecular associations) on a molecule-by-molecule basis revealing the protein population and spatial heterogeneity inherent in protein interfacial behavior. By employing single-molecule total internal reflection fluorescence microscopy (SM-TIRFM), we have developed SMT methods to directly observe protein interfacial dynamics at the solid-liquid interface to build a better mechanistic understanding of protein adsorption. First, we examined the effects of surface chemistry (e.g. hydrophobicity, hydrogen-bonding capacity), temperature, and electrostatics on isolated protein desorption and interfacial diffusion for fibrinogen (Fg) and bovine serum albumin (BSA). Next, we directly and indirectly probed the effects of protein-protein interactions on interfacial desorption, diffusion, aggregation, and surface spatial heterogeneity on model and polymeric thin films. These studies provided many useful insights into interfacial protein dynamics including the following observations. First, protein adsorption was reversible, with the majority of proteins desorbing from all surface chemistries within seconds. Isolated protein-surface interactions were relatively weak on both hydrophobic and hydrophilic surfaces (apparent desorption activation energies of only a few kBT). However, proteins could dynamically and reversibly associate at the interface, and these interfacial associations led to proteins remaining on the surface for longer time intervals. Surface chemistry and surface spatial heterogeneity (i.e. surface sites with different binding strengths) were shown to influence adsorption, desorption, and interfacial protein-protein associations. For example, faster protein diffusion on hydrophobic surfaces increased protein-protein associations and, at higher protein surface coverage, led to proteins remaining on hydrophobic surfaces longer than on hydrophilic surfaces. Ultimately these studies suggested that surface properties (chemistry, heterogeneity) influence not only protein-surface interactions but also interfacial mobility and protein-protein associations, implying that surfaces that better control protein adsorption can be designed by accounting for these processes.

Protein-Nanoparticle Interactions: Improving Immobilized Lytic Enzyme Activity and Surface Energy Effects

NASA Astrophysics Data System (ADS)

Downs, Emily Elizabeth

Protein-nanostructure conjugates, particularly particles, are a subject of significant interest due to changes in their fundamental behavior compared to bulk surfaces. As the size scale of nano-structured materials and proteins are on the same order of magnitude, nanomaterial properties can heavily influence how proteins adsorb and conform to the surface. Previous work has demonstrated the ability of nanoscale surfaces to modulate protein activity, conformation, and retention by modifying the particle surface curvature, morphology, and surface charge. This work has improved our understanding of the protein material interactions, but a complete understanding is still lacking. The goal of this thesis is to investigate two missing areas of understanding using two distinct systems. The first system utilizes a particle with controlled surface energy to observe the impact of surface energy on protein-particle interactions, while the second system uses a modified Listeria-specific protein to determine how protein structure and flexibility affects protein adsorption and activity on particles. Spherical, amorphous, and uniformly doped Zn-silica particles with tailored surface energies were synthesized to understand the impact of surface energy on protein adsorption behavior. Particle surface energy increased with a decrease in particle size and greater dopant concentrations. Protein adsorption and structural loss increased with both particle size and particle surface energy. Higher surface energies promoted protein-particle association and increased protein unfolding. Particle curvature and protein steric hindrance effects limited adsorption and structural loss on smaller particles. Protein surface charge heterogeneity was also found to be linked to both protein adsorption and unfolding behavior on larger particles. Greater surface charge heterogeneity led to higher adsorption concentrations and multilayer formation. These multilayers transitioned from protein-particle interactions to protein-protein interactions and were thicker with greater surface energy, which resulted in the recovery of secondary structure in the outermost layer. To help understand the impact of protein structure on nano-bio conjugate interactions, a listeria specific protein was used. This system was chosen as it has applications in the food industry in preventing bacterial contamination. The insertion of an amino acid linker between the enzymatic and binding domain of the protein improved the flexibility between domains, leading to increased adsorption, and improved activity in both cell-wall and plating assays. Additionally, linker modified protein incorporated into the silica-polymer nanocomposite showed significant activity in a real-world example of contaminated lettuce. This thesis study has isolated the impact of surface energy and protein flexibility on protein adsorption and structure. Particle surface energy affects adsorbed protein concentration and conformation. Coupled with protein surface charge, surface energy was also found to dictate multilayer thickness. The conformational flexibility of the protein was shown to help in controlling not only protein adsorption concentration but also in retaining protein activity after immobilization. Also, a controllable synthesis method for particles with adjustable surface energy, an ideal platform for studying protein-particle interactions, has been established.

Upper ocean bubble measurements from the NE Pacific and estimates of their role in air-sea gas transfer of the weakly soluble gases nitrogen and oxygen

NASA Astrophysics Data System (ADS)

Vagle, Svein; McNeil, Craig; Steiner, Nadja

2010-12-01

Simultaneous observations of upper-ocean bubble clouds, and dissolved gaseous nitrogen (N2) and oxygen (O2) from three winter storms are presented and analyzed. The data were collected on the Canadian Surface Ocean Lower Atmosphere Study (C-SOLAS) mooring located near Ocean Station Papa (OSP) at 50°N, 145°W in the NE Pacific during winter of 2003/2004. The bubble field was measured using an upward looking 200 kHz echosounder. Direct estimates of bubble mediated gas fluxes were made using assumed bubble size spectra and the upward looking echosounder data. A one-dimensional biogeochemical model was used to help compare data and various existing models of bubble mediated air-sea gas exchange. The direct bubble flux calculations show an approximate quadratic/cubic dependence on mean bubble penetration depth. After scaling from N2/O2 to carbon dioxide, near surface, nonsupersaturating, air-sea transfer rates, KT, for U10 > 12 m s-1 fall between quadratic and cubic relationships. Estimates of the subsurface bubble induced air injection flux, VT, show an approximate quadratic/cubic dependence on mean bubble penetration depth. Both KT and VT are much higher than those measured during Hurricane Frances over the wind speed range 12 < U10 < 23 m s-1. This result implies that over the open ocean and this wind speed range, older and more developed seas which occur during winter storms are more effective in exchanging gases between the atmosphere and ocean than younger less developed seas which occur during the rapid passage of a hurricane.

Water at protein surfaces studied with femtosecond nonlinear spectroscopy

NASA Astrophysics Data System (ADS)

Bakker, Huib J.

We report on an investigation of the structure and dynamics of water molecules near protein surfaces with femtosecond nonlinear spectroscopic techniques. We measured the reorientation dynamics of water molecules near the surface of several globular protein surfaces, using polarization-resolved femtosecond infrared spectroscopy. We found that water molecules near the protein surface have a much slower reorientation than water molecules in bulk liquid water. The number of slow water molecules scales scales with the size of the hydrophobic surface of the protein. When we denature the proteins by adding an increasing amount of urea to the protein solution, we observe that the water-exposed surface increases by 50% before the secondary structure of the proteins changes. This finding indicates that protein unfolding starts with the protein structure becoming less tight, thereby allowing water to enter. With surface vibrational sum frequency generation (VSFG) spectroscopy, we studied the structure of water at the surface of antifreeze protein III. The measured VSFG spectra showed the presence of ice-like water layers at the ice-binding site of the protein in aqueous solution, at temperatures well above the freezing point. This ordered ice-like hydration layers at the protein surface likely plays an important role in the specific recognition and binding of anti-freeze protein III to nascent ice crystallites, and thus in its anti-freeze mechanism. This research is supported by the ''Nederlandse organisatie voor Wetenschappelijk Onderzoek (NWO).

Modulation of protein stability and aggregation properties by surface charge engineering.

PubMed

Raghunathan, Govindan; Sokalingam, Sriram; Soundrarajan, Nagasundarapandian; Madan, Bharat; Munussami, Ganapathiraman; Lee, Sun-Gu

2013-09-01

An attempt to alter protein surface charges through traditional protein engineering approaches often affects the native protein structure significantly and induces misfolding. This limitation is a major hindrance in modulating protein properties through surface charge variations. In this study, as a strategy to overcome such a limitation, we attempted to co-introduce stabilizing mutations that can neutralize the destabilizing effect of protein surface charge variation. Two sets of rational mutations were designed; one to increase the number of surface charged amino acids and the other to decrease the number of surface charged amino acids by mutating surface polar uncharged amino acids and charged amino acids, respectively. These two sets of mutations were introduced into Green Fluorescent Protein (GFP) together with or without stabilizing mutations. The co-introduction of stabilizing mutations along with mutations for surface charge modification allowed us to obtain functionally active protein variants (s-GFP(+15-17) and s-GFP(+5-6)). When the protein properties such as fluorescent activity, folding rate and kinetic stability were assessed, we found the possibility that the protein stability can be modulated independently of activity and folding by engineering protein surface charges. The aggregation properties of GFP could also be altered through the surface charge engineering.

Effects of surface wettability and contact time on protein adhesion to biomaterial surfaces

PubMed Central

Xu, Li-Chong; Siedlecki, Christopher A.

2013-01-01

Atomic force microscopy (AFM) was used to directly measure the adhesion forces between three test proteins and low density polyethylene (LDPE) surfaces treated by glow discharge plasma to yield various levels of water wettability. The adhesion of proteins to the LDPE substrates showed a step dependence on the wettability of surfaces as measured by the water contact angle (θ). For LDPE surfaces with θ > ∼60–65°, stronger adhesion forces were observed for bovine serum albumin, fibrinogen and human FXII than for the surfaces with θ < 60°. Smaller adhesion forces were observed for FXII than for the other two proteins on all surfaces although trends were identical. Increasing the contact time from 0 to 50 s for each protein–surface combination increased the adhesion force regardless of surface wettability. Time varying adhesion data was fit to an exponential model and free energies of protein unfolding were calculated. This data, viewed in light of previously published studies, suggests a 2-step model of protein denaturation, an early stage on the order of seconds to minutes where the outer surface of the protein interacts with the substrate and a second stage involving movement of hydrophobic amino acids from the protein core to the protein/surface interface. Impact statement The work described in this manuscript shows a stark transition between protein adherent and protein non-adherent materials in the range of water contact angles 60–65°, consistent with known changes in protein adsorption and activity. Time-dependent changes in adhesion force were used to calculate unfolding energies relating to protein–surface interactions. This analysis provides justification for a 2-step model of protein denaturation on surfaces. PMID:17466368

A mobile precursor determines protein resistance on nanostructured surfaces.

PubMed

Wang, Kang; Chen, Ye; Gong, Xiangjun; Xia, Jianlong; Zhao, Junpeng; Shen, Lei

2018-05-09

Biomaterials are often engineered with nanostructured surfaces to control interactions with proteins and thus regulate their biofunctions. However, the mechanism of how nanostructured surfaces resist or attract proteins together with the underlying design rules remains poorly understood at a molecular level, greatly limiting attempts to develop high-performance biomaterials and devices through the rational design of nanostructures. Here, we study the dynamics of nonspecific protein adsorption on block copolymer nanostructures of varying adhesive domain areas in a resistant matrix. Using surface plasmon resonance and single molecule tracking techniques, we show that weakly adsorbed proteins with two-dimensional diffusivity are critical precursors to protein resistance on nanostructured surfaces. The adhesive domain areas must be more than tens or hundreds of times those of the protein footprints to slow down the 2D-mobility of the precursor proteins for their irreversible adsorption. This precursor model can be used to quantitatively analyze the kinetics of nonspecific protein adsorption on nanostructured surfaces. Our method is applicable to precisely manipulate protein adsorption and resistance on various nanostructured surfaces, e.g., amphiphilic, low-surface-energy, and charged nanostructures, for the design of protein-compatible materials.

Surface Mediated Protein Disaggregation

NASA Astrophysics Data System (ADS)

Radhakrishna, Mithun; Kumar, Sanat K.

2014-03-01

Preventing protein aggregation is of both biological and industrial importance. Biologically these aggregates are known to cause amyloid type diseases like Alzheimer's and Parkinson's disease. Protein aggregation leads to reduced activity of the enzymes in industrial applications. Inter-protein interactions between the hydrophobic residues of the protein are known to be the major driving force for protein aggregation. In the current paper we show how surface chemistry and curvature can be tuned to mitigate these inter-protein interactions. Our results calculated in the framework of the Hydrophobic-Polar (HP) lattice model show that, inter-protein interactions can be drastically reduced by increasing the surface hydrophobicity to a critical value corresponding to the adsorption transition of the protein. At this value of surface hydrophobicity, proteins lose inter-protein contacts to gain surface contacts and thus the surface helps in reducing the inter-protein interactions. Further, we show that the adsorption of the proteins inside hydrophobic pores of optimal sizes are most efficient both in reducing inter-protein contacts and simultaneously retaining most of the native-contacts due to strong protein-surface interactions coupled with stabilization due to the confinement. Department of Energy (Grant No DE-FG02-11ER46811).

Online formative assessments: exploring their educational value

PubMed Central

NAGANDLA, KAVITHA; SULAIHA, SHARIFAH; NALLIAH, SIVALINGAM

2018-01-01

Introduction: Online formative assessments (OFA’s) have been increasingly recognised in medical education as resources that promote self-directed learning. Formative assessments are used to support the self-directed learning of students. Online formative assessments have been identified to be less time consuming with automated feedback. This pilot study aimed to determine whether participation and performance in online formative assessments (OFA’s) had measurable effects on learning and evaluate the students’ experience of using the OFA’s in the department of Obstetrics and Gynaecology. Methods: This is a cross-sectional study conducted among fourth year medical students (n=92) during their seven week postings in Obstetrics and Gynaecology. Five sets of online formative assessments in the format of one best answers (OBA), Objective structured practical examination (OSPE) and Short answer question (SAQ) with feedback were delivered over five weeks through the online portal. The mean scores of the end of posting summative exam (EOP) of those who participated in the assessments (OFA users) and of those who did not (non-OFA users) were compared, using Students t test. The frequency of tool usage was analysed and satisfaction surveys were utilized at the end of the course by survey questionnaire using the five point Likert scale. Results: The mean scores of the students in end of posting summative examination marks for students who had participated in the online formative assessment (OFA users) and for those who had not (non OFA users) showed no significant difference in all the three components OBA, SAQ and OSPE (p=0.902, 0.633, 0.248). Majority of the students perceived that OFAs fulfilled the stated aims and objectives and so they would persuade their peers to participate in the OFAs. Conclusions: Online formative assessments are perceived as tools that promote self-directed learning, improved knowledge and tailor learning for individual learning needs and style. PMID:29607332

Enzootic origins for clinical manifestations of Lyme borreliosis.

PubMed

Jahfari, Setareh; Krawczyk, Aleksandra; Coipan, E Claudia; Fonville, Manoj; Hovius, Joppe W; Sprong, Hein; Takumi, Katsuhisa

2017-04-01

Both early localized and late disseminated forms of Lyme borreliosis are caused by Borrelia burgdorferi senso lato. Differentiating between the spirochetes that only cause localized skin infection from those that cause disseminated infection, and tracing the group of medically-important spirochetes to a specific vertebrate host species, are two critical issues in disease risk assessment and management. Borrelia burgdorferi senso lato isolates from Lyme borreliosis cases with distinct clinical manifestations (erythema migrans, neuroborreliosis, acrodermatitis chronica atrophicans, and Lyme arthritis) and isolates from Ixodes ricinus ticks feeding on rodents, birds and hedgehogs were typed to the genospecies level by sequencing part of the intergenic spacer region. In-depth molecular typing was performed by sequencing eight additional loci with different characteristics (plasmid-bound, regulatory, and housekeeping genes). The most abundant genospecies and genotypes in the clinical isolates were identified by using odds ratio as a measure of dominance. Borrelia afzelii was the most common genospecies in acrodermatitis patients and engorged ticks from rodents. Borrelia burgdorferi senso stricto was widespread in erythema migrans patients. Borrelia bavariensis was widespread in neuroborreliosis patients and in ticks from hedgehogs, but rare in erythema migrans patients. Borrelia garinii was the dominant genospecies in ticks feeding on birds. Spirochetes in ticks feeding on hedgehogs were overrepresented in genotypes of the plasmid gene ospC from spirochetes in erythema migrans patients. Spirochetes in ticks feeding on hedgehogs were overrepresented in genotypes of ospA from spirochetes in acrodermatitis patients. Spirochetes from ticks feeding on birds were overrepresented in genotypes of the plasmid and regulatory genes dbpA, rpoN and rpoS from spirochetes in neuroborreliosis patients. Overall, the analyses of our datasets support the existence of at least three transmission pathways from an enzootic cycle to a clinical manifestation of Lyme borreliosis. Based on the observations with these nine loci, it seems to be justified to consider the population structure of B. burgdorferi senso lato as being predominantly clonal. Copyright © 2017 Elsevier B.V. All rights reserved.

Remediation of hexavalent chromium contamination in chromite ore processing residue by sodium dithionite and sodium phosphate addition and its mechanism.

PubMed

Li, Yunyi; Cundy, Andrew B; Feng, Jingxuan; Fu, Hang; Wang, Xiaojing; Liu, Yangsheng

2017-05-01

Large amounts of chromite ore processing residue (COPR) wastes have been deposited in many countries worldwide, generating significant contamination issues from the highly mobile and toxic hexavalent chromium species (Cr(VI)). In this study, sodium dithionite (Na 2 S 2 O 4 ) was used to reduce Cr(VI) to Cr(III) in COPR containing high available Fe, and then sodium phosphate (Na 3 PO 4 ) was utilized to further immobilize Cr(III), via a two-step procedure (TSP). Remediation and immobilization processes and mechanisms were systematically investigated using batch experiments, sequential extraction studies, X-ray diffraction (XRD) and X-ray Photoelectron Spectroscopy (XPS). Results showed that Na 2 S 2 O 4 effectively reduced Cr(VI) to Cr(III), catalyzed by Fe(III). The subsequent addition of Na 3 PO 4 further immobilized Cr(III) by the formation of crystalline CrPO 4 ·6H 2 O. However, addition of Na 3 PO 4 simultaneously with Na 2 S 2 O 4 (via a one-step procedure, OSP) impeded Cr(VI) reduction due to the competitive reaction of Na 3 PO 4 and Na 2 S 2 O 4 with Fe(III). Thus, the remediation efficiency of the TSP was much higher than the corresponding OSP. Using an optimal dosage in the two-step procedure (Na 2 S 2 O 4 at a dosage of 12× the stoichiometric requirement for 15 days, and then Na 3 PO 4 in a molar ratio (i.e. Na 3 PO 4 : initial Cr(VI)) of 4:1 for another 15 days), the total dissolved Cr in the leachate determined via Toxicity Characteristic Leaching Procedure (TCLP Cr) testing of our samples was reduced to 3.8 mg/L (from an initial TCLP Cr of 112.2 mg/L, i.e. at >96% efficiency). Copyright © 2017 Elsevier Ltd. All rights reserved.

Rapid comparison of properties on protein surface

PubMed Central

Sael, Lee; La, David; Li, Bin; Rustamov, Raif; Kihara, Daisuke

2008-01-01

The mapping of physicochemical characteristics onto the surface of a protein provides crucial insights into its function and evolution. This information can be further used in the characterization and identification of similarities within protein surface regions. We propose a novel method which quantitatively compares global and local properties on the protein surface. We have tested the method on comparison of electrostatic potentials and hydrophobicity. The method is based on 3D Zernike descriptors, which provides a compact representation of a given property defined on a protein surface. Compactness and rotational invariance of this descriptor enable fast comparison suitable for database searches. The usefulness of this method is exemplified by studying several protein families including globins, thermophilic and mesophilic proteins, and active sites of TIM β/α barrel proteins. In all the cases studied, the descriptor is able to cluster proteins into functionally relevant groups. The proposed approach can also be easily extended to other surface properties. This protein surface-based approach will add a new way of viewing and comparing proteins to conventional methods, which compare proteins in terms of their primary sequence or tertiary structure. PMID:18618695

Rapid comparison of properties on protein surface.

PubMed

Sael, Lee; La, David; Li, Bin; Rustamov, Raif; Kihara, Daisuke

2008-10-01

The mapping of physicochemical characteristics onto the surface of a protein provides crucial insights into its function and evolution. This information can be further used in the characterization and identification of similarities within protein surface regions. We propose a novel method which quantitatively compares global and local properties on the protein surface. We have tested the method on comparison of electrostatic potentials and hydrophobicity. The method is based on 3D Zernike descriptors, which provides a compact representation of a given property defined on a protein surface. Compactness and rotational invariance of this descriptor enable fast comparison suitable for database searches. The usefulness of this method is exemplified by studying several protein families including globins, thermophilic and mesophilic proteins, and active sites of TIM beta/alpha barrel proteins. In all the cases studied, the descriptor is able to cluster proteins into functionally relevant groups. The proposed approach can also be easily extended to other surface properties. This protein surface-based approach will add a new way of viewing and comparing proteins to conventional methods, which compare proteins in terms of their primary sequence or tertiary structure.

Quantitative proteomic view on secreted, cell surface-associated, and cytoplasmic proteins of the methicillin-resistant human pathogen Staphylococcus aureus under iron-limited conditions.

PubMed

Hempel, Kristina; Herbst, Florian-Alexander; Moche, Martin; Hecker, Michael; Becher, Dörte

2011-04-01

Staphylococcus aureus is capable of colonizing and infecting humans by its arsenal of surface-exposed and secreted proteins. Iron-limited conditions in mammalian body fluids serve as a major environmental signal to bacteria to express virulence determinants. Here we present a comprehensive, gel-free, and GeLC-MS/MS-based quantitative proteome profiling of S. aureus under this infection-relevant situation. (14)N(15)N metabolic labeling and three complementing approaches were combined for relative quantitative analyses of surface-associated proteins. The surface-exposed and secreted proteome profiling approaches comprise trypsin shaving, biotinylation, and precipitation of the supernatant. By analysis of the outer subproteomic and cytoplasmic protein fraction, 1210 proteins could be identified including 221 surface-associated proteins. Thus, access was enabled to 70% of the predicted cell wall-associated proteins, 80% of the predicted sortase substrates, two/thirds of lipoproteins and more than 50% of secreted and cytoplasmic proteins. For iron-deficiency, 158 surface-associated proteins were quantified. Twenty-nine proteins were found in altered amounts showing particularly surface-exposed proteins strongly induced, such as the iron-regulated surface determinant proteins IsdA, IsdB, IsdC and IsdD as well as lipid-anchored iron compound-binding proteins. The work presents a crucial subject for understanding S. aureus pathophysiology by the use of methods that allow quantitative surface proteome profiling.

Constitutive expression of a plant ferredoxin-like protein (pflp) enhances capacity of photosynthetic carbon assimilation in rice (Oryza sativa).

PubMed

Chang, Hsiang; Huang, Hsiang-En; Cheng, Chin-Fu; Ho, Mei-Hsuan; Ger, Mang-Jye

2017-04-01

The plant ferredoxin-like protein (PFLP) gene, cloned from sweet peppers predicted as an electron carrier in photosynthesis, shows high homology to the Fd-I sequence of Arabidopsis thaliana, Lycopersicon esculentum, Oryza sativa and Spinacia oleracea. Most of pflp related studies focused on anti-pathogenic effects, while less understanding for the effects in photosynthesis with physiological aspects, such as photosynthesis rate, and levels of carbohydrate metabolites. This project focuses on the effects of pflp overexpression on photosynthesis by physiological evaluations of carbon assimilation with significant higher levels of carbohydrates with higher photosynthesis efficiency. In this report, two independent transgenic lines of rice plants (designated as pflp-1 and pflp-2) were generated from non-transgenic TNG67 rice plant (WT). Both transgenic pflp rice plants exhibited enhanced photosynthesis efficiency, and gas exchange rates of photosynthesis were 1.3- and 1.2-fold higher for pflp-1 and pflp-2 than WT respectively. Significantly higher electron transport rates of pflp rice plants were observed. Moreover, photosynthetic products, such as fructose, glucose, sucrose and starch contents of pflp transgenic lines were increased accordingly. Molecular evidences of carbohydrate metabolism related genes activities (osHXK5, osHXK6, osAGPL3, osAGPS2α, osSPS, ospFBPase, oscFBPase, and osSBPase) in transgenic lines were higher than those of WT. For performance of crop production, 1000-grain weight for pflp-1 and pflp-2 rice plants were 52.9 and 41.1 g that were both significantly higher than 31.6 g for WT, and panicles weights were 1.4- and 1.2-fold higher than WT. Panicle number, tiller number per plants for pflp rice plants were all significantly higher compared with those of WT where there was no significant difference observed between two pflp rice plants. Taken altogether; this study demonstrated that constitutive pflp expression can improve rice production by enhancing the capacity of photosynthetic carbon assimilation.

Role of NleH, a Type III Secreted Effector from Attaching and Effacing Pathogens, in Colonization of the Bovine, Ovine, and Murine Gut▿

PubMed Central

Hemrajani, Cordula; Marches, Olivier; Wiles, Siouxsie; Girard, Francis; Dennis, Alison; Dziva, Francis; Best, Angus; Phillips, Alan D.; Berger, Cedric N.; Mousnier, Aurelie; Crepin, Valerie F.; Kruidenier, Laurens; Woodward, Martin J.; Stevens, Mark P.; La Ragione, Roberto M.; MacDonald, Thomas T.; Frankel, Gad

2008-01-01

The human pathogen enterohemorrhagic Escherichia coli (EHEC) O157:H7 colonizes human and animal gut via formation of attaching and effacing lesions. EHEC strains use a type III secretion system to translocate a battery of effector proteins into the mammalian host cell, which subvert diverse signal transduction pathways implicated in actin dynamics, phagocytosis, and innate immunity. The genomes of sequenced EHEC O157:H7 strains contain two copies of the effector protein gene nleH, which share 49% sequence similarity with the gene for the Shigella effector OspG, recently implicated in inhibition of migration of the transcriptional regulator NF-κB to the nucleus. In this study we investigated the role of NleH during EHEC O157:H7 infection of calves and lambs. We found that while EHEC ΔnleH colonized the bovine gut more efficiently than the wild-type strain, in lambs the wild-type strain exhibited a competitive advantage over the mutant during mixed infection. Using the mouse pathogen Citrobacter rodentium, which shares many virulence factors with EHEC O157:H7, including NleH, we observed that the wild-type strain exhibited a competitive advantage over the mutant during mixed infection. We found no measurable differences in T-cell infiltration or hyperplasia in colons of mice inoculated with the wild-type or the nleH mutant strain. Using NF-κB reporter mice carrying a transgene containing a luciferase reporter driven by three NF-κB response elements, we found that NleH causes an increase in NF-κB activity in the colonic mucosa. Consistent with this, we found that the nleH mutant triggered a significantly lower tumor necrosis factor alpha response than the wild-type strain. PMID:18725419

Modulators of heterogeneous protein surface water dynamics

NASA Astrophysics Data System (ADS)

Han, Songi

The hydration water that solvates proteins is a major factor in driving or enabling biological events, including protein-protein and protein-ligand interactions. We investigate the role of the protein surface in modulating the hydration water fluctuations on both the picosecond and nanosecond timescale with an emerging experimental NMR technique known as Overhauser Dynamic Nuclear Polarization (ODNP). We carry out site-specific ODNP measurements of the hydration water fluctuations along the surface of Chemotaxis Y (CheY), and correlate the measured fluctuations to hydropathic and topological properties of the CheY surface as derived from molecular dynamics (MD) simulation. Furthermore, we compare hydration water fluctuations measured on the CheY surface to that of other globular proteins, as well as intrinsically disordered proteins, peptides, and liposome surfaces to systematically test characteristic effects of the biomolecular surface on the hydration water dynamics. Our results suggest that the labile (ps) hydration water fluctuations are modulated by the chemical nature of the surface, while the bound (ns) water fluctuations are present on surfaces that feature a rough topology and chemical heterogeneity such as the surface of a folded and structured protein. In collaboration with: Ryan Barnes, Dept of Chemistry and Biochemistry, University of California Santa Barbara

Protein Adsorption in Three Dimensions

PubMed Central

Vogler, Erwin A.

2011-01-01

Recent experimental and theoretical work clarifying the physical chemistry of blood-protein adsorption from aqueous-buffer solution to various kinds of surfaces is reviewed and interpreted within the context of biomaterial applications, especially toward development of cardiovascular biomaterials. The importance of this subject in biomaterials surface science is emphasized by reducing the “protein-adsorption problem” to three core questions that require quantitative answer. An overview of the protein-adsorption literature identifies some of the sources of inconsistency among many investigators participating in more than five decades of focused research. A tutorial on the fundamental biophysical chemistry of protein adsorption sets the stage for a detailed discussion of the kinetics and thermodynamics of protein adsorption, including adsorption competition between two proteins for the same adsorbent immersed in a binary-protein mixture. Both kinetics and steady-state adsorption can be rationalized using a single interpretive paradigm asserting that protein molecules partition from solution into a three-dimensional (3D) interphase separating bulk solution from the physical-adsorbent surface. Adsorbed protein collects in one-or-more adsorbed layers, depending on protein size, solution concentration, and adsorbent surface energy (water wettability). The adsorption process begins with the hydration of an adsorbent surface brought into contact with an aqueous-protein solution. Surface hydration reactions instantaneously form a thin, pseudo-2D interface between the adsorbent and protein solution. Protein molecules rapidly diffuse into this newly-formed interface, creating a truly 3D interphase that inflates with arriving proteins and fills to capacity within milliseconds at mg/mL bulk-solution concentrations CB. This inflated interphase subsequently undergoes time-dependent (minutes-to-hours) decrease in volume VI by expulsion of either-or-both interphase water and initially-adsorbed protein. Interphase protein concentration CI increases as VI decreases, resulting in slow reduction in interfacial energetics. Steady-state is governed by a net partition coefficient P=(/CBCI). In the process of occupying space within the interphase, adsorbing protein molecules must displace an equivalent volume of interphase water. Interphase water is itself associated with surface-bound water through a network of transient hydrogen bonds. Displacement of interphase water thus requires an amount of energy that depends on the adsorbent surface chemistry/energy. This “adsorption-dehydration” step is the significant free-energy cost of adsorption that controls the maximum amount of protein that can be adsorbed at steady state to a unit adsorbent-surface area (the adsorbent capacity). As adsorbent hydrophilicity increases, protein adsorption monotonically decreases because the energetic cost of surface dehydration increases, ultimately leading to no protein adsorption near an adsorbent water wettability (surface energy) characterized by a water contact angle θ → 65°. Consequently, protein does not adsorb (accumulate at interphase concentrations greater than bulk solution) to more hydrophilic adsorbents exhibiting θ < 65° . For adsorbents bearing strong Lewis acid/base chemistry such as ion-exchange resins, protein/surface interactions can be highly favorable, causing protein to adsorb in multilayers in a relatively thick interphase. A straightforward, three-component free energy relationship captures salient features of protein adsorption to all surfaces predicting that the overall free energy of protein adsorption ΔGadso is a relatively small multiple of thermal energy for any surface chemistry (except perhaps for bioengineered surfaces bearing specific ligands for adsorbing protein) because a surface chemistry that interacts chemically with proteins must also interact with water through hydrogen bonding. In this way, water moderates protein adsorption to any surface by competing with adsorbing protein molecules. This Leading Opinion ends by proposing several changes to the protein-adsorption paradigm that might advance answers to the three core questions that frame the “protein-adsorption problem” that is so fundamental to biomaterials surface science. PMID:22088888

Mapping Protein Surface Accessibility via an Electron Transfer Dissociation Selectively Cleavable Hydrazone Probe*

PubMed Central

Vasicek, Lisa; O'Brien, John P.; Browning, Karen S.; Tao, Zhihua; Liu, Hung-Wen; Brodbelt, Jennifer S.

2012-01-01

A protein's surface influences its role in protein-protein interactions and protein-ligand binding. Mass spectrometry can be used to give low resolution structural information about protein surfaces and conformations when used in combination with derivatization methods that target surface accessible amino acid residues. However, pinpointing the resulting modified peptides upon enzymatic digestion of the surface-modified protein is challenging because of the complexity of the peptide mixture and low abundance of modified peptides. Here a novel hydrazone reagent (NN) is presented that allows facile identification of all modified surface residues through a preferential cleavage upon activation by electron transfer dissociation coupled with a collision activation scan to pinpoint the modified residue in the peptide sequence. Using this approach, the correlation between percent reactivity and surface accessibility is demonstrated for two biologically active proteins, wheat eIF4E and PARP-1 Domain C. PMID:22393264

Predicting stability of alpha-helical, orthogonal-bundle proteins on surfaces

NASA Astrophysics Data System (ADS)

Wei, Shuai; Knotts, Thomas A.

2010-09-01

The interaction of proteins with surfaces is a key phenomenon in many applications, but current understanding of the biophysics involved is lacking. At present, rational design of such emerging technologies is difficult as no methods or theories exist that correctly predict how surfaces influence protein behavior. Using molecular simulation and a coarse-grain model, this study illustrates for the first time that stability of proteins on surfaces can be correlated with tertiary structural elements for alpha-helical, orthogonal-bundle proteins. Results show that several factors contribute to stability on surfaces including the nature of the loop region where the tether is placed and the ability of the protein to freely rotate on the surface. A thermodynamic analysis demonstrates that surfaces stabilize proteins entropically and that any destabilization is an enthalpic effect. Moreover, the entropic effects are concentrated on the unfolded state of the protein while the ethalpic effects are focused on the folded state.

Deciphering fine molecular details of proteins' structure and function with a Protein Surface Topography (PST) method.

PubMed

Koromyslova, Anna D; Chugunov, Anton O; Efremov, Roman G

2014-04-28

Molecular surfaces are the key players in biomolecular recognition and interactions. Nowadays, it is trivial to visualize a molecular surface and surface-distributed properties in three-dimensional space. However, such a representation trends to be biased and ambiguous in case of thorough analysis. We present a new method to create 2D spherical projection maps of entire protein surfaces and manipulate with them--protein surface topography (PST). It permits visualization and thoughtful analysis of surface properties. PST helps to easily portray conformational transitions, analyze proteins' properties and their dynamic behavior, improve docking performance, and reveal common patterns and dissimilarities in molecular surfaces of related bioactive peptides. This paper describes basic usage of PST with an example of small G-proteins conformational transitions, mapping of caspase-1 intersubunit interface, and intrinsic "complementarity" in the conotoxin-acetylcholine binding protein complex. We suggest that PST is a beneficial approach for structure-function studies of bioactive peptides and small proteins.

Protein quantification on dendrimer-activated surfaces by using time-of-flight secondary ion mass spectrometry and principal component regression

NASA Astrophysics Data System (ADS)

Kim, Young-Pil; Hong, Mi-Young; Shon, Hyun Kyong; Chegal, Won; Cho, Hyun Mo; Moon, Dae Won; Kim, Hak-Sung; Lee, Tae Geol

2008-12-01

Interaction between streptavidin and biotin on poly(amidoamine) (PAMAM) dendrimer-activated surfaces and on self-assembled monolayers (SAMs) was quantitatively studied by using time-of-flight secondary ion mass spectrometry (ToF-SIMS). The surface protein density was systematically varied as a function of protein concentration and independently quantified using the ellipsometry technique. Principal component analysis (PCA) and principal component regression (PCR) were used to identify a correlation between the intensities of the secondary ion peaks and the surface protein densities. From the ToF-SIMS and ellipsometry results, a good linear correlation of protein density was found. Our study shows that surface protein densities are higher on dendrimer-activated surfaces than on SAMs surfaces due to the spherical property of the dendrimer, and that these surface protein densities can be easily quantified with high sensitivity in a label-free manner by ToF-SIMS.

ABI domain containing proteins contribute to surface protein display and cell division in Staphylococcus aureus

PubMed Central

Frankel, Matthew B.; Wojcik, Brandon; DeDent, Andrea C.; Missiakas, Dominique M.; Schneewind, Olaf

2012-01-01

Summary The human pathogen Staphyloccocus aureus requires cell wall anchored surface proteins to cause disease. During cell division, surface proteins with YSIRK signal peptides are secreted into the cross wall, a layer of newly synthesized peptidoglycan between separating daughter cells. The molecular determinants for the trafficking of surface proteins are, however, still unknown. We screened mutants with non-redundant transposon insertions by fluorescence-activated cell sorting for reduced deposition of protein A (SpA) into the staphylococcal envelope. Three mutants, each of which harbored transposon insertions in genes for transmembrane proteins, displayed greatly reduced envelope abundance of SpA and surface proteins with YSIRK signal peptides. Characterization of the corresponding mutations identified three transmembrane proteins with abortive infectivity (ABI) domains, elements first described in lactococci for their role in phage exclusion. Mutations in genes for ABI domain proteins, designated spdA, spdB and spdC (surface protein display), diminish the expression of surface proteins with YSIRK signal peptides, but not of precursor proteins with conventional signal peptides. spdA, spdB and spdC mutants display an increase in the thickness of cross walls and in the relative abundance of staphylococci with cross walls, suggesting that spd mutations may represent a possible link between staphylococcal cell division and protein secretion. PMID:20923422

ABI domain-containing proteins contribute to surface protein display and cell division in Staphylococcus aureus.

PubMed

Frankel, Matthew B; Wojcik, Brandon M; DeDent, Andrea C; Missiakas, Dominique M; Schneewind, Olaf

2010-10-01

The human pathogen Staphylococcus aureus requires cell wall anchored surface proteins to cause disease. During cell division, surface proteins with YSIRK signal peptides are secreted into the cross-wall, a layer of newly synthesized peptidoglycan between separating daughter cells. The molecular determinants for the trafficking of surface proteins are, however, still unknown. We screened mutants with non-redundant transposon insertions by fluorescence-activated cell sorting for reduced deposition of protein A (SpA) into the staphylococcal envelope. Three mutants, each of which harboured transposon insertions in genes for transmembrane proteins, displayed greatly reduced envelope abundance of SpA and surface proteins with YSIRK signal peptides. Characterization of the corresponding mutations identified three transmembrane proteins with abortive infectivity (ABI) domains, elements first described in lactococci for their role in phage exclusion. Mutations in genes for ABI domain proteins, designated spdA, spdB and spdC (surface protein display), diminish the expression of surface proteins with YSIRK signal peptides, but not of precursor proteins with conventional signal peptides. spdA, spdB and spdC mutants display an increase in the thickness of cross-walls and in the relative abundance of staphylococci with cross-walls, suggesting that spd mutations may represent a possible link between staphylococcal cell division and protein secretion. © 2010 Blackwell Publishing Ltd.

Enhanced protein retention on poly(caprolactone) via surface initiated polymerization of acrylamide

NASA Astrophysics Data System (ADS)

Ma, Yuhao; Cai, Mengtan; He, Liu; Luo, Xianglin

2016-01-01

To enhance the biocompatibility or extend the biomedical application of poly(caprolactone) (PCL), protein retention on PCL surface is often required. In this study, poly(acrylamide) (PAAm) brushes were grown from PCL surface via surface-initiated atom transfer radical polymerization (SI-ATRP) and served as a protein-capturing platform. Grafted PAAm was densely packed on surface and exhibited superior protein retention ability. Captured protein was found to be resistant to washing under detergent environment. Furthermore, protein structure after being captured was investigated by circular dichroism (CD) spectroscopy, and the CD spectra verified that secondary structure of captured proteins was maintained, indicating no denaturation of protein happened for retention process.

Significance of surface charge and shell material of superparamagnetic iron oxide nanoparticle (SPION) based core/shell nanoparticles on the composition of the protein corona.

PubMed

Sakulkhu, Usawadee; Mahmoudi, Morteza; Maurizi, Lionel; Coullerez, Geraldine; Hofmann-Amtenbrink, Margarethe; Vries, Marcel; Motazacker, Mahdi; Rezaee, Farhad; Hofmann, Heinrich

2015-02-01

As nanoparticles (NPs) are increasingly used in many applications their safety and efficient applications in nanomedicine have become concerns. Protein coronas on nanomaterials' surfaces can influence how the cell "recognizes" nanoparticles, as well as the in vitro and in vivo NPs' behaviors. The SuperParamagnetic Iron Oxide Nanoparticle (SPION) is one of the most prominent agents because of its superparamagnetic properties, which is useful for separation applications. To mimic surface properties of different types of NPs, a core-shell SPION library was prepared by coating with different surfaces: polyvinyl alcohol polymer (PVA) (positive, neutral and negative), SiO2 (positive and negative), titanium dioxide and metal gold. The SPIONs with different surfaces were incubated at a fixed serum : nanoparticle surface ratio, magnetically trapped and washed. The tightly bound proteins were quantified and identified. The surface charge has a great impact on protein adsorption, especially on PVA and silica where proteins preferred binding to the neutral and positively charged surfaces. The importance of surface material on protein adsorption was also revealed by preferential binding on TiO2 and gold coated SPION, even negatively charged. There is no correlation between the protein net charge and the nanoparticle surface charge on protein binding, nor direct correlation between the serum proteins' concentration and the proteins detected in the coronas.

3D-SURFER 2.0: web platform for real-time search and characterization of protein surfaces.

PubMed

Xiong, Yi; Esquivel-Rodriguez, Juan; Sael, Lee; Kihara, Daisuke

2014-01-01

The increasing number of uncharacterized protein structures necessitates the development of computational approaches for function annotation using the protein tertiary structures. Protein structure database search is the basis of any structure-based functional elucidation of proteins. 3D-SURFER is a web platform for real-time protein surface comparison of a given protein structure against the entire PDB using 3D Zernike descriptors. It can smoothly navigate the protein structure space in real-time from one query structure to another. A major new feature of Release 2.0 is the ability to compare the protein surface of a single chain, a single domain, or a single complex against databases of protein chains, domains, complexes, or a combination of all three in the latest PDB. Additionally, two types of protein structures can now be compared: all-atom-surface and backbone-atom-surface. The server can also accept a batch job for a large number of database searches. Pockets in protein surfaces can be identified by VisGrid and LIGSITE (csc) . The server is available at http://kiharalab.org/3d-surfer/.

Single-molecule resolution of protein dynamics on polymeric membrane surfaces: the roles of spatial and population heterogeneity.

PubMed

Langdon, Blake B; Mirhossaini, Roya B; Mabry, Joshua N; Sriram, Indira; Lajmi, Ajay; Zhang, Yanxia; Rojas, Orlando J; Schwartz, Daniel K

2015-02-18

Although polymeric membranes are widely used in the purification of protein pharmaceuticals, interactions between biomolecules and membrane surfaces can lead to reduced membrane performance and damage to the product. In this study, single-molecule fluorescence microscopy provided direct observation of bovine serum albumin (BSA) and human monoclonal antibody (IgG) dynamics at the interface between aqueous buffer and polymeric membrane materials including regenerated cellulose and unmodified poly(ether sulfone) (PES) blended with either polyvinylpyrrolidone (PVP), polyvinyl acetate-co-polyvinylpyrrolidone (PVAc-PVP), or polyethylene glycol methacrylate (PEGM) before casting. These polymer surfaces were compared with model surfaces composed of hydrophilic bare fused silica and hydrophobic trimethylsilane-coated fused silica. At extremely dilute protein concentrations (10(-3)-10(-7) mg/mL), protein surface exchange was highly dynamic with protein monomers desorbing from the surface within ∼1 s after adsorption. Protein oligomers (e.g., nonspecific dimers, trimers, or larger aggregates), although less common, remained on the surface for 5 times longer than monomers. Using newly developed super-resolution methods, we could localize adsorption sites with ∼50 nm resolution and quantify the spatial heterogeneity of the various surfaces. On a small anomalous subset of the adsorption sites, proteins adsorbed preferentially and tended to reside for significantly longer times (i.e., on "strong" sites). Proteins resided for shorter times overall on surfaces that were more homogeneous and exhibited fewer strong sites (e.g., PVAc-PVP/PES). We propose that strong surface sites may nucleate protein aggregation, initiated preferentially by protein oligomers, and accelerate ultrafiltration membrane fouling. At high protein concentrations (0.3-1.0 mg/mL), fewer strong adsorption sites were observed, and surface residence times were reduced. This suggests that at high concentrations, adsorbed proteins block strong sites from further protein adsorption. Importantly, this demonstrates that strong binding sites can be modified by changing solution conditions. Membrane surfaces are intrinsically heterogeneous; by employing single-molecule techniques, we have provided a new framework for understanding protein interactions with such surfaces.

Disturbed vesicular trafficking of membrane proteins in prion disease.

PubMed

Uchiyama, Keiji; Miyata, Hironori; Sakaguchi, Suehiro

2013-01-01

The pathogenic mechanism of prion diseases remains unknown. We recently reported that prion infection disturbs post-Golgi trafficking of certain types of membrane proteins to the cell surface, resulting in reduced surface expression of membrane proteins and abrogating the signal from the proteins. The surface expression of the membrane proteins was reduced in the brains of mice inoculated with prions, well before abnormal symptoms became evident. Prions or pathogenic prion proteins were mainly detected in endosomal compartments, being particularly abundant in recycling endosomes. Some newly synthesized membrane proteins are delivered to the surface from the Golgi apparatus through recycling endosomes, and some endocytosed membrane proteins are delivered back to the surface through recycling endosomes. These results suggest that prions might cause neuronal dysfunctions and cell loss by disturbing post-Golgi trafficking of membrane proteins via accumulation in recycling endosomes. Interestingly, it was recently shown that delivery of a calcium channel protein to the cell surface was impaired and its function was abrogated in a mouse model of hereditary prion disease. Taken together, these results suggest that impaired delivery of membrane proteins to the cell surface is a common pathogenic event in acquired and hereditary prion diseases.

Plasma protein adsorption to zwitterionic poly (carboxybetaine methacrylate) modified surfaces: chain chemistry and end-group effects on protein adsorption kinetics, adsorbed amounts and immunoblots.

PubMed

Abraham, Sinoj; Bahniuk, Markian S; Unsworth, Larry D

2012-12-01

Protein-surface interactions are crucial to the overall biocompatability of biomaterials, and are thought to be the impetus towards the adverse host responses such as blood coagulation and complement activation. Only a few studies hint at the ultra-low fouling potential of zwitterionic poly(carboxybetaine methacrylate) (PCBMA) grafted surfaces and, of those, very few systematically investigate their non-fouling behavior. In this work, single protein adsorption studies as well as protein adsorption from complex solutions (i.e. human plasma) were used to evaluate the non-fouling potential of PCBMA grafted silica wafers prepared by nitroxide-mediated free radical polymerization. PCBMAs used for surface grafting varied in charge separating spacer groups that influence the overall surface charges, and chain end-groups that influence the overall hydrophilicity, thereby, allows a better understanding of these effects towards the protein adsorption for these materials. In situ ellipsometry was used to quantify the adsorbed layer thickness and adsorption kinetics for the adsorption of four proteins from single protein buffer solutions, viz, lysozyme, α-lactalbumin, human serum albumin and fibrinogen. Total amount of protein adsorbed on surfaces differed as a function of surface properties and protein characteristics. Finally, immunoblots results showed that human plasma protein adsorption to these surfaces resulted, primarily, in the adsorption of human serum albumin, with total protein adsorbed amounts being the lowest for PCBMA-3 (TEMPO). It was apparent that surface charge and chain hydrophilicity directly influenced protein adsorption behavior of PCBMA systems and are promising materials for biomedical applications.

Precise control of surface electrostatic forces on polymer brush layers with opposite charges for resistance to protein adsorption.

PubMed

Sakata, Sho; Inoue, Yuuki; Ishihara, Kazuhiko

2016-10-01

Various molecular interaction forces are generated during protein adsorption process on material surfaces. Thus, it is necessary to control them to suppress protein adsorption and the subsequent cell and tissue responses. A series of binary copolymer brush layers were prepared via surface-initiated atom transfer radical polymerization, by mixing the cationic monomer unit and anionic monomer unit randomly in various ratios. Surface characterization revealed that the constructed copolymer brush layers exhibited an uniform super-hydrophilic nature and different surface potentials. The strength of the electrostatic interaction forces operating on these mixed-charge copolymer brush surfaces was evaluated quantitatively using force-versus-distance (f-d) curve measurements by atomic force microscopy (AFM) and probes modified by negatively charged carboxyl groups or positively charged amino groups. The electrostatic interaction forces were determined based on the charge ratios of the copolymer brush layers. Notably, the surface containing equivalent cationic/anionic monomer units hardly interacted with both the charged groups. Furthermore, the protein adsorption force and the protein adsorption mass on these surfaces were examined by AFM f-d curve measurement and surface plasmon resonance measurement, respectively. To clarify the influence of the electrostatic interaction on the protein adsorption behavior on the surface, three kinds of proteins having negative, positive, and relatively neutral net charges under physiological conditions were used in this study. We quantitatively demonstrated that the amount of adsorbed proteins on the surfaces would have a strong correlation with the strength of surface-protein interaction forces, and that the strength of surface-protein interaction forces would be determined from the combination between the properties of the electrostatic interaction forces on the surfaces and the charge properties of the proteins. Especially, the copolymer brush surface composed of equivalent cationic/anionic monomer units exhibited no significant interaction forces, and dramatically suppressed the adsorption of proteins regardless of their charge properties. We conclude that the established methodology could elucidate relationship between the protein adsorption behavior and molecular interaction, especially the electrostatic interaction forces, and demonstrated that the suppression of the electrostatic interactions with the ionic functional groups would be important for the development of new polymeric biomaterials with a high repellency of protein adsorption. Copyright © 2016 Elsevier Ltd. All rights reserved.

Unraveling Gardnerella vaginalis Surface Proteins Using Cell Shaving Proteomics.

PubMed

Marín, Elvira; Haesaert, Annelies; Padilla, Laura; Adán, Jaume; Hernáez, María L; Monteoliva, Lucía; Gil, Concha

2018-01-01

Gardnerella vaginalis is one of the main etiologic agents of bacterial vaginosis (BV). This infection is responsible for a wide range of public health costs and is associated with several adverse outcomes during pregnancy. Improving our understanding of G. vaginalis protein cell surface will assist in BV diagnosis. This study represents the first proteomic approach that has analyzed the exposed proteins on G. vaginalis cell surface using a shaving approach. The 261 G. vaginalis proteins identified using this approach were analyzed with bioinformatic tools to detect characteristic motifs from surface-exposed proteins, such as signal peptides (36 proteins), lipobox domains (17 proteins), LPXTG motifs (5 proteins) and transmembrane alpha-helices (66 proteins). One third of the identified proteins were found to have at least one typical motif of surface-exposed proteins. Furthermore, the subcellular location was examined using two predictors (PSORT and Gpos-mPLoc). These bioinformatic tools classified 17% of the identified proteins as surface-associated proteins. Interestingly, we identified 13 members of the ATP-binding cassette (ABC) superfamily, which were mainly involved in the translocation of various substrates across membranes. To validate the location of the G. vaginalis surface-exposed proteins, an immunofluorescence assay with antibodies against Escherichia coli GroEL was performed to reveal the extracellular location of the moonlighting GroEL. In addition, monoclonal antibodies (mAb) against G. vaginalis Cna protein were produced and used to validate the location of Cna on the surface of the G. vaginalis . These high affinity anti-Cna mAb represent a useful tool for the study of this pathogenic microorganism and the BV.

Unraveling Gardnerella vaginalis Surface Proteins Using Cell Shaving Proteomics

PubMed Central

Marín, Elvira; Haesaert, Annelies; Padilla, Laura; Adán, Jaume; Hernáez, María L.; Monteoliva, Lucía; Gil, Concha

2018-01-01

Gardnerella vaginalis is one of the main etiologic agents of bacterial vaginosis (BV). This infection is responsible for a wide range of public health costs and is associated with several adverse outcomes during pregnancy. Improving our understanding of G. vaginalis protein cell surface will assist in BV diagnosis. This study represents the first proteomic approach that has analyzed the exposed proteins on G. vaginalis cell surface using a shaving approach. The 261 G. vaginalis proteins identified using this approach were analyzed with bioinformatic tools to detect characteristic motifs from surface-exposed proteins, such as signal peptides (36 proteins), lipobox domains (17 proteins), LPXTG motifs (5 proteins) and transmembrane alpha-helices (66 proteins). One third of the identified proteins were found to have at least one typical motif of surface-exposed proteins. Furthermore, the subcellular location was examined using two predictors (PSORT and Gpos-mPLoc). These bioinformatic tools classified 17% of the identified proteins as surface-associated proteins. Interestingly, we identified 13 members of the ATP-binding cassette (ABC) superfamily, which were mainly involved in the translocation of various substrates across membranes. To validate the location of the G. vaginalis surface-exposed proteins, an immunofluorescence assay with antibodies against Escherichia coli GroEL was performed to reveal the extracellular location of the moonlighting GroEL. In addition, monoclonal antibodies (mAb) against G. vaginalis Cna protein were produced and used to validate the location of Cna on the surface of the G. vaginalis. These high affinity anti-Cna mAb represent a useful tool for the study of this pathogenic microorganism and the BV. PMID:29867878

Fundamentals of nanoscale polymer-protein interactions and potential contributions to solid-state nanobioarrays.

PubMed

Hahm, Jong-in

2014-08-26

Protein adsorption onto polymer surfaces is a very complex, ubiquitous, and integrated process, impacting essential areas of food processing and packaging, health devices, diagnostic tools, and medical products. The nature of protein-surface interactions is becoming much more complicated with continuous efforts toward miniaturization, especially for the development of highly compact protein detection and diagnostic devices. A large body of literature reports on protein adsorption from the perspective of ensemble-averaged behavior on macroscopic, chemically homogeneous, polymeric surfaces. However, protein-surface interactions governing the nanoscale size regime may not be effectively inferred from their macroscopic and microscopic characteristics. Recently, research efforts have been made to produce periodically arranged, nanoscopic protein patterns on diblock copolymer surfaces solely through self-assembly. Intriguing protein adsorption phenomena are directly probed on the individual biomolecule level for a fundamental understanding of protein adsorption on nanoscale surfaces exhibiting varying degrees of chemical heterogeneity. Insight gained from protein assembly on diblock copolymers can be effectively used to control the surface density, conformation, orientation, and biofunctionality of prebound proteins in highly miniaturized applications, now approaching the nanoscale. This feature article will highlight recent experimental and theoretical advances made on these fronts while focusing on single-biomolecule-level investigations of protein adsorption behavior combined with surface chemical heterogeneity on the length scale commensurate with a single protein. This article will also address advantages and challenges of the self-assembly-driven patterning technology used to produce protein nanoarrays and its implications for ultrahigh density, functional, and quantifiable protein detection in a highly miniaturized format.

Non-interacting surface solvation and dynamics in protein-protein interactions.

PubMed

Visscher, Koen M; Kastritis, Panagiotis L; Bonvin, Alexandre M J J

2015-03-01

Protein-protein interactions control a plethora of cellular processes, including cell proliferation, differentiation, apoptosis, and signal transduction. Understanding how and why proteins interact will inevitably lead to novel structure-based drug design methods, as well as design of de novo binders with preferred interaction properties. At a structural and molecular level, interface and rim regions are not enough to fully account for the energetics of protein-protein binding, even for simple lock-and-key rigid binders. As we have recently shown, properties of the global surface might also play a role in protein-protein interactions. Here, we report on molecular dynamics simulations performed to understand solvent effects on protein-protein surfaces. We compare properties of the interface, rim, and non-interacting surface regions for five different complexes and their free components. Interface and rim residues become, as expected, less mobile upon complexation. However, non-interacting surface appears more flexible in the complex. Fluctuations of polar residues are always lower compared with charged ones, independent of the protein state. Further, stable water molecules are often observed around polar residues, in contrast to charged ones. Our analysis reveals that (a) upon complexation, the non-interacting surface can have a direct entropic compensation for the lower interface and rim entropy and (b) the mobility of the first hydration layer, which is linked to the stability of the protein-protein complex, is influenced by the local chemical properties of the surface. These findings corroborate previous hypotheses on the role of the hydration layer in shielding protein-protein complexes from unintended protein-protein interactions. © 2014 Wiley Periodicals, Inc.

Three steps to gold: mechanism of protein adsorption revealed by Brownian and molecular dynamics simulations.

PubMed

Ozboyaci, M; Kokh, D B; Wade, R C

2016-04-21

The addition of three N-terminal histidines to β-lactamase inhibitor protein was shown experimentally to increase its binding potency to an Au(111) surface substantially but the binding mechanism was not resolved. Here, we propose a complete adsorption mechanism for this fusion protein by means of a multi-scale simulation approach and free energy calculations. We find that adsorption is a three-step process: (i) recognition of the surface predominantly by the histidine fusion peptide and formation of an encounter complex facilitated by a reduced dielectric screening of water in the interfacial region, (ii) adsorption of the protein on the surface and adoption of a specific binding orientation, and (iii) adaptation of the protein structure on the metal surface accompanied by induced fit. We anticipate that the mechanistic features of protein adsorption to an Au(111) surface revealed here can be extended to other inorganic surfaces and proteins and will therefore aid the design of specific protein-surface interactions.

The electrokinetic behavior of calcium oxalate monohydrate in macromolecular solutions

NASA Technical Reports Server (NTRS)

Curreri, P. A.; Onoda, G. Y., Jr.; Finlayson, B.

1988-01-01

Electrophoretic mobilities were measured for calcium oxalate monohydrate (COM) in solutions containing macromolecules. Two mucopolysaccharides (sodium heparin and chrondroitin sulfate) and two proteins (positively charged lysozyme and negatively charged bovine serum albumin) were studied as adsorbates. The effects of pH, calcium oxalate surface charge (varied by calcium or oxalate ion activity), and citrate concentration were investigated. All four macromolecules showed evidence for chemical adsorption. The macromolecule concentrations needed for reversing the surface charge indicated that the mucopopolysacchrides have greater affinity for the COM surface than the proteins. The amount of proteins that can chemically adsorb appears to be limited to approximately one monomolecular layer. When the surface charge is high, an insufficient number of proteins can chemically adsorb to neutralize or reverse the surface charge. The remaining surface charge is balanced by proteins held near the surface by longer range electrostatic forces only. Citrate ions at high concentrations appear to compete effectively with the negative protein for surface sites but show no evidence for competing with the positively charged protein.

A coarse grain model for protein-surface interactions

NASA Astrophysics Data System (ADS)

Wei, Shuai; Knotts, Thomas A.

2013-09-01

The interaction of proteins with surfaces is important in numerous applications in many fields—such as biotechnology, proteomics, sensors, and medicine—but fundamental understanding of how protein stability and structure are affected by surfaces remains incomplete. Over the last several years, molecular simulation using coarse grain models has yielded significant insights, but the formalisms used to represent the surface interactions have been rudimentary. We present a new model for protein surface interactions that incorporates the chemical specificity of both the surface and the residues comprising the protein in the context of a one-bead-per-residue, coarse grain approach that maintains computational efficiency. The model is parameterized against experimental adsorption energies for multiple model peptides on different types of surfaces. The validity of the model is established by its ability to quantitatively and qualitatively predict the free energy of adsorption and structural changes for multiple biologically-relevant proteins on different surfaces. The validation, done with proteins not used in parameterization, shows that the model produces remarkable agreement between simulation and experiment.

Binding ligand prediction for proteins using partial matching of local surface patches.

PubMed

Sael, Lee; Kihara, Daisuke

2010-01-01

Functional elucidation of uncharacterized protein structures is an important task in bioinformatics. We report our new approach for structure-based function prediction which captures local surface features of ligand binding pockets. Function of proteins, specifically, binding ligands of proteins, can be predicted by finding similar local surface regions of known proteins. To enable partial comparison of binding sites in proteins, a weighted bipartite matching algorithm is used to match pairs of surface patches. The surface patches are encoded with the 3D Zernike descriptors. Unlike the existing methods which compare global characteristics of the protein fold or the global pocket shape, the local surface patch method can find functional similarity between non-homologous proteins and binding pockets for flexible ligand molecules. The proposed method improves prediction results over global pocket shape-based method which was previously developed by our group.

Binding Ligand Prediction for Proteins Using Partial Matching of Local Surface Patches

PubMed Central

Sael, Lee; Kihara, Daisuke

2010-01-01

Functional elucidation of uncharacterized protein structures is an important task in bioinformatics. We report our new approach for structure-based function prediction which captures local surface features of ligand binding pockets. Function of proteins, specifically, binding ligands of proteins, can be predicted by finding similar local surface regions of known proteins. To enable partial comparison of binding sites in proteins, a weighted bipartite matching algorithm is used to match pairs of surface patches. The surface patches are encoded with the 3D Zernike descriptors. Unlike the existing methods which compare global characteristics of the protein fold or the global pocket shape, the local surface patch method can find functional similarity between non-homologous proteins and binding pockets for flexible ligand molecules. The proposed method improves prediction results over global pocket shape-based method which was previously developed by our group. PMID:21614188

Noninvasive noble metal nanoparticle arrays for surface-enhanced Raman spectroscopy of proteins

NASA Astrophysics Data System (ADS)

Inya-Agha, Obianuju; Forster, Robert J.; Keyes, Tia E.

2007-02-01

Noble metal nanoparticles arrays are well established substrates for surface enhanced Raman spectroscopy (SERS). Their ability to enhance optical fields is based on the interaction of their surface valence electrons with incident electromagnetic radiation. In the array configuration, noble metal nanoparticles have been used to produce SER spectral enhancements of up to 10 8 orders of magnitude, making them useful for the trace analysis of physiologically relevant analytes such as proteins and peptides. Electrostatic interactions between proteins and metal surfaces result in the preferential adsorption of positively charged protein domains onto metal surfaces. This preferential interaction has the effect of disrupting the native conformation of the protein fold, with a concomitant loss of protein function. A major historic advantage of Raman microspectroscopy has been is its non-invasive nature; protein denaturation on the metal surfaces required for SER spectroscopy renders it a much more invasive technique. Further, part of the analytical power of Raman spectroscopy lies in its use as a secondary conformation probe. The protein structural loss which occurs on the metal surface results in secondary conformation readings which are not true to the actual native state of the analyte. This work presents a method for chemical fabrication of noble metal SERS arrays with surface immobilized layers which can protect protein native conformation without excessively mitigating the electromagnetic enhancements of spectra. Peptide analytes are used as model systems for proteins. Raman spectra of alpha lactalbumin on surfaces and when immobilized on these novel arrays are compared. We discuss the ability of the surface layer to protect protein structure whilst improving signal intensity.

Reversible Immobilization of Proteins in Sensors and Solid-State Nanopores.

PubMed

Ananth, Adithya; Genua, María; Aissaoui, Nesrine; Díaz, Leire; Eisele, Nico B; Frey, Steffen; Dekker, Cees; Richter, Ralf P; Görlich, Dirk

2018-05-01

The controlled functionalization of surfaces with proteins is crucial for many analytical methods in life science research and biomedical applications. Here, a coating for silica-based surfaces is established which enables stable and selective immobilization of proteins with controlled orientation and tunable surface density. The coating is reusable, retains functionality upon long-term storage in air, and is applicable to surfaces of complex geometry. The protein anchoring method is validated on planar surfaces, and then a method is developed to measure the anchoring process in real time using silicon nitride solid-state nanopores. For surface attachment, polyhistidine tags that are site specifically introduced into recombinant proteins are exploited, and the yeast nucleoporin Nsp1 is used as model protein. Contrary to the commonly used covalent thiol chemistry, the anchoring of proteins via polyhistidine tag is reversible, permitting to take proteins off and replace them by other ones. Such switching in real time in experiments on individual nanopores is monitored using ion conductivity. Finally, it is demonstrated that silica and gold surfaces can be orthogonally functionalized to accommodate polyhistidine-tagged proteins on silica but prevent protein binding to gold, which extends the applicability of this surface functionalization method to even more complex sensor devices. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A broad-range survey of ticks from livestock in Northern Xinjiang: changes in tick distribution and the isolation of Borrelia burgdorferi sensu stricto.

PubMed

Wang, Yuan-Zhi; Mu, Lu-Meng; Zhang, Ke; Yang, Mei-Hua; Zhang, Lin; Du, Jing-Yun; Liu, Zhi-Qiang; Li, Yong-Xiang; Lu, Wei-Hua; Chen, Chuang-Fu; Wang, Yan; Chen, Rong-Gui; Xu, Jun; Yuan, Li; Zhang, Wan-Jiang; Zuo, Wei-Ze; Shao, Ren-Fu

2015-09-04

Borreliosis is highly prevalent in Xinjiang Uygur Autonomous Region, China. However, little is known about the presence of Borrelia pathogens in tick species in this region, in addition Borrelia pathogens have not been isolated from domestic animals. We collected adult ticks from domestic animals at 19 sampling sites in 14 counties in northern Xinjiang from 2012 to 2014. Ticks were identified to species by morphology and were molecularly analysed by sequences of mitochondrial 16S rDNA gene; 4-8 ticks of each species at every sampling site were sequenced. 112 live adult ticks were selected for each species in every county, and were used to culture Borrelia pathogens; the genotypes were then determined by sequences of the 5S-23S rRNA intergenic spacer and the outer surface protein A (ospA) gene. A total of 5257 adult ticks, belonging to four genera and seven species, were collected. Compared with three decades ago, the abundance of the five common tick species during the peak ixodid tick season has changed. Certain tick species, such as Rhipicephalus turanicus (Rh. turanicus), was found at Jimusaer, Yining, Fukang, and Chabuchaer Counties for the first time. Additionally, the sequence analyses showed that the Hyalomma asiaticum (Hy. asiaticum), Haemaphysalis punctata (Ha. punctata), and Dermacentor marginatus (D. marginatus) that were collected from different sampling sites (≥3 sites) shared identical 16S rDNA sequences respectively. For the tick species that were collected from the same county, such as Hy. asiaticum from Shihezi County and Rh. turanicus from Yining County, their 16S rDNA sequences showed genetic diversity. In addition, sixteen Borrelia isolates were found in Hy. asiaticum, Ha. punctata, D. marginatus and Rh. turanicus, which infested cattle, sheep, horse and camel in Yining, Chabuchaer, Shihezi and Shawan Counties. All of the isolates were genetically identified as B. Burgdorferi sensu stricto. Warmer and wetter climate may have contributed to the altered distribution and abundance of the five most common ticks in northern Xinjiang. The genetic analyses showed that certain tick species, such as Hy. asiaticum or Rh. turanicus, exhibit genetic commonness or diversity. Additionally, this study is the first to isolate B. burgdorferi sensu stricto in Hy. asiaticum asiaticum, H. punctata, D. nuttalli and D. marginatus ticks from domestic animals. These ticks may transmit borreliosis among livestock.

Impact of hydrophilic and hydrophobic functionalization of flat TiO2/Ti surfaces on proteins adsorption

NASA Astrophysics Data System (ADS)

Fabre, Héloïse; Mercier, Dimitri; Galtayries, Anouk; Portet, David; Delorme, Nicolas; Bardeau, Jean-François

2018-02-01

Controlling adsorption of proteins onto medical devices is a key issue for implant-related infections. As self-assembled monolayers (SAMs) on titanium oxide represent a good model to study the surface-protein interactions, TiO2 surface properties were modified by grafting bisphosphonate molecules terminated with hydrophilic poly(ethylene glycol) groups and hydrophobic perfluoropolyether ones, respectively. Characterisation of the surface chemistry and surface topography of the modified surfaces was performed using XPS and atomic force microscopy (AFM). Quartz-crystal microbalance with dissipation (QCM-D) was used to determine the mass of adsorbed proteins as well as its kinetics. Poly(ethylene glycol)-terminated SAMs were the most effective surfaces to limit the adsorption of both BSA and fibrinogen in comparison to perfluorinated-terminated SAMs and non-modified TiO2 surfaces, as expected. The adsorption was not reversible in the case of BSA, while a partial reversibility was observed with Fg, most probably due to multilayers of proteins. The grafted surfaces adsorbed about the same quantity of proteins in terms of molecules per surface area, most probably in monolayer or island-like groups of adsorbed proteins. The adsorption on pristine TiO2 reveals a more important, non-specific adsorption of proteins.

Selective binding of proteins on functional nanoparticles via reverse charge parity model: an in vitro study

NASA Astrophysics Data System (ADS)

Ghosh, Goutam; Panicker, Lata; Barick, K. C.

2014-03-01

The conformation of proteins absorbed on nanoparticles surface plays a crucial role in applications of nanoparticles in biomedicine. The surface protein conformation depends on several factors, namely, nature of protein-nanoparticles interaction, chemical composition of the surface of nanoparticles etc. A model of the electrostatic binding of proteins on charged surface nanoparticles has been proposed earlier (Ghosh et al 2013 Colloids Surf. B 103 267). Also, the irreversible denaturation of the protein conformation due to binding of counterions was reported. In this paper, we have used this model, involving reverse charge parity, to show selective binding of proteins on charged surface iron oxide nanoparticles (IONPs). IONPs were surface functionalized with cetylpyridinium chloride (CPC), cetyl(trimethyl)ammonium bromide (CTAB) and cetylpyridinium iodide (CPI). The effect of counterions (Cl-, Br- and I-) on protein conformation has also been investigated. Several proteins such as α-lactalbumin (ALA), β-lactoglobulin (BLG), ovalbumin (OVA), bovin serum albumin (BSA) and HEWL were chosen for this investigation.

Surface modification of protein enhances encapsulation in chitosan nanoparticles

NASA Astrophysics Data System (ADS)

Koyani, Rina D.; Andrade, Mariana; Quester, Katrin; Gaytán, Paul; Huerta-Saquero, Alejandro; Vazquez-Duhalt, Rafael

2018-04-01

Chitosan nanoparticles have a huge potential as nanocarriers for environmental and biomedical purposes. Protein encapsulation in nano-sized chitosan provides protection against inactivation, proteolysis, and other alterations due to environmental conditions, as well as the possibility to be targeted to specific tissues by ligand functionalization. In this work, we demonstrate that the chemical modification of the protein surface enhances the protein loading in chitosan nanocarriers. Encapsulation of green fluorescent protein and the cytochrome P450 was studied. The increase of electrostatic interactions between the free amino groups of chitosan and the increased number of free carboxylic groups in the protein surface enhance the protein loading, protein retention, and, thus, the enzymatic activity of chitosan nanoparticles. The chemical modification of protein surface with malonic acid moieties reduced drastically the protein isoelectric point increasing the protein interaction with the polycationic biomaterial and chitosan. The chemical modification of protein does not alter the morphology of chitosan nanoparticles that showed an average diameter of 18 nm, spheroidal in shape, and smooth surfaced. The strategy of chemical modification of protein surface, shown here, is a simple and efficient technique to enhance the protein loading in chitosan nanoparticles. This technique could be used for other nanoparticles based on polycationic or polyanionic materials. The increase of protein loading improves, doubtless, the performance of protein-loaded chitosan nanoparticles for biotechnological and biomedical applications.

Presence of closely spaced protein thiols on the surface of mammalian cells.

PubMed Central

Donoghue, N.; Yam, P. T.; Jiang, X. M.; Hogg, P. J.

2000-01-01

It has been proposed that certain cell-surface proteins undergo redox reactions, that is, transfer of hydrogens and electrons between closely spaced cysteine thiols that can lead to reduction, formation, or interchange of disulfide bonds. This concept was tested using a membrane-impermeable trivalent arsenical to identify closely spaced thiols in cell-surface proteins. We attached the trivalent arsenical, phenylarsenoxide, to the thiol of reduced glutathione to produce 4-(N-(S-glutathionylacetyl)amino)phenylarsenoxide (GSAO). GSAO bound tightly to synthetic, peptide, and protein dithiols like thioredoxin, but not to monothiols. To identify cell-surface proteins that contain closely spaced thiols, we attached a biotin moiety through a spacer arm to the primary amino group of the gamma-glutamyl residue of GSAO (GSAO-B). Incorporation of GSAO-B into proteins was assessed by measuring the biotin using streptavidin-peroxidase. Up to 12 distinct proteins were labeled with GSAO-B on the surface of endothelial and fibrosarcoma cells. The pattern of labeled proteins differed between the different cell types. Protein disulfide isomerase was one of the proteins on the endothelial and fibrosarcoma cell surface that incorporated GSAO-B. These findings demonstrate that the cell-surface environment can support the existence of closely spaced protein thiols and suggest that at least some of these thiols are redox active. PMID:11206065

The effect of polymer surface modification on polymer-protein interaction via interfacial polymerization and hydrophilic polymer grafting

USDA-ARS?s Scientific Manuscript database

Protein membrane separation is prone to fouling on the membrane surface resulting from protein adsorption onto the surface. Surface modification of synthetic membranes is one way to reduce fouling. We investigated surface modification of polyethersulfone (PES) as a way of improving hydrophilicity ...

Effect of fullerenol surface chemistry on nanoparticle binding-induced protein misfolding

NASA Astrophysics Data System (ADS)

Radic, Slaven; Nedumpully-Govindan, Praveen; Chen, Ran; Salonen, Emppu; Brown, Jared M.; Ke, Pu Chun; Ding, Feng

2014-06-01

Fullerene and its derivatives with different surface chemistry have great potential in biomedical applications. Accordingly, it is important to delineate the impact of these carbon-based nanoparticles on protein structure, dynamics, and subsequently function. Here, we focused on the effect of hydroxylation -- a common strategy for solubilizing and functionalizing fullerene -- on protein-nanoparticle interactions using a model protein, ubiquitin. We applied a set of complementary computational modeling methods, including docking and molecular dynamics simulations with both explicit and implicit solvent, to illustrate the impact of hydroxylated fullerenes on the structure and dynamics of ubiquitin. We found that all derivatives bound to the model protein. Specifically, the more hydrophilic nanoparticles with a higher number of hydroxyl groups bound to the surface of the protein via hydrogen bonds, which stabilized the protein without inducing large conformational changes in the protein structure. In contrast, fullerene derivatives with a smaller number of hydroxyl groups buried their hydrophobic surface inside the protein, thereby causing protein denaturation. Overall, our results revealed a distinct role of surface chemistry on nanoparticle-protein binding and binding-induced protein misfolding.Fullerene and its derivatives with different surface chemistry have great potential in biomedical applications. Accordingly, it is important to delineate the impact of these carbon-based nanoparticles on protein structure, dynamics, and subsequently function. Here, we focused on the effect of hydroxylation -- a common strategy for solubilizing and functionalizing fullerene -- on protein-nanoparticle interactions using a model protein, ubiquitin. We applied a set of complementary computational modeling methods, including docking and molecular dynamics simulations with both explicit and implicit solvent, to illustrate the impact of hydroxylated fullerenes on the structure and dynamics of ubiquitin. We found that all derivatives bound to the model protein. Specifically, the more hydrophilic nanoparticles with a higher number of hydroxyl groups bound to the surface of the protein via hydrogen bonds, which stabilized the protein without inducing large conformational changes in the protein structure. In contrast, fullerene derivatives with a smaller number of hydroxyl groups buried their hydrophobic surface inside the protein, thereby causing protein denaturation. Overall, our results revealed a distinct role of surface chemistry on nanoparticle-protein binding and binding-induced protein misfolding. Electronic supplementary information (ESI) is available: Fluorescence spectra, ITC, CD spectra and other data as described in the text. See DOI: 10.1039/c4nr01544d

Fundamentals of Nanoscale Polymer–Protein Interactions and Potential Contributions to Solid-State Nanobioarrays

PubMed Central

2015-01-01

Protein adsorption onto polymer surfaces is a very complex, ubiquitous, and integrated process, impacting essential areas of food processing and packaging, health devices, diagnostic tools, and medical products. The nature of protein–surface interactions is becoming much more complicated with continuous efforts toward miniaturization, especially for the development of highly compact protein detection and diagnostic devices. A large body of literature reports on protein adsorption from the perspective of ensemble-averaged behavior on macroscopic, chemically homogeneous, polymeric surfaces. However, protein–surface interactions governing the nanoscale size regime may not be effectively inferred from their macroscopic and microscopic characteristics. Recently, research efforts have been made to produce periodically arranged, nanoscopic protein patterns on diblock copolymer surfaces solely through self-assembly. Intriguing protein adsorption phenomena are directly probed on the individual biomolecule level for a fundamental understanding of protein adsorption on nanoscale surfaces exhibiting varying degrees of chemical heterogeneity. Insight gained from protein assembly on diblock copolymers can be effectively used to control the surface density, conformation, orientation, and biofunctionality of prebound proteins in highly miniaturized applications, now approaching the nanoscale. This feature article will highlight recent experimental and theoretical advances made on these fronts while focusing on single-biomolecule-level investigations of protein adsorption behavior combined with surface chemical heterogeneity on the length scale commensurate with a single protein. This article will also address advantages and challenges of the self-assembly-driven patterning technology used to produce protein nanoarrays and its implications for ultrahigh density, functional, and quantifiable protein detection in a highly miniaturized format. PMID:24456577

Restricted mobility of side chains on concave surfaces of solenoid proteins may impart heightened potential for intermolecular interactions.

PubMed

Ramya, L; Gautham, N; Chaloin, Laurent; Kajava, Andrey V

2015-09-01

Significant progress has been made in the determination of the protein structures with their number today passing over a hundred thousand structures. The next challenge is the understanding and prediction of protein-protein and protein-ligand interactions. In this work we address this problem by analyzing curved solenoid proteins. Many of these proteins are considered as "hub molecules" for their high potential to interact with many different molecules and to be a scaffold for multisubunit protein machineries. Our analysis of these structures through molecular dynamics simulations reveals that the mobility of the side-chains on the concave surfaces of the solenoids is lower than on the convex ones. This result provides an explanation to the observed preferential binding of the ligands, including small and flexible ligands, to the concave surface of the curved solenoid proteins. The relationship between the landscapes and dynamic properties of the protein surfaces can be further generalized to the other types of protein structures and eventually used in the computer algorithms, allowing prediction of protein-ligand interactions by analysis of protein surfaces. © 2015 Wiley Periodicals, Inc.

Correlation between surface morphology and surface forces of protein A adsorbed on mica.

PubMed Central

Ohnishi, S; Murata, M; Hato, M

1998-01-01

We have investigated the morphology and surface forces of protein A adsorbed on mica surface in the protein solutions of various concentrations. The force-distance curves, measured with a surface force apparatus (SFA), were interpreted in terms of two different regimens: a "large-distance" regimen in which an electrostatic double-layer force dominates, and an "adsorbed layer" regimen in which a force of steric origin dominates. To further clarify the forces of steric origin, the surface morphology of the adsorbed protein layer was investigated with an atomic force microscope (AFM) because the steric repulsive forces are strongly affected by the adsorption mode of protein A molecules on mica. At lower protein concentrations (2 ppm, 10 ppm), protein A molecules were adsorbed "side-on" parallel to the mica surfaces, forming a monolayer of approximately 2.5 nm. AFM images at higher concentrations (30 ppm, 100 ppm) showed protruding structures over the monolayer, which revealed that the adsorbed protein A molecules had one end oriented into the solution, with the remainder of each molecule adsorbed side-on to the mica surface. These extending ends of protein A overlapped each other and formed a "quasi-double layer" over the mica surface. These AFM images proved the existence of a monolayer of protein A molecules at low concentrations and a "quasi-double layer" with occasional protrusions at high concentrations, which were consistent with the adsorption mode observed in the force-distance curves. PMID:9449346

Quantification of the Influence of Protein-Protein Interactions on Adsorbed Protein Structure and Bioactivity

PubMed Central

Wei, Yang; Thyparambil, Aby A.; Latour, Robert A.

2013-01-01

While protein-surface interactions have been widely studied, relatively little is understood at this time regarding how protein-surface interaction effects are influenced by protein-protein interactions and how these effects combine with the internal stability of a protein to influence its adsorbed-state structure and bioactivity. The objectives of this study were to develop a method to study these combined effects under widely varying protein-protein interaction conditions using hen egg-white lysozyme (HEWL) adsorbed on silica glass, poly(methyl methacrylate), and polyethylene as our model systems. In order to vary protein-protein interaction effects over a wide range, HEWL was first adsorbed to each surface type under widely varying protein solution concentrations for 2 h to saturate the surface, followed by immersion in pure buffer solution for 15 h to equilibrate the adsorbed protein layers in the absence of additionally adsorbing protein. Periodic measurements were made at selected time points of the areal density of the adsorbed protein layer as an indicator of the level of protein-protein interaction effects within the layer, and these values were then correlated with measurements of the adsorbed protein’s secondary structure and bioactivity. The results from these studies indicate that protein-protein interaction effects help stabilize the structure of HEWL adsorbed on silica glass, have little influence on the structural behavior of HEWL on HDPE, and actually serve to destabilize HEWL’s structure on PMMA. The bioactivity of HEWL on silica glass and HDPE was found to decrease in direct proportion to the degree of adsorption-induce protein unfolding. A direct correlation between bioactivity and the conformational state of adsorbed HEWL was less apparent on PMMA, thus suggesting that other factors influenced HEWL’s bioactivity on this surface, such as the accessibility of HEWL’s bioactive site being blocked by neighboring proteins or the surface itself. The developed methods provide an effective means to characterize the influence of protein-protein interaction effects and provide new molecular-level insights into how protein-protein interaction effects combine with protein-surface interaction and internal protein stability effects to influence the structure and bioactivity of adsorbed protein. PMID:23751416

Protein Surface Mimetics: Understanding How Ruthenium Tris(Bipyridines) Interact with Proteins.

PubMed

Hewitt, Sarah H; Filby, Maria H; Hayes, Ed; Kuhn, Lars T; Kalverda, Arnout P; Webb, Michael E; Wilson, Andrew J

2017-01-17

Protein surface mimetics achieve high-affinity binding by exploiting a scaffold to project binding groups over a large area of solvent-exposed protein surface to make multiple cooperative noncovalent interactions. Such recognition is a prerequisite for competitive/orthosteric inhibition of protein-protein interactions (PPIs). This paper describes biophysical and structural studies on ruthenium(II) tris(bipyridine) surface mimetics that recognize cytochrome (cyt) c and inhibit the cyt c/cyt c peroxidase (CCP) PPI. Binding is electrostatically driven, with enhanced affinity achieved through enthalpic contributions thought to arise from the ability of the surface mimetics to make a greater number of noncovalent interactions than CCP with surface-exposed basic residues on cyt c. High-field natural abundance 1 H, 15 N HSQC NMR experiments are consistent with surface mimetics binding to cyt c in similar manner to CCP. This provides a framework for understanding recognition of proteins by supramolecular receptors and informing the design of ligands superior to the protein partners upon which they are inspired. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

AFM study of adsorption of protein A on a poly(dimethylsiloxane) surface

NASA Astrophysics Data System (ADS)

Yu, Ling; Lu, Zhisong; Gan, Ye; Liu, Yingshuai; Li, Chang Ming

2009-07-01

In this paper, the morphology and kinetics of adsorption of protein A on a PDMS surface is studied by AFM. The results of effects of pH, protein concentration and contact time of the adsorption reveal that the morphology of adsorbed protein A is significantly affected by pH and adsorbed surface concentration, in which the pH away from the isoelectric point (IEP) of protein A could produce electrical repulsion to change the protein conformation, while the high adsorbed surface protein volume results in molecular networks. Protein A can form an adsorbed protein film on PDMS with a maximum volume of 2.45 × 10-3 µm3. This work enhances our fundamental understanding of protein A adsorption on PDMS, a frequently used substrate component in miniaturized immunoassay devices.

Proteins at the Biomaterial Electrolyte Interface

NASA Astrophysics Data System (ADS)

Tengvall, Pentti

2005-03-01

Proteins adsorb rapidly onto solid and polymeric surfaces because the association process is in the vast majority of cases energetically favourable, i.e. exothermic. The most common exceptions to this rule are hydrophilic interfaces with low net charge and high mobility, e.g. immobilized PEGs. Current research in the research area tries to understand and control unwanted and wanted adsorption by studying the adsorption kinetics, protein surface binding specificity, protein exchange at interfaces, and surface protein repulsion mechanisms. In blood plasma model systems humoral cascade reactions such as surface mediated coagulation and immune complement raise considerable interest due to the immediate association to blood compatibility, and in tissue applications the binding between surfaces and membrane receptors in cells and tissues. Thus, the understanding of interfacial events at the protein level is of large importance in applications such as blood and tissue contacting biomaterials, in vitro medical and biological diagnostics, food industry and in marine anti-fouling technology. Well described consequences of adsorption are a lowered system energy, increased system entropy, irreversible binding, conformational changes, specific surface/protein interactions, and in biomedical materials applications surface opsonization followed by cell-surface interactions and a host tissue response. This lecture will deal with some mechanisms known to be of importance for the adsorption processes, such as the influence of surface chemistry and surface energy, the composition of the protein solution, the Vroman effect, and residence time. Examples will be shown from ellipsometric experiments using different model surfaces in single/few protein solutions, and specific attention be given to blood serum and plasma experiments on coagulation and immune complement at interfaces.

Understanding the interfacial behavior of lysozyme on Au (111) surfaces with multiscale simulations

NASA Astrophysics Data System (ADS)

Samieegohar, Mohammadreza; Ma, Heng; Sha, Feng; Jahan Sajib, Md Symon; Guerrero-García, G. Iván; Wei, Tao

2017-02-01

The understanding of the adsorption and interfacial behavior of proteins is crucial to the development of novel biosensors and biomaterials. By using bottom-up atomistic multiscale simulations, we study here the adsorption of lysozyme on Au(111) surfaces in an aqueous environment. Atomistic simulations are used to calculate the inhomogeneous polarization of the gold surface, which is induced by the protein adsorption, and by the presence of an interfacial layer of water molecules and monovalent salts. The corresponding potential of mean force between the protein and the gold surface including polarization effects is used in Langevin Dynamics simulations to study the time dependent behavior of proteins at finite concentration. These simulations display a rapid adsorption and formation of a first-layer of proteins at the interface. Proteins are initially adsorbed directly on the gold surface due to the strong protein-surface attractive interaction. A subsequent interfacial weak aggregation of proteins leading to multilayer build-up is also observed at long times.

Surface Proteins of Gram-Positive Bacteria and Mechanisms of Their Targeting to the Cell Wall Envelope

PubMed Central

Navarre, William Wiley; Schneewind, Olaf

1999-01-01

The cell wall envelope of gram-positive bacteria is a macromolecular, exoskeletal organelle that is assembled and turned over at designated sites. The cell wall also functions as a surface organelle that allows gram-positive pathogens to interact with their environment, in particular the tissues of the infected host. All of these functions require that surface proteins and enzymes be properly targeted to the cell wall envelope. Two basic mechanisms, cell wall sorting and targeting, have been identified. Cell well sorting is the covalent attachment of surface proteins to the peptidoglycan via a C-terminal sorting signal that contains a consensus LPXTG sequence. More than 100 proteins that possess cell wall-sorting signals, including the M proteins of Streptococcus pyogenes, protein A of Staphylococcus aureus, and several internalins of Listeria monocytogenes, have been identified. Cell wall targeting involves the noncovalent attachment of proteins to the cell surface via specialized binding domains. Several of these wall-binding domains appear to interact with secondary wall polymers that are associated with the peptidoglycan, for example teichoic acids and polysaccharides. Proteins that are targeted to the cell surface include muralytic enzymes such as autolysins, lysostaphin, and phage lytic enzymes. Other examples for targeted proteins are the surface S-layer proteins of bacilli and clostridia, as well as virulence factors required for the pathogenesis of L. monocytogenes (internalin B) and Streptococcus pneumoniae (PspA) infections. In this review we describe the mechanisms for both sorting and targeting of proteins to the envelope of gram-positive bacteria and review the functions of known surface proteins. PMID:10066836

Surface Passivation for Single-molecule Protein Studies

PubMed Central

Chandradoss, Stanley D.; Haagsma, Anna C.; Lee, Young Kwang; Hwang, Jae-Ho; Nam, Jwa-Min; Joo, Chirlmin

2014-01-01

Single-molecule fluorescence spectroscopy has proven to be instrumental in understanding a wide range of biological phenomena at the nanoscale. Important examples of what this technique can yield to biological sciences are the mechanistic insights on protein-protein and protein-nucleic acid interactions. When interactions of proteins are probed at the single-molecule level, the proteins or their substrates are often immobilized on a glass surface, which allows for a long-term observation. This immobilization scheme may introduce unwanted surface artifacts. Therefore, it is essential to passivate the glass surface to make it inert. Surface coating using polyethylene glycol (PEG) stands out for its high performance in preventing proteins from non-specifically interacting with a glass surface. However, the polymer coating procedure is difficult, due to the complication arising from a series of surface treatments and the stringent requirement that a surface needs to be free of any fluorescent molecules at the end of the procedure. Here, we provide a robust protocol with step-by-step instructions. It covers surface cleaning including piranha etching, surface functionalization with amine groups, and finally PEG coating. To obtain a high density of a PEG layer, we introduce a new strategy of treating the surface with PEG molecules over two rounds, which remarkably improves the quality of passivation. We provide representative results as well as practical advice for each critical step so that anyone can achieve the high quality surface passivation. PMID:24797261

Role of Transbilayer Distribution of Lipid Molecules on the Structure and Protein-Lipid Interaction of an Amyloidogenic Protein on the Membrane Surface

NASA Astrophysics Data System (ADS)

Cheng, Kwan; Cheng, Sara

We used molecular dynamics simulations to examine the effects of transbilayer distribution of lipid molecules, particularly anionic lipids with negatively charged headgroups, on the structure and binding kinetics of an amyloidogenic protein on the membrane surface and subsequent protein-induced structural disruption of the membrane. Our systems consisted of a model beta-sheet rich dimeric protein absorbed on asymmetric bilayers with neutral and anionic lipids and symmetric bilayers with neutral lipids. We observed larger folding, domain aggregation, and tilt angle of the absorbed protein on the asymmetric bilayer surfaces. We also detected more focused bilayer thinning in the asymmetric bilayer due to weak lipid-protein interactions. Our results support the mechanism that the higher lipid packing in the protein-contacting lipid leaflet promotes stronger protein-protein but weaker protein-lipid interactions of an amyloidogenic protein on the membrane surface. We speculate that the observed surface-induced structural and protein-lipid interaction of our model amyloidogenic protein may play a role in the early membrane-associated amyloid cascade pathway that leads to membrane structural damage of neurons in Alzheimer's disease. NSF ACI-1531594.

Titanium Surface Roughing Treatments contribute to Higher Interaction with Salivary Proteins MG2 and Lactoferrin.

PubMed

Cavalcanti, Yuri Wanderley; Soare, Rodrigo Villamarim; Leite Assis, Marina Araújo; Zenóbio, Elton Gonçalves; Girundi, Francisco Mauro da Silva

2015-02-01

Some surface treatments performed on titanium can alter the composition of salivary pellicle formed on this abiotic surface. Such treatments modify the titanium's surface properties and can promote higher adsorption of proteins, which allow better integration of titanium to the biotic system. This study aimed to evaluate the interactions between salivary proteins and titanium disks with different surface treatments. Machined titanium disks (n = 48) were divided into four experimental groups (n = 12), according to their surface treatments: surface polishing (SP); acid etching (A); spot-blasting plus acid etching (SB-A); spot-blasting followed by acid etching and nano-functionalization (SB-A-NF). Titanium surfaces were characterized by surface roughness and scanning electron microscopy (SEM). Specimens were incubated with human saliva extracted from submandibular and sublingual glands. Total salivary protein adsorbed to titanium was quantified and samples were submitted to western blotting for mucin glycoprotein 2 (MG2) and lactoferrin identification. Surface roughness was statistically higher for SB-A and SB-A-NF groups. Scanning electron microscopy images confirmed that titanium surface treatments increased surface roughness with higher number of porous and scratches for SB-A and SB-A-NF groups. Total protein adsorption was significantly higher for SB-A and SB-A-NF groups (p < 0.05), which also presented higher interactions with MG2 and lactoferrin proteins. The roughing of titanium surface by spot-blasting plus acid etching treatments contribute to higher interaction with salivary proteins, such as MG2 and lactoferrin. Titanium surface roughing increases the interactions of the substratum with salivary proteins, which can influence the integration of dental implants and their components to the oral environment. However, those treatments should be used carefully intraorally, avoiding increase biofilm formation.

Targeted Proteomics and Absolute Protein Quantification for the Construction of a Stoichiometric Host-Pathogen Surface Density Model*

PubMed Central

Sjöholm, Kristoffer; Kilsgård, Ola; Teleman, Johan; Happonen, Lotta; Malmström, Lars; Malmström, Johan

2017-01-01

Sepsis is a systemic immune response responsible for considerable morbidity and mortality. Molecular modeling of host-pathogen interactions in the disease state represents a promising strategy to define molecular events of importance for the transition from superficial to invasive infectious diseases. Here we used the Gram-positive bacterium Streptococcus pyogenes as a model system to establish a mass spectrometry based workflow for the construction of a stoichiometric surface density model between the S. pyogenes surface, the surface virulence factor M-protein, and adhered human blood plasma proteins. The workflow relies on stable isotope labeled reference peptides and selected reaction monitoring mass spectrometry analysis of a wild-type strain and an M-protein deficient mutant strain, to generate absolutely quantified protein stoichiometry ratios between S. pyogenes and interacting plasma proteins. The stoichiometry ratios in combination with a novel targeted mass spectrometry method to measure cell numbers enabled the construction of a stoichiometric surface density model using protein structures available from the protein data bank. The model outlines the topology and density of the host-pathogen protein interaction network on the S. pyogenes bacterial surface, revealing a dense and highly organized protein interaction network. Removal of the M-protein from S. pyogenes introduces a drastic change in the network topology, validated by electron microscopy. We propose that the stoichiometric surface density model of S. pyogenes in human blood plasma represents a scalable framework that can continuously be refined with the emergence of new results. Future integration of new results will improve the understanding of protein-protein interactions and their importance for bacterial virulence. Furthermore, we anticipate that the general properties of the developed workflow will facilitate the production of stoichiometric surface density models for other types of host-pathogen interactions. PMID:28183813

Controlling adsorption and passivation properties of bovine serum albumin on silica surfaces by ionic strength modulation and cross-linking.

PubMed

Park, Jae Hyeon; Sut, Tun Naw; Jackman, Joshua A; Ferhan, Abdul Rahim; Yoon, Bo Kyeong; Cho, Nam-Joon

2017-03-29

Understanding the physicochemical factors that influence protein adsorption onto solid supports holds wide relevance for fundamental insights into protein structure and function as well as for applications such as surface passivation. Ionic strength is a key parameter that influences protein adsorption, although how its modulation might be utilized to prepare well-coated protein adlayers remains to be explored. Herein, we investigated how ionic strength can be utilized to control the adsorption and passivation properties of bovine serum albumin (BSA) on silica surfaces. As protein stability in solution can influence adsorption kinetics, the size distribution and secondary structure of proteins in solution were first characterized by dynamic light scattering (DLS), nanoparticle tracking analysis (NTA), and circular dichroism (CD) spectroscopy. A non-monotonic correlation between ionic strength and protein aggregation was observed and attributed to colloidal agglomeration, while the primarily α-helical character of the protein in solution was maintained in all cases. Quartz crystal microbalance-dissipation (QCM-D) experiments were then conducted in order to track protein adsorption onto silica surfaces as a function of ionic strength, and the measurement responses indicated that total protein uptake at saturation coverage is lower with increasing ionic strength. In turn, the QCM-D data and the corresponding Voigt-Voinova model analysis support that the surface area per bound protein molecule is greater with increasing ionic strength. While higher protein uptake under lower ionic strengths by itself did not result in greater surface passivation under subsequent physiologically relevant conditions, the treatment of adsorbed protein layers with a gluteraldehyde cross-linking agent stabilized the bound protein in this case and significantly improved surface passivation. Collectively, our findings demonstrate that ionic strength modulation influences BSA adsorption uptake on account of protein spreading and can be utilized in conjunction with covalent cross-linking strategies to prepare well-coated protein adlayers for improved surface passivation.

RS-88 Pad Abort Demonstrator Thrust Chamber Assembly Testing at NASA Marshall Space Flight Center

NASA Technical Reports Server (NTRS)

Farr, Rebecca A.; Sanders, Timothy M.

1990-01-01

This paper documents the effort conducted to collect hot-tire dynamic and acoustics environments data during 50,000-lb thrust lox-ethanol hot-fire rocket testing at NASA Marshall Space Flight Center (MSFC) in November-December 2003. This test program was conducted during development testing of the Boeing Rocketdyne RS-88 development engine thrust chamber assembly (TCA) in support of the Orbital Space Plane (OSP) Crew Escape System Propulsion (CESP) Program Pad Abort Demonstrator (PAD). In addition to numerous internal TCA and nozzle measurements, induced acoustics environments data were also collected. Provided here is an overview of test parameters, a discussion of the measurements, test facility systems and test operations, and a quality assessment of the data collected during this test program.

Time series of the northeast Pacific

NASA Astrophysics Data System (ADS)

Peña, M. Angelica; Bograd, Steven J.

2007-10-01

In July 2006, the North Pacific Marine Science Organization (PICES) and Fisheries & Oceans Canada sponsored the symposium “Time Series of the Northeast Pacific: A symposium to mark the 50th anniversary of Line P”. The symposium, which celebrated 50 years of oceanography along Line P and at Ocean Station Papa (OSP), explored the scientific value of the Line P and other long oceanographic time series of the northeast Pacific (NEP). Overviews of the principal NEP time-series were presented, which facilitated regional comparisons and promoted interaction and exchange of information among investigators working in the NEP. More than 80 scientists from 8 countries attended the symposium. This introductory essay is a brief overview of the symposium and the 10 papers that were selected for this special issue of Progress in Oceanography.

Optical monitoring of proteins at solid interfaces

NASA Astrophysics Data System (ADS)

Dunne, G.; McDonnell, L.; Miller, R.; McMillan, N. D.; O'Rourke, B.; Mitchell, C. I.

2005-06-01

The adsorption properties of polymers are of great importance for implant studies. A better understanding of these properties can lead to improved implant materials. In this study the surface energy of different polymers was derived from contact angle measurements taken using profile analysis tensiometry (PAT) of sessile drops of water. The contact angles were measured for advancing and receding water drops on polished polymer surfaces and also on polymer surfaces modified by adsorbing protein to the surface prior to analysis of the sessile drop. The protein used was bovine serum albumin (BSA) and the surfaces were poly-methylmethacrylate (PMMA), poly-ether-ether-ketone (PEEK) and stainless steel. The polymer surfaces were also studied using atomic force microscopy (AFM). Images of the surfaces were taken in different states: rough, smooth and with albumin adsorbed. As a method to identify the proteins on the surface easier, anti-albumin antibodies with 30nm nano gold particles attached were adsorbed to the albumin on the surfaces. Using nano gold particles made the imaging more straightforward and thus made identification of the protein on the surface easier. The results from this work show the differing hydrophobicities of polymer surfaces under different conditions and a new nanotechnological method of protein identification.

Effects of carbohydrate/protein ratio on the microstructure and the barrier and sorption properties of wheat starch-whey protein blend edible films.

PubMed

Basiak, Ewelina; Lenart, Andrzej; Debeaufort, Frédéric

2017-02-01

Starch and whey protein isolate and their mixtures were used for making edible films. Moisture sorption isotherms, water vapour permeability, sorption of aroma compounds, microstructure, water contact angle and surface properties were investigated. With increasing protein content, the microstructure changes became more homogeneous. The water vapour permeability increases with both the humidity gradient and the starch content. For all films, the hygroscopicity increases with starch content. Surface properties change according to the starch/whey protein ratio and are mainly related to the polar component of the surface tension. Films composed of 80% starch and 20% whey proteins have more hydrophobic surfaces than the other films due to specific interactions. The effect of carbohydrate/protein ratio significantly influences the microstructure, the surface wettability and the barrier properties of wheat starch-whey protein blend films. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Silica surface characterization as a function of formation and surface treatment using traditional methods and proteins as surface probes

NASA Astrophysics Data System (ADS)

Korwin-Edson, Michelle Lynn

Previous works have shown that cells proliferate differently depending on the chemistry of the glass on which they are growing. Since proteins form the bonds between cells and glass, the hypothesis of this study is that proteins can distinguish between surface chemical variations of glass. This theory was examined through the use of various silica forms, a few select proteins, four surface treatment procedures, and a variety of characterization techniques. The silica forms include amorphous slides, cane, fiber, microspheres, fumed silica and quartz crystal terminals. The proteins selected were human serum albumin, mouse Immunoglobulin G, streptavidin, antimouse IgG, and biotin. The surface treatments utilized to bring about chemical variation on the silica surface were HF acid etching, ethanol cleaning, water plasma treatments, and 1000°C heat treatments. The characterization techniques encompassed both traditional material techniques and biological methods. The techniques studied were atomic force microscopy (AFM), chemical force microscopy (CFM), glancing incidence X-ray analysis (GIXA), fluorescence spectrometry, polyacrylamide gel electrophoresis (SDS-PAGE), and bicinchoninic acid (BCA) assay. It was the main goal of this project to determine the feasibility of these techniques in utilizing proteins as glass surface probes. Proteins were adsorbed to all of the various forms and the binding ability was studied by either stripping off the protein and quantifying them, or by deductive reasoning through the use of "depleted" protein solutions. Fluorimetry and BCA assay both utilized the depleted solutions, but the high error associated with this protocol was prohibitive. SDS-PAGE with streptavidin was very difficult due to staining problems, however the IgG proteins were able to be quantified with some success. GIXA showed that the protein layer thickness is monolayer in nature, which agrees well with the AFM fluid tapping data on protein height, but in addition showed features on the order of ten protein agglomerations. CFM is by far the most promising technique for utilizing proteins as surface probes. Functionalized tips of -COOH, streptavidin and -CH3 are able to discern between surface treatments, but not forms. A general conclusion is that adhesion forces are greatest for -COOH, then streptavidin, and least for -CH3.

A novel Pfs38 protein complex on the surface of Plasmodium falciparum blood-stage merozoites.

PubMed

Paul, Gourab; Deshmukh, Arunaditya; Kaur, Inderjeet; Rathore, Sumit; Dabral, Surbhi; Panda, Ashutosh; Singh, Susheel Kumar; Mohmmed, Asif; Theisen, Michael; Malhotra, Pawan

2017-02-16

The Plasmodium genome encodes for a number of 6-Cys proteins that contain a module of six cysteine residues forming three intramolecular disulphide bonds. These proteins have been well characterized at transmission as well as hepatic stages of the parasite life cycle. In the present study, a large complex of 6-Cys proteins: Pfs41, Pfs38 and Pfs12 and three other merozoite surface proteins: Glutamate-rich protein (GLURP), SERA5 and MSP-1 were identified on the Plasmodium falciparum merozoite surface. Recombinant 6-cys proteins i.e. Pfs38, Pfs12, Pfs41 as well as PfMSP-1 65 were expressed and purified using Escherichia coli expression system and antibodies were raised against each of these proteins. These antibodies were used to immunoprecipitate the native proteins and their associated partners from parasite lysate. ELISA, Far western, surface plasmon resonance and glycerol density gradient fractionation were carried out to confirm the respective interactions. Furthermore, erythrocyte binding assay with 6-cys proteins were undertaken to find out their possible role in host-parasite infection and seropositivity was assessed using Indian and Liberian sera. Immunoprecipitation of parasite-derived polypeptides, followed by LC-MS/MS analysis, identified a large Pfs38 complex comprising of 6-cys proteins: Pfs41, Pfs38, Pfs12 and other merozoite surface proteins: GLURP, SERA5 and MSP-1. The existence of such a complex was further corroborated by several protein-protein interaction tools, co-localization and co-sedimentation analysis. Pfs38 protein of Pfs38 complex binds to host red blood cells (RBCs) directly via glycophorin A as a receptor. Seroprevalence analysis showed that of the six antigens, prevalence varied from 40 to 99%, being generally highest for MSP-1 65 and GLURP proteins. Together the data show the presence of a large Pfs38 protein-associated complex on the parasite surface which is involved in RBC binding. These results highlight the complex molecular interactions among the P. falciparum merozoite surface proteins and advocate the development of a multi-sub-unit malaria vaccine based on some of these protein complexes on merozoite surface.

Effect of mechanical denaturation on surface free energy of protein powders.

PubMed

Mohammad, Mohammad Amin; Grimsey, Ian M; Forbes, Robert T; Blagbrough, Ian S; Conway, Barbara R

2016-10-01

Globular proteins are important both as therapeutic agents and excipients. However, their fragile native conformations can be denatured during pharmaceutical processing, which leads to modification of the surface energy of their powders and hence their performance. Lyophilized powders of hen egg-white lysozyme and β-galactosidase from Aspergillus oryzae were used as models to study the effects of mechanical denaturation on the surface energies of basic and acidic protein powders, respectively. Their mechanical denaturation upon milling was confirmed by the absence of their thermal unfolding transition phases and by the changes in their secondary and tertiary structures. Inverse gas chromatography detected differences between both unprocessed protein powders and the changes induced by their mechanical denaturation. The surfaces of the acidic and basic protein powders were relatively basic, however the surface acidity of β-galactosidase was higher than that of lysozyme. Also, the surface of β-galactosidase powder had a higher dispersive energy compared to lysozyme. The mechanical denaturation decreased the dispersive energy and the basicity of the surfaces of both protein powders. The amino acid composition and molecular conformation of the proteins explained the surface energy data measured by inverse gas chromatography. The biological activity of mechanically denatured protein powders can either be reversible (lysozyme) or irreversible (β-galactosidase) upon hydration. Our surface data can be exploited to understand and predict the performance of protein powders within pharmaceutical dosage forms. Copyright © 2016 Elsevier B.V. All rights reserved.

Druggable protein interaction sites are more predisposed to surface pocket formation than the rest of the protein surface.

PubMed

Johnson, David K; Karanicolas, John

2013-01-01

Despite intense interest and considerable effort via high-throughput screening, there are few examples of small molecules that directly inhibit protein-protein interactions. This suggests that many protein interaction surfaces may not be intrinsically "druggable" by small molecules, and elevates in importance the few successful examples as model systems for improving our fundamental understanding of druggability. Here we describe an approach for exploring protein fluctuations enriched in conformations containing surface pockets suitable for small molecule binding. Starting from a set of seven unbound protein structures, we find that the presence of low-energy pocket-containing conformations is indeed a signature of druggable protein interaction sites and that analogous surface pockets are not formed elsewhere on the protein. We further find that ensembles of conformations generated with this biased approach structurally resemble known inhibitor-bound structures more closely than equivalent ensembles of unbiased conformations. Collectively these results suggest that "druggability" is a property encoded on a protein surface through its propensity to form pockets, and inspire a model in which the crude features of the predisposed pocket(s) restrict the range of complementary ligands; additional smaller conformational changes then respond to details of a particular ligand. We anticipate that the insights described here will prove useful in selecting protein targets for therapeutic intervention.

A synergistic approach to protein crystallization: Combination of a fixed-arm carrier with surface entropy reduction

PubMed Central

Moon, Andrea F; Mueller, Geoffrey A; Zhong, Xuejun; Pedersen, Lars C

2010-01-01

Protein crystallographers are often confronted with recalcitrant proteins not readily crystallizable, or which crystallize in problematic forms. A variety of techniques have been used to surmount such obstacles: crystallization using carrier proteins or antibody complexes, chemical modification, surface entropy reduction, proteolytic digestion, and additive screening. Here we present a synergistic approach for successful crystallization of proteins that do not form diffraction quality crystals using conventional methods. This approach combines favorable aspects of carrier-driven crystallization with surface entropy reduction. We have generated a series of maltose binding protein (MBP) fusion constructs containing different surface mutations designed to reduce surface entropy and encourage crystal lattice formation. The MBP advantageously increases protein expression and solubility, and provides a streamlined purification protocol. Using this technique, we have successfully solved the structures of three unrelated proteins that were previously unattainable. This crystallization technique represents a valuable rescue strategy for protein structure solution when conventional methods fail. PMID:20196072

Regulation of Macrophage Recognition through the Interplay of Nanoparticle Surface Functionality and Protein Corona.

PubMed

Saha, Krishnendu; Rahimi, Mehran; Yazdani, Mahdieh; Kim, Sung Tae; Moyano, Daniel F; Hou, Singyuk; Das, Ridhha; Mout, Rubul; Rezaee, Farhad; Mahmoudi, Morteza; Rotello, Vincent M

2016-04-26

Using a family of cationic gold nanoparticles (NPs) with similar size and charge, we demonstrate that proper surface engineering can control the nature and identity of protein corona in physiological serum conditions. The protein coronas were highly dependent on the hydrophobicity and arrangement of chemical motifs on NP surface. The NPs were uptaken in macrophages in a corona-dependent manner, predominantly through recognition of specific complement proteins in the NP corona. Taken together, this study shows that surface functionality can be used to tune the protein corona formed on NP surface, dictating the interaction of NPs with macrophages.

Applications of yeast surface display for protein engineering

PubMed Central

Cherf, Gerald M.; Cochran, Jennifer R.

2015-01-01

The method of displaying recombinant proteins on the surface of Saccharomyces cerevisiae via genetic fusion to an abundant cell wall protein, a technology known as yeast surface display, or simply, yeast display, has become a valuable protein engineering tool for a broad spectrum of biotechnology and biomedical applications. This review focuses on the use of yeast display for engineering protein affinity, stability, and enzymatic activity. Strategies and examples for each protein engineering goal are discussed. Additional applications of yeast display are also briefly presented, including protein epitope mapping, identification of protein-protein interactions, and uses of displayed proteins in industry and medicine. PMID:26060074

SFG analysis of surface bound proteins: a route towards structure determination.

PubMed

Weidner, Tobias; Castner, David G

2013-08-14

The surface of a material is rapidly covered with proteins once that material is placed in a biological environment. The structure and function of these bound proteins play a key role in the interactions and communications of the material with the biological environment. Thus, it is crucial to gain a molecular level understanding of surface bound protein structure. While X-ray diffraction and solution phase NMR methods are well established for determining the structure of proteins in the crystalline or solution phase, there is not a corresponding single technique that can provide the same level of structural detail about proteins at surfaces or interfaces. However, recent advances in sum frequency generation (SFG) vibrational spectroscopy have significantly increased our ability to obtain structural information about surface bound proteins and peptides. A multi-technique approach of combining SFG with (1) protein engineering methods to selectively introduce mutations and isotopic labels, (2) other experimental methods such as time-of-flight secondary ion mass spectrometry (ToF-SIMS) and near edge X-ray absorption fine structure (NEXAFS) to provide complementary information, and (3) molecular dynamic (MD) simulations to extend the molecular level experimental results is a particularly promising route for structural characterization of surface bound proteins and peptides. By using model peptides and small proteins with well-defined structures, methods have been developed to determine the orientation of both backbone and side chains to the surface.

SFG analysis of surface bound proteins: A route towards structure determination

PubMed Central

Weidner, Tobias; Castner, David G.

2013-01-01

The surface of a material is rapidly covered with proteins once that material is placed in a biological environment. The structure and function of these bound proteins play a key role in the interactions and communications of the material with the biological environment. Thus, it is crucial to gain a molecular level understanding of surface bound protein structure. While X-ray diffraction and solution phase NMR methods are well established for determining the structure of proteins in the crystalline or solution phase, there is not a corresponding single technique that can provide the same level of structural detail about proteins at surfaces or interfaces. However, recent advances in sum frequency generation (SFG) vibrational spectroscopy have significantly increased our ability to obtain structural information about surface bound proteins and peptides. A multi-technique approach of combining SFG with (1) protein engineering methods to selectively introduce mutations and isotopic labels, (2) other experimental methods such as time-of-flight secondary ion mass spectrometry (ToF-SIMS) and near edge x-ray absorption fine structure (NEXAFS) to provide complementary information, and (3) molecular dynamic (MD) simulations to extend the molecular level experimental results is a particularly promising route for structural characterization of surface bound proteins and peptides. By using model peptides and small proteins with well-defined structures, methods have been developed to determine the orientation of both backbone and side chains to the surface. PMID:23727992

Improving biocompatibility by surface modification techniques on implantable bioelectronics.

PubMed

Lin, Peter; Lin, Chii-Wann; Mansour, Raafat; Gu, Frank

2013-09-15

For implantable bioelectronic devices, the interface between the device and the biological environment requires significant attention as it dictates the device performance in vivo. Non-specific protein adsorption onto the device surface is the initial stage of many degradation mechanisms that will ultimately compromise the functionality of the device. In order to preserve the functionality of any implanted bioelectronics overtime, protein adsorption must be controlled. This review paper outlines two major approaches to minimize protein adsorption onto the surface of implantable electronics. The first approach is surface coating, which minimizes close proximity interactions between proteins and device surfaces by immobilizing electrically neutral hydrophilic polymers as surface coating. These coatings reduce protein fouling by steric repulsion and formation of a hydration layer which acts as both a physical and energetic barrier that minimize protein adsorption onto the device. Relevant performances of various conventional hydrophilic coatings are discussed. The second approach is surface patterning using arrays of hydrophobic nanostructures through photolithography techniques. By establishing a large slip length via super hydrophobic surfaces, the amount of proteins adsorbed to the surface of the device can be reduced. The last section discusses emerging surface coating techniques utilizing zwitterionic polymers where ultralow-biofouling surfaces have been demonstrated. These surface modification techniques may significantly improve the long-term functionality of implantable bioelectronics, thus allowing researchers to overcome challenges to diagnose and treat chronic neurological and cardiovascular diseases. Copyright © 2013 Elsevier B.V. All rights reserved.

Water organization between oppositely charged surfaces: Implications for protein sliding along DNA a)

NASA Astrophysics Data System (ADS)

Marcovitz, Amir; Naftaly, Aviv; Levy, Yaakov

2015-02-01

Water molecules are abundant in protein-DNA interfaces, especially in their nonspecific complexes. In this study, we investigated the organization and energetics of the interfacial water by simplifying the geometries of the proteins and the DNA to represent them as two equally and oppositely charged planar surfaces immersed in water. We found that the potential of mean force for bringing the two parallel surfaces into close proximity comprises energetic barriers whose properties strongly depend on the charge density of the surfaces. We demonstrated how the organization of the water molecules into discretized layers and the corresponding energetic barriers to dehydration can be modulated by the charge density on the surfaces, salt, and the structure of the surfaces. The 1-2 layers of ordered water are tightly bound to the charged surfaces representing the nonspecific protein-DNA complex. This suggests that water might mediate one-dimensional diffusion of proteins along DNA (sliding) by screening attractive electrostatic interactions between the positively charged molecular surface on the protein and the negatively charged DNA backbone and, in doing so, reduce intermolecular friction in a manner that smoothens the energetic landscape for sliding, and facilitates the 1D diffusion of the protein.

A role for surface hydrophobicity in protein-protein recognition.

PubMed Central

Young, L.; Jernigan, R. L.; Covell, D. G.

1994-01-01

The role of hydrophobicity as a determinant of protein-protein interactions is examined. Surfaces of apo-protein targets comprising 9 classes of enzymes, 7 antibody fragments, hirudin, growth hormone, and retinol-binding protein, and their associated ligands with available X-ray structures for their complexed forms, are scanned to determine clusters of surface-accessible amino acids. Clusters of surface residues are ranked on the basis of the hydrophobicity of their constituent amino acids. The results indicate that the location of the co-crystallized ligand is commonly found to correspond with one of the strongest hydrophobic clusters on the surface of the target molecule. In 25 of 38 cases, the correspondence is exact, with the position of the most hydrophobic cluster coinciding with more than one-third of the surface buried by the bound ligand. The remaining 13 cases demonstrate this correspondence within the top 6 hydrophobic clusters. These results suggest that surface hydrophobicity can be used to identify regions of a protein's surface most likely to interact with a binding ligand. This fast and simple procedure may be useful for identifying small sets of well-defined loci for possible ligand attachment. PMID:8061602

Membrane Fusion Promoted by Increasing Surface Densities of the Paramyxovirus F and HN Proteins: Comparison of Fusion Reactions Mediated by Simian Virus 5 F, Human Parainfluenza Virus Type 3 F, and Influenza Virus HA

PubMed Central

Dutch, Rebecca Ellis; Joshi, Sangeeta Bagai; Lamb, Robert A.

1998-01-01

The membrane fusion reaction promoted by the paramyxovirus simian virus 5 (SV5) and human parainfluenza virus type 3 (HPIV-3) fusion (F) proteins and hemagglutinin-neuraminidase (HN) proteins was characterized when the surface densities of F and HN were varied. Using a quantitative content mixing assay, it was found that the extent of SV5 F-mediated fusion was dependent on the surface density of the SV5 F protein but independent of the density of SV5 HN protein, indicating that HN serves only a binding function in the reaction. However, the extent of HPIV-3 F protein promoted fusion reaction was found to be dependent on surface density of HPIV-3 HN protein, suggesting that the HPIV-3 HN protein is a direct participant in the fusion reaction. Analysis of the kinetics of lipid mixing demonstrated that both initial rates and final extents of fusion increased with rising SV5 F protein surface densities, suggesting that multiple fusion pores can be active during SV5 F protein-promoted membrane fusion. Initial rates and extent of lipid mixing were also found to increase with increasing influenza virus hemagglutinin protein surface density, suggesting parallels between the mechanism of fusion promoted by these two viral fusion proteins. PMID:9733810

Protein-surfactant interactions at hydrophobic interfaces studied with total internal reflection fluorescence correlation spectroscopy (TIR-FCS).

PubMed

Sonesson, Andreas W; Blom, Hans; Hassler, Kai; Elofsson, Ulla M; Callisen, Thomas H; Widengren, Jerker; Brismar, Hjalmar

2008-01-15

The aim of this work was to study the dynamics of proteins near solid surfaces in the presence or absence of competing surfactants by means of total internal reflection fluorescence correlation spectroscopy (TIR-FCS). Two different proteins were studied, bovine serum albumin (BSA) and Thermomyces lanuginosus lipase (TLL). A nonionic/anionic (C12E6/LAS) surfactant composition was used to mimic a detergent formulation and the surfaces used were C18 terminated glass. It was found that with increasing surfactant concentrations the term in the autocorrelation function (ACF) representing surface binding decreased. This suggested that the proteins were competed off the hydrophobic surface by the surfactant. When fitting the measured ACF to a model for surface kinetics, it was seen that with raised C12E6/LAS concentration, the surface interaction rate increased for both proteins. Under these experimental conditions this meant that the time the protein was bound to the surface decreased. At 10 microM C12E6/LAS the surface interaction was not visible for BSA, whereas it was still distinguishable in the ACF for TLL. This indicated that TLL had a higher affinity than BSA for the C18 surface. The study showed that TIR-FCS provides a useful tool to quantify the surfactant effect on proteins adsorption.

Pooled protein immunization for identification of cell surface antigens in Streptococcus sanguinis.

PubMed

Ge, Xiuchun; Kitten, Todd; Munro, Cindy L; Conrad, Daniel H; Xu, Ping

2010-07-26

Available bacterial genomes provide opportunities for screening vaccines by reverse vaccinology. Efficient identification of surface antigens is required to reduce time and animal cost in this technology. We developed an approach to identify surface antigens rapidly in Streptococcus sanguinis, a common infective endocarditis causative species. We applied bioinformatics for antigen prediction and pooled antigens for immunization. Forty-seven surface-exposed proteins including 28 lipoproteins and 19 cell wall-anchored proteins were chosen based on computer algorithms and comparative genomic analyses. Eight proteins among these candidates and 2 other proteins were pooled together to immunize rabbits. The antiserum reacted strongly with each protein and with S. sanguinis whole cells. Affinity chromatography was used to purify the antibodies to 9 of the antigen pool components. Competitive ELISA and FACS results indicated that these 9 proteins were exposed on S. sanguinis cell surfaces. The purified antibodies had demonstrable opsonic activity. The results indicate that immunization with pooled proteins, in combination with affinity purification, and comprehensive immunological assays may facilitate cell surface antigen identification to combat infectious diseases.

Pooled Protein Immunization for Identification of Cell Surface Antigens in Streptococcus sanguinis

PubMed Central

Ge, Xiuchun; Kitten, Todd; Munro, Cindy L.; Conrad, Daniel H.; Xu, Ping

2010-01-01

Background Available bacterial genomes provide opportunities for screening vaccines by reverse vaccinology. Efficient identification of surface antigens is required to reduce time and animal cost in this technology. We developed an approach to identify surface antigens rapidly in Streptococcus sanguinis, a common infective endocarditis causative species. Methods and Findings We applied bioinformatics for antigen prediction and pooled antigens for immunization. Forty-seven surface-exposed proteins including 28 lipoproteins and 19 cell wall-anchored proteins were chosen based on computer algorithms and comparative genomic analyses. Eight proteins among these candidates and 2 other proteins were pooled together to immunize rabbits. The antiserum reacted strongly with each protein and with S. sanguinis whole cells. Affinity chromatography was used to purify the antibodies to 9 of the antigen pool components. Competitive ELISA and FACS results indicated that these 9 proteins were exposed on S. sanguinis cell surfaces. The purified antibodies had demonstrable opsonic activity. Conclusions The results indicate that immunization with pooled proteins, in combination with affinity purification, and comprehensive immunological assays may facilitate cell surface antigen identification to combat infectious diseases. PMID:20668678

Candida albicans Shaving to Profile Human Serum Proteins on Hyphal Surface

PubMed Central

Marín, Elvira; Parra-Giraldo, Claudia M.; Hernández-Haro, Carolina; Hernáez, María L.; Nombela, César; Monteoliva, Lucía; Gil, Concha

2015-01-01

Candida albicans is a human opportunistic fungus and it is responsible for a wide variety of infections, either superficial or systemic. C. albicans is a polymorphic fungus and its ability to switch between yeast and hyphae is essential for its virulence. Once C. albicans obtains access to the human body, the host serum constitutes a complex environment of interaction with C. albicans cell surface in bloodstream. To draw a comprehensive picture of this relevant step in host-pathogen interaction during invasive candidiasis, we have optimized a gel-free shaving proteomic strategy to identify both, human serum proteins coating C. albicans cells and fungi surface proteins simultaneously. This approach was carried out with normal serum (NS) and heat inactivated serum (HIS). We identified 214 human and 372 C. albicans unique proteins. Proteins identified in C. albicans included 147 which were described as located at the cell surface and 52 that were described as immunogenic. Interestingly, among these C. albicans proteins, we identified 23 GPI-anchored proteins, Gpd2 and Pra1, which are involved in complement system evasion and 7 other proteins that are able to attach plasminogen to C. albicans surface (Adh1, Eno1, Fba1, Pgk1, Tdh3, Tef1, and Tsa1). Furthermore, 12 proteins identified at the C. albicans hyphae surface induced with 10% human serum were not detected in other hypha-induced conditions. The most abundant human proteins identified are involved in complement and coagulation pathways. Remarkably, with this strategy, all main proteins belonging to complement cascades were identified on the C. albicans surface. Moreover, we identified immunoglobulins, cytoskeletal proteins, metabolic proteins such as apolipoproteins and others. Additionally, we identified more inhibitors of complement and coagulation pathways, some of them serpin proteins (serine protease inhibitors), in HIS vs. NS. On the other hand, we detected a higher amount of C3 at the C. albicans surface in NS than in HIS, as validated by immunofluorescence. PMID:26696967

Competitive Protein Adsorption on Polysaccharide and Hyaluronate Modified Surfaces

PubMed Central

Ombelli, Michela; Costello, Lauren; Postle, Corinne; Anantharaman, Vinod; Meng, Qing Cheng; Composto, Russell J.; Eckmann, David M.

2011-01-01

We measured adsorption of bovine serum albumin (BSA) and fibrinogen (Fg) onto six distinct bare and dextran- and hyaluronate-modified silicon surfaces created using two dextran grafting densities and three hyaluronic acid (HA) sodium salts derived from human umbilical cord, rooster comb and streptococcus zooepidemicus. Film thickness and surface morphology depended on HA molecular weight and concentration. BSA coverage was enhanced on surfaces upon competitive adsorption of BSA:Fg mixtures. Dextranization differentially reduced protein adsorption onto surfaces based on oxidation state. Hyaluronization was demonstrated to provide the greatest resistance to protein coverage, equivalent to that of the most resistant dextranized surface. Resistance to protein adsorption was independent of the type of hyaluronic acid utilized. With changing bulk protein concentration from 20 to 40 µg ml−1 for each species, Fg coverage on silicon increased by 4×, whereas both BSA and Fg adsorption on dextran and HA were far less dependent of protein bulk concentration. PMID:21623481

Atomic force microscopy – looking at mechanosensors on the cell surface

PubMed Central

Heinisch, Jürgen J.; Lipke, Peter N.; Beaussart, Audrey; El Kirat Chatel, Sofiane; Dupres, Vincent; Alsteens, David; Dufrêne, Yves F.

2012-01-01

Summary Living cells use cell surface proteins, such as mechanosensors, to constantly sense and respond to their environment. However, the way in which these proteins respond to mechanical stimuli and assemble into large complexes remains poorly understood at the molecular level. In the past years, atomic force microscopy (AFM) has revolutionized the way in which biologists analyze cell surface proteins to molecular resolution. In this Commentary, we discuss how the powerful set of advanced AFM techniques (e.g. live-cell imaging and single-molecule manipulation) can be integrated with the modern tools of molecular genetics (i.e. protein design) to study the localization and molecular elasticity of individual mechanosensors on the surface of living cells. Although we emphasize recent studies on cell surface proteins from yeasts, the techniques described are applicable to surface proteins from virtually all organisms, from bacteria to human cells. PMID:23077172

Surface Proteins of Gram-Positive Pathogens: Using Crystallography to Uncover Novel Features in Drug and Vaccine Candidates

NASA Astrophysics Data System (ADS)

Baker, Edward N.; Proft, Thomas; Kang, Haejoo

Proteins displayed on the cell surfaces of pathogenic organisms are the front-line troops of bacterial attack, playing critical roles in colonization, infection and virulence. Although such proteins can often be recognized from genome sequence data, through characteristic sequence motifs, their functions are often unknown. One such group of surface proteins is attached to the cell surface of Gram-positive pathogens through the action of sortase enzymes. Some of these proteins are now known to form pili: long filamentous structures that mediate attachment to human cells. Crystallographic analyses of these and other cell surface proteins have uncovered novel features in their structure, assembly and stability, including the presence of inter- and intramolecular isopeptide crosslinks. This improved understanding of structures on the bacterial cell surface offers opportunities for the development of some new drug targets and for novel approaches to vaccine design.

Modulating the activity of protein conjugated to gold nanoparticles by site-directed orientation and surface density of bound protein.

PubMed

Liu, Feng; Wang, Lei; Wang, Hongwei; Yuan, Lin; Li, Jingwen; Brash, John Law; Chen, Hong

2015-02-18

The key property of protein-nanoparticle conjugates is the bioactivity of the protein. The ability to accurately modulate the activity of protein on the nanoparticles at the interfaces is important in many applications. In the work reported here, modulation of the activity of protein-gold nanoparticle (AuNP) conjugates by specifically orienting the protein and by varying the surface density of the protein was investigated. Different orientations were achieved by introducing cysteine (Cys) residues at specific sites for binding to gold. We chose Escherichia coli inorganic pyrophosphatase (PPase) as a model protein and used site-directed mutagenesis to generate two mutant types (MTs) with a single Cys residue on the surface: MT1 with Cys near the active center and MT2 with Cys far from the active center. The relative activities of AuNP conjugates with wild type (WT), MT1, and MT2 were found to be 44.8%, 68.8%, and 91.2% of native PPase in aqueous solution. Site-directed orientation with the binding site far from the active center thus allowed almost complete preservation of the protein activity. The relative activity of WT and MT2 conjugates did not change with the surface density of the protein, while that of MT1 increased significantly with increasing surface density. These results demonstrate that site-directed orientation and surface density can both modulate the activity of proteins conjugated to AuNP and that orientation has a greater effect than density. Furthermore, increasing the surface density of the specifically oriented protein MT2, while having no significant effect on the specific activity of the protein, still allowed increased protein loading on the AuNP and thus increased the total protein activity. This is of great importance in the study on the interface of protein and nanoparticle and the applications for enzyme immobilization, drug delivery, and biocatalysis.

Platelet Adhesion and Activation on Chiral Surfaces: The Influence of Protein Adsorption.

PubMed

Fan, Yonghong; Luo, Rifang; Han, Honghong; Weng, Yajun; Wang, Hong; Li, Jing'an; Yang, Ping; Wang, Yunbing; Huang, Nan

2017-10-03

Adsorbed proteins and their conformational change on blood-contacting biomaterials will determine their final hemocompatibility. It has frequently been reported that surface chirality of biomaterials may highly influence their protein adsorption behavior. Here, lysine and tartaric acid with different chirality were immobilized onto TiO 2 films respectively, and the influence of surface chirality on protein adsorption, platelet adhesion, and activation was also investigated. It showed that the l- and d-molecule grafted samples had almost the same grafting density, surface topography, chemical components, and hydrophilicity in this study. However, biological behaviors such as protein adsorption, platelet adhesion, and activation were quite different. The d-lysine grafted surface had a greater ability to inhibit both bovine serum albumin and fibrinogen adsorption, along with less degeneration of fibrinogen compared to the l-lysine anchored surface. However, the d-tartaric acid grafted surface adsorbed more protein but with less denatured fibrinogen compared to the l-tartaric acid grafted one. Further studies showed that the secondary structural change of the adsorbed albumin and fibrinogen on all surfaces with deduction of the α-helix content and increase of disordered structure, while the changing degree was apparently varied. As a result, the d-lysine immobilized surface absorbed less platelets and red blood cells and achieved slightly increased platelet activation. For tartaric acid anchored surfaces, a larger number of platelets adhered to the D-surface but were less activated compared to the L-surface. In conclusion, the surface chirality significantly influenced the adsorption and conformational change of blood plasma protein, which in turn influenced both platelet adhesion and activation.

Selectivity by Small-Molecule Inhibitors of Protein Interactions Can Be Driven by Protein Surface Fluctuations

PubMed Central

Johnson, David K.; Karanicolas, John

2015-01-01

Small-molecules that inhibit interactions between specific pairs of proteins have long represented a promising avenue for therapeutic intervention in a variety of settings. Structural studies have shown that in many cases, the inhibitor-bound protein adopts a conformation that is distinct from its unbound and its protein-bound conformations. This plasticity of the protein surface presents a major challenge in predicting which members of a protein family will be inhibited by a given ligand. Here, we use biased simulations of Bcl-2-family proteins to generate ensembles of low-energy conformations that contain surface pockets suitable for small molecule binding. We find that the resulting conformational ensembles include surface pockets that mimic those observed in inhibitor-bound crystal structures. Next, we find that the ensembles generated using different members of this protein family are overlapping but distinct, and that the activity of a given compound against a particular family member (ligand selectivity) can be predicted from whether the corresponding ensemble samples a complementary surface pocket. Finally, we find that each ensemble includes certain surface pockets that are not shared by any other family member: while no inhibitors have yet been identified to take advantage of these pockets, we expect that chemical scaffolds complementing these “distinct” pockets will prove highly selective for their targets. The opportunity to achieve target selectivity within a protein family by exploiting differences in surface fluctuations represents a new paradigm that may facilitate design of family-selective small-molecule inhibitors of protein-protein interactions. PMID:25706586

NMR identification of the binding surfaces involved in the Salmonella and Shigella Type III secretion tip-translocon protein-protein interactions.

PubMed

McShan, Andrew C; Kaur, Kawaljit; Chatterjee, Srirupa; Knight, Kevin M; De Guzman, Roberto N

2016-08-01

The type III secretion system (T3SS) is essential for the pathogenesis of many bacteria including Salmonella and Shigella, which together are responsible for millions of deaths worldwide each year. The structural component of the T3SS consists of the needle apparatus, which is assembled in part by the protein-protein interaction between the tip and the translocon. The atomic detail of the interaction between the tip and the translocon proteins is currently unknown. Here, we used NMR methods to identify that the N-terminal domain of the Salmonella SipB translocon protein interacts with the SipD tip protein at a surface at the distal region of the tip formed by the mixed α/β domain and a portion of its coiled-coil domain. Likewise, the Shigella IpaB translocon protein and the IpaD tip protein interact with each other using similar surfaces identified for the Salmonella homologs. Furthermore, removal of the extreme N-terminal residues of the translocon protein, previously thought to be important for the interaction, had little change on the binding surface. Finally, mutations at the binding surface of SipD reduced invasion of Salmonella into human intestinal epithelial cells. Together, these results reveal the binding surfaces involved in the tip-translocon protein-protein interaction and advance our understanding of the assembly of the T3SS needle apparatus. Proteins 2016; 84:1097-1107. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Studying protein structural changes based on surface plasmon resonance and surface-enhanced Raman scattering

NASA Astrophysics Data System (ADS)

Hu, Wen-Pin; Chen, Shean-Jen; Yih, Jenq-Nan; Lin, G.-Y.; Chang, Guan L.

2004-06-01

The ability to recognize the conformational changes and structural variations of a protein when immobilized in a solid surface is of great importance in a variety of applications. Surface plasmon resonance (SPR) sensing is an appropriate technique for investigating interfacial phenomena, and enables the conformational changes of proteins to be monitored through the variation in the SPR angle shift. Meanwhile, the surface-enhanced Raman scattering (SERS) system can also assist in clarifying the changes in protein structure. The present study utilizes a 1 mM CrO3 phosphate buffer solution (PBS) to induce conformational changes of human serum albumin (HSA). Monitoring the corresponding SPR angle shifts and the SPR reflectivity spectrum enables the relationships between the conformational changes of the surface-immobilized protein and the thickness and dielectric constants of the protein layer to be estimated. The experimental SPR results indicate that the Cr6+ ions cause significant conformational change of the protein. It is established that the ions are not merely absorbed into the protein as a result of electrostatic forces, but that complex protein refolding events also take place. Furthermore, the data acquired from the SERS system yield valuable information regarding the changes which take place in the protein structure.

Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags

PubMed Central

2017-01-01

We report oriented immobilization of proteins using the standard hexahistidine (His6)-Ni2+:NTA (nitrilotriacetic acid) methodology, which we systematically tuned to give control of surface coverage. Fluorescence microscopy and surface plasmon resonance measurements of self-assembled monolayers (SAMs) of red fluorescent proteins (TagRFP) showed that binding strength increased by 1 order of magnitude for each additional His6-tag on the TagRFP proteins. All TagRFP variants with His6-tags located on only one side of the barrel-shaped protein yielded a 1.5 times higher surface coverage compared to variants with His6-tags on opposite sides of the so-called β-barrel. Time-resolved fluorescence anisotropy measurements supported by polarized infrared spectroscopy verified that the orientation (and thus coverage and functionality) of proteins on surfaces can be controlled by strategic placement of a His6-tag on the protein. Molecular dynamics simulations show how the differently tagged proteins reside at the surface in “end-on” and “side-on” orientations with each His6-tag contributing to binding. Also, not every dihistidine subunit in a given His6-tag forms a full coordination bond with the Ni2+:NTA SAMs, which varied with the position of the His6-tag on the protein. At equal valency but different tag positions on the protein, differences in binding were caused by probing for Ni2+:NTA moieties and by additional electrostatic interactions between different fractions of the β-barrel structure and charged NTA moieties. Potential of mean force calculations indicate there is no specific single-protein interaction mode that provides a clear preferential surface orientation, suggesting that the experimentally measured preference for the end-on orientation is a supra-protein, not a single-protein, effect. PMID:28850777

Jet blown PTFE for control of biocompatibility

NASA Astrophysics Data System (ADS)

Leibner, Evan Scott

The development of fully hemocompatible cardiovascular biomaterials will have a major impact on the practice of modern medicine. Current artificial surfaces, unlike native vascular surfaces, are not able to control clot and thrombus formation. Protein interactions are an important component in hemocompatibility and can result in decreased patency due to thrombus formation or surface passivation which can improve endothelization. It is believed that controlling these properties, specifically the nanometer sizes of the fibers on the material's surface, will allow for better control of biological responses. The biocompatibility of Teflon, a widely used polymer for vascular grafts, would be improved with nanostructured control of surface features. Due to the difficultly in processing polytetrafluoroethylene (PTFE), it has not been possible to create nanofibrous PTFE surfaces. The novel technique of Jet Blowing allows for the formation of nanostructured PTFE (nPTFE). A systematic investigation into controlling polymer properties by varying the processing conditions of temperature, pressure, and gas used in the Jet Blowing allows for an increased understanding of the effects of plasticization on the material's properties. This fundamental understanding of the material science behind the Jet Blowing process has enabled control of the micro and nanoscale structure of nPTFE. While protein adsorption, a key component of biocompatibility, has been widely studied, it is not fully understood. Major problems in the field of biomaterials include a lack of standard protocols to measure biocompatibility, and inconstant literature on protein adsorption. A reproducible protocol for measuring protein adsorption onto superhydrophobic surfaces (ePTFE and nPTFE) has been developed. Both degassing of PBS buffer solutions and evacuation of the air around the expanded PTFE (ePTFE) prior to contact with protein solutions are essential. Protein adsorption experiments show a four-fold difference in the measure of proteins adsorbed using radiometry (I-125 labeled human serum albumin (HSA)) and electrophoresis (unlabeled HSA). This provides evidence that the standard method of radiolabeled protein for measuring adsorption does not fully account for changes to the HSA molecules due to labeling. The differences between measured protein values can be attributed to the radiolabel affecting the HSA hydrophobicity resulting in a change in the protein's interactions with the hydrophobic surface. Additionally, our work has provided repeatable results showing that the amount of protein adsorbed onto the polymer surface, after washing, accounted for only 65% of the amount of protein that was removed from solution based on depletion analysis. This implies that measurement of the amount of strongly bound protein on the material significantly underestimates the actual amount of protein adsorbing into the surface region of the material interface. HSA adsorption isotherms demonstrate an increase in protein adsorption capacity on the nPTFE surface compared to adsorption on the same surface area of ePTFE. Preliminary cell work shows that the nPTFE surfaces had a larger number of cells growing on the surface of the material when compared to ePTFE surfaces. The research also shows that while most endothelial cells were not viable on the ePTFE surface after 96 hours, they remained alive on the nPTFE surface during that same time period. Surface functionalization using ammonia plasma has been performed. X-ray photoelectron spectroscopy (XPS) analysis revealed the presence of amine groups on the nPTFE surface. The amine groups can be used to couple polypeptides onto the PTFE surface in the future. The selection of different peptides will allow for selective control of cell adhesion. This research shows that nPTFE has potential for improved biocompatibility over standard ePTFE, based on increased protein adsorption capacity, increased viability of endothelial cells, and the ability to plasma modify the PTFE surface.

Shape Complementarity of Protein-Protein Complexes at Multiple Resolutions

PubMed Central

Zhang, Qing; Sanner, Michel; Olson, Arthur J.

2010-01-01

Biological complexes typically exhibit intermolecular interfaces of high shape complementarity. Many computational docking approaches use this surface complementarity as a guide in the search for predicting the structures of protein-protein complexes. Proteins often undergo conformational changes in order to create a highly complementary interface when associating. These conformational changes are a major cause of failure for automated docking procedures when predicting binding modes between proteins using their unbound conformations. Low resolution surfaces in which high frequency geometric details are omitted have been used to address this problem. These smoothed, or blurred, surfaces are expected to minimize the differences between free and bound structures, especially those that are due to side chain conformations or small backbone deviations. In spite of the fact that this approach has been used in many docking protocols, there has yet to be a systematic study of the effects of such surface smoothing on the shape complementarity of the resulting interfaces. Here we investigate this question by computing shape complementarity of a set of 66 protein-protein complexes represented by multi-resolution blurred surfaces. Complexed and unbound structures are available for these protein-protein complexes. They are a subset of complexes from a non-redundant docking benchmark selected for rigidity (i.e. the proteins undergo limited conformational changes between their bound and unbound states). In this work we construct the surfaces by isocontouring a density map obtained by accumulating the densities of Gaussian functions placed at all atom centers of the molecule. The smoothness or resolution is specified by a Gaussian fall-off coefficient, termed “blobbyness”. Shape complementarity is quantified using a histogram of the shortest distances between two proteins' surface mesh vertices for both the crystallographic complexes and the complexes built using the protein structures in their unbound conformation. The histograms calculated for the bound complex structures demonstrate that medium resolution smoothing (blobbyness=−0.9) can reproduce about 88% of the shape complementarity of atomic resolution surfaces. Complexes formed from the free component structures show a partial loss of shape complementarity (more overlaps and gaps) with the atomic resolution surfaces. For surfaces smoothed to low resolution (blobbyness=−0.3), we find more consistency of shape complementarity between the complexed and free cases. To further reduce bad contacts without significantly impacting the good contacts we introduce another blurred surface, in which the Gaussian densities of flexible atoms are reduced. From these results we discuss the use of shape complementarity in protein-protein docking. PMID:18837463

Surface derivatization strategy for combinatorial analysis of cell response to mixtures of protein domains.

PubMed

Chiang, Chunyi; Karuri, Stella W; Kshatriya, Pradnya P; Schwartz, Jeffrey; Schwarzbauer, Jean E; Karuri, Nancy W

2012-01-10

We report a robust strategy for conjugating mixtures of two or more protein domains to nonfouling polyurethane surfaces. In our strategy, the carbamate groups of polyurethane are reacted with zirconium alkoxide from the vapor phase to give a surface-bound oxide that serves as a chemical layer that can be used to bond organics to the polymer substrate. A hydroxyalkylphosphonate monolayer was synthesized on this layer, which was then used to covalently bind primary amine groups in protein domains using chloroformate-derived cross-linking. The effectiveness of this synthesis strategy was gauged by using an ELISA to measure competitive, covalent bonding of cell-binding (III(9-10)) and fibronectin-binding (III(1-2)) domains of the cell adhesion protein fibronectin. Cell adhesion, spreading, and fibronectin matrix assembly were examined on surfaces conjugated with single domains, a 1:1 surface mixture of III(1-2) and III(9-10), and a recombinant protein "duplex" containing both domains in one fusion protein. The mixture performed as well as or better than the other surfaces in these assays. Our surface activation strategy is amenable to a wide range of polymer substrates and free amino group-containing protein fragments. As such, this technique may be used to create biologically specific materials through the immobilization of specific protein groups or mixtures thereof on a substrate surface.

Effect of tissue scaffold topography on protein structure monitored by fluorescence spectroscopy.

PubMed

Portugal, Carla A M; Truckenmüller, Roman; Stamatialis, Dimitrios; Crespo, João G

2014-11-10

The impact of surface topography on the structure of proteins upon adhesion was assessed through non-invasive fluorescence monitoring. This study aimed at obtaining a better understanding about the role of protein structural status on cell-scaffold interactions. The changes induced upon adsorption of two model proteins with different geometries, trypsin (globular conformation) and fibrinogen (rod-shaped conformation) on poly-l-lactic acid (PLLA) scaffolds with different surface topographies, flat, fibrous and surfaces with aligned nanogrooves, were assessed by fluorescence spectroscopy monitoring, using tryptophan as structural probe. Hence, the maximum emission blue shift and the increase of fluorescence anisotropy observed after adsorption of globular and rod-like shaped proteins on surfaces with parallel nanogrooves were ascribed to more intense protein-surface interactions. Furthermore, the decrease of fluorescence anisotropy observed upon adsorption of proteins to scaffolds with fibrous morphology was more significant for rod-shaped proteins. This effect was associated to the ability of these proteins to adjust to curved surfaces. The additional unfolding of proteins induced upon adsorption on scaffolds with a fibrous morphology may be the reason for better cell attachment there, promoting an easier access of cell receptors to initially hidden protein regions (e.g. RGDS sequence), which are known to have a determinant role in cell attaching processes. Copyright © 2014 Elsevier B.V. All rights reserved.

Druggable Protein Interaction Sites Are More Predisposed to Surface Pocket Formation than the Rest of the Protein Surface

PubMed Central

Johnson, David K.; Karanicolas, John

2013-01-01

Despite intense interest and considerable effort via high-throughput screening, there are few examples of small molecules that directly inhibit protein-protein interactions. This suggests that many protein interaction surfaces may not be intrinsically “druggable” by small molecules, and elevates in importance the few successful examples as model systems for improving our fundamental understanding of druggability. Here we describe an approach for exploring protein fluctuations enriched in conformations containing surface pockets suitable for small molecule binding. Starting from a set of seven unbound protein structures, we find that the presence of low-energy pocket-containing conformations is indeed a signature of druggable protein interaction sites and that analogous surface pockets are not formed elsewhere on the protein. We further find that ensembles of conformations generated with this biased approach structurally resemble known inhibitor-bound structures more closely than equivalent ensembles of unbiased conformations. Collectively these results suggest that “druggability” is a property encoded on a protein surface through its propensity to form pockets, and inspire a model in which the crude features of the predisposed pocket(s) restrict the range of complementary ligands; additional smaller conformational changes then respond to details of a particular ligand. We anticipate that the insights described here will prove useful in selecting protein targets for therapeutic intervention. PMID:23505360

Protein consensus-based surface engineering (ProCoS): a computer-assisted method for directed protein evolution.

PubMed

Shivange, Amol V; Hoeffken, Hans Wolfgang; Haefner, Stefan; Schwaneberg, Ulrich

2016-12-01

Protein consensus-based surface engineering (ProCoS) is a simple and efficient method for directed protein evolution combining computational analysis and molecular biology tools to engineer protein surfaces. ProCoS is based on the hypothesis that conserved residues originated from a common ancestor and that these residues are crucial for the function of a protein, whereas highly variable regions (situated on the surface of a protein) can be targeted for surface engineering to maximize performance. ProCoS comprises four main steps: ( i ) identification of conserved and highly variable regions; ( ii ) protein sequence design by substituting residues in the highly variable regions, and gene synthesis; ( iii ) in vitro DNA recombination of synthetic genes; and ( iv ) screening for active variants. ProCoS is a simple method for surface mutagenesis in which multiple sequence alignment is used for selection of surface residues based on a structural model. To demonstrate the technique's utility for directed evolution, the surface of a phytase enzyme from Yersinia mollaretii (Ymphytase) was subjected to ProCoS. Screening just 1050 clones from ProCoS engineering-guided mutant libraries yielded an enzyme with 34 amino acid substitutions. The surface-engineered Ymphytase exhibited 3.8-fold higher pH stability (at pH 2.8 for 3 h) and retained 40% of the enzyme's specific activity (400 U/mg) compared with the wild-type Ymphytase. The pH stability might be attributed to a significantly increased (20 percentage points; from 9% to 29%) number of negatively charged amino acids on the surface of the engineered phytase.

Modeling the surface of Campylobacter fetus: protein surface layer stability and resistance to cationic antimicrobial peptides.

PubMed

Roberts, James M D; Graham, Lori L; Quinn, Bonnie; Pink, David A

2013-03-01

Campylobacter fetus is a Gram negative bacterium recognized for its virulence in animals and humans. This bacterium possesses a paracrystalline array of high molecular weight proteins known as surface-layer proteins covering its cell surface. A mathematical model has been made of the outer membrane of this bacterium, both with its surface-layer proteins (S+) and without (S-). Monte Carlo computer simulation was used to understand the stability of the surface-layer protein structure as a function of ionic concentration. The interactions of an electrically-charged antimicrobial agent, the cationic antimicrobial peptide protamine, with surface-layer proteins and with the lipopolysaccharides of the outer membrane were modeled and analyzed. We found that (1) divalent ions stabilize the surface-layer protein array by reducing the fluctuations perpendicular and parallel to the membrane plane thereby promoting adhesion to the LPS region. This was achieved via (2) divalent ions bridging the negatively-charged LPS Core. The effect of this bridging is to bring individual Core regions closer together so that the O-antigens can (3) increase their attractive van der Waals interactions and "collapse" to form a surface with reduced perpendicular fluctuations. These findings provide support for the proposal of Yang et al. [1]. (4) No evidence for a significant increase in Ca(2+) concentration in the region of the surface-layer protein subunits was observed in S+ simulations compared to S- simulations. (5) We predicted the trends of protamine MIC tests performed on C. fetus and these were in good agreement with our experimental results. Copyright © 2012 Elsevier B.V. All rights reserved.

Targeted Proteomics and Absolute Protein Quantification for the Construction of a Stoichiometric Host-Pathogen Surface Density Model.

PubMed

Sjöholm, Kristoffer; Kilsgård, Ola; Teleman, Johan; Happonen, Lotta; Malmström, Lars; Malmström, Johan

2017-04-01

Sepsis is a systemic immune response responsible for considerable morbidity and mortality. Molecular modeling of host-pathogen interactions in the disease state represents a promising strategy to define molecular events of importance for the transition from superficial to invasive infectious diseases. Here we used the Gram-positive bacterium Streptococcus pyogenes as a model system to establish a mass spectrometry based workflow for the construction of a stoichiometric surface density model between the S. pyogenes surface, the surface virulence factor M-protein, and adhered human blood plasma proteins. The workflow relies on stable isotope labeled reference peptides and selected reaction monitoring mass spectrometry analysis of a wild-type strain and an M-protein deficient mutant strain, to generate absolutely quantified protein stoichiometry ratios between S. pyogenes and interacting plasma proteins. The stoichiometry ratios in combination with a novel targeted mass spectrometry method to measure cell numbers enabled the construction of a stoichiometric surface density model using protein structures available from the protein data bank. The model outlines the topology and density of the host-pathogen protein interaction network on the S. pyogenes bacterial surface, revealing a dense and highly organized protein interaction network. Removal of the M-protein from S. pyogenes introduces a drastic change in the network topology, validated by electron microscopy. We propose that the stoichiometric surface density model of S. pyogenes in human blood plasma represents a scalable framework that can continuously be refined with the emergence of new results. Future integration of new results will improve the understanding of protein-protein interactions and their importance for bacterial virulence. Furthermore, we anticipate that the general properties of the developed workflow will facilitate the production of stoichiometric surface density models for other types of host-pathogen interactions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Global phenotypic characterisation of human platelet lysate expanded MSCs by high-throughput flow cytometry.

PubMed

Reis, Monica; McDonald, David; Nicholson, Lindsay; Godthardt, Kathrin; Knobel, Sebastian; Dickinson, Anne M; Filby, Andrew; Wang, Xiao-Nong

2018-03-02

Mesenchymal stromal cells (MSCs) are a promising cell source to develop cell therapy for many diseases. Human platelet lysate (PLT) is increasingly used as an alternative to foetal calf serum (FCS) for clinical-scale MSC production. To date, the global surface protein expression of PLT-expended MSCs (MSC-PLT) is not known. To investigate this, paired MSC-PLT and MSC-FCS were analysed in parallel using high-throughput flow cytometry for the expression of 356 cell surface proteins. MSC-PLT showed differential surface protein expression compared to their MSC-FCS counterpart. Higher percentage of positive cells was observed in MSC-PLT for 48 surface proteins, of which 13 were significantly enriched on MSC-PLT. This finding was validated using multiparameter flow cytometry and further confirmed by quantitative staining intensity analysis. The enriched surface proteins are relevant to increased proliferation and migration capacity, as well as enhanced chondrogenic and osteogenic differentiation properties. In silico network analysis revealed that these enriched surface proteins are involved in three distinct networks that are associated with inflammatory responses, carbohydrate metabolism and cellular motility. This is the first study reporting differential cell surface protein expression between MSC-PLT and MSC-FSC. Further studies are required to uncover the impact of those enriched proteins on biological functions of MSC-PLT.

Modulation of Protein Fouling and Interfacial Properties at Carbon Surfaces via Immobilization of Glycans Using Aryldiazonium Chemistry

PubMed Central

Zen, Federico; Angione, M. Daniela; Behan, James A.; Cullen, Ronan J.; Duff, Thomas; Vasconcelos, Joana M.; Scanlan, Eoin M.; Colavita, Paula E.

2016-01-01

Carbon materials and nanomaterials are of great interest for biological applications such as implantable devices and nanoparticle vectors, however, to realize their potential it is critical to control formation and composition of the protein corona in biological media. In this work, protein adsorption studies were carried out at carbon surfaces functionalized with aryldiazonium layers bearing mono- and di-saccharide glycosides. Surface IR reflectance absorption spectroscopy and quartz crystal microbalance were used to study adsorption of albumin, lysozyme and fibrinogen. Protein adsorption was found to decrease by 30–90% with respect to bare carbon surfaces; notably, enhanced rejection was observed in the case of the tested di-saccharide vs. simple mono-saccharides for near-physiological protein concentration values. ζ-potential measurements revealed that aryldiazonium chemistry results in the immobilization of phenylglycosides without a change in surface charge density, which is known to be important for protein adsorption. Multisolvent contact angle measurements were used to calculate surface free energy and acid-base polar components of bare and modified surfaces based on the van Oss-Chaudhury-Good model: results indicate that protein resistance in these phenylglycoside layers correlates positively with wetting behavior and Lewis basicity. PMID:27108562

Modulation of Protein Fouling and Interfacial Properties at Carbon Surfaces via Immobilization of Glycans Using Aryldiazonium Chemistry

NASA Astrophysics Data System (ADS)

Zen, Federico; Angione, M. Daniela; Behan, James A.; Cullen, Ronan J.; Duff, Thomas; Vasconcelos, Joana M.; Scanlan, Eoin M.; Colavita, Paula E.

2016-04-01

Carbon materials and nanomaterials are of great interest for biological applications such as implantable devices and nanoparticle vectors, however, to realize their potential it is critical to control formation and composition of the protein corona in biological media. In this work, protein adsorption studies were carried out at carbon surfaces functionalized with aryldiazonium layers bearing mono- and di-saccharide glycosides. Surface IR reflectance absorption spectroscopy and quartz crystal microbalance were used to study adsorption of albumin, lysozyme and fibrinogen. Protein adsorption was found to decrease by 30-90% with respect to bare carbon surfaces; notably, enhanced rejection was observed in the case of the tested di-saccharide vs. simple mono-saccharides for near-physiological protein concentration values. ζ-potential measurements revealed that aryldiazonium chemistry results in the immobilization of phenylglycosides without a change in surface charge density, which is known to be important for protein adsorption. Multisolvent contact angle measurements were used to calculate surface free energy and acid-base polar components of bare and modified surfaces based on the van Oss-Chaudhury-Good model: results indicate that protein resistance in these phenylglycoside layers correlates positively with wetting behavior and Lewis basicity.

Construction of a cell-surface display system based on the N-terminal domain of ice nucleation protein and its application in identification of mycoplasma adhesion proteins.

PubMed

Bao, S; Yu, S; Guo, X; Zhang, F; Sun, Y; Tan, L; Duan, Y; Lu, F; Qiu, X; Ding, C

2015-07-01

To construct and demonstrate a surface display system that could be used to identify mycoplasma adhesion proteins. Using the N-terminal domain of InaZ (InaZN) as the anchoring motif and the enhanced green fluorescent protein (EGFP) as the reporter, the surface display system pET-InaZN-EGFP was constructed. Then, the mgc2 gene which encodes an adhesin and the holB gene which encodes DNA polymerase III subunit delta' (nonadhesin, negative control) of Mycoplasma gallisepticum were cloned into the pET-InaZN-EGFP respectively. The fusion proteins were expressed in Escherichia coli BL21 (DE3). The distribution of the fusion proteins in E. coli cells was determined using SDS-PAGE followed by Western blotting, based on cell fractionation. Escherichia coli cell surface display of the fusion protein was confirmed by immunofluorescence microscopy. The results indicated that the fusion proteins were not only anchored to the outer membrane fraction but also were successfully displayed on the surface of E. coli cells. Adhesion analysis of E. coli harbouring InaZN-EGFP-mgc2 to host cells showed that the MGC2-positive E. coli cells can effectively adhere to the surfaces of DF-1 cells. A surface display system using the InaZN as the anchoring motif and EGFP as the reporter was developed to identify putative adhesins of mycoplasma. Results indicated that adhesion by the cytadhesin-like protein MGC2 of mycoplasma can be reproduced using this surface display system. This is the first construction of surface display system which could be used to identify the adhesion proteins of mycoplasma. The method developed in this study can even be used to select and identify the adhesion proteins of other pathogens. © 2015 The Society for Applied Microbiology.

Revealing Surface Waters on an Antifreeze Protein by Fusion Protein Crystallography Combined with Molecular Dynamic Simulations.

PubMed

Sun, Tianjun; Gauthier, Sherry Y; Campbell, Robert L; Davies, Peter L

2015-10-08

Antifreeze proteins (AFPs) adsorb to ice through an extensive, flat, relatively hydrophobic surface. It has been suggested that this ice-binding site (IBS) organizes surface waters into an ice-like clathrate arrangement that matches and fuses to the quasi-liquid layer on the ice surface. On cooling, these waters join the ice lattice and freeze the AFP to its ligand. Evidence for the generality of this binding mechanism is limited because AFPs tend to crystallize with their IBS as a preferred protein-protein contact surface, which displaces some bound waters. Type III AFP is a 7 kDa globular protein with an IBS made up two adjacent surfaces. In the crystal structure of the most active isoform (QAE1), the part of the IBS that docks to the primary prism plane of ice is partially exposed to solvent and has clathrate waters present that match this plane of ice. The adjacent IBS, which matches the pyramidal plane of ice, is involved in protein-protein crystal contacts with few surface waters. Here we have changed the protein-protein contacts in the ice-binding region by crystallizing a fusion of QAE1 to maltose-binding protein. In this 1.9 Å structure, the IBS that fits the pyramidal plane of ice is exposed to solvent. By combining crystallography data with MD simulations, the surface waters on both sides of the IBS were revealed and match well with the target ice planes. The waters on the pyramidal plane IBS were loosely constrained, which might explain why other isoforms of type III AFP that lack the prism plane IBS are less active than QAE1. The AFP fusion crystallization method can potentially be used to force the exposure to solvent of the IBS on other AFPs to reveal the locations of key surface waters.

Unique surface adsorption behaviors of serum proteins on chemically uniform and alternating surfaces

NASA Astrophysics Data System (ADS)

Song, Sheng

With increasing interests of studying proteins adsorption on the surfaces with nanoscale features in biomedical field, it is crucial to have fundamental understandings on how the proteins are adsorbed on such a surface and what factors contribute to the driving forces of adsorption. Besides, exploring more available nanoscale templates would greatly offer more possibilities one could design surface bio-detection methods with favorable protein-surface interactions. Thus, to fulfill the purpose, the work in this dissertation has been made into three major sections. First, to probe the intermediate states which possibly exist between stable and unstable phases described in mean-field theory diagram, a solvent vapor annealing method is chosen to slowly induce the copolymer polystyrene-block-polyvinylpyridine (PS-b-PVP)'s both blocks undergoing micro-phase separations from initial spherical nanodomains into terminal cylindrical nanodomains. During this process, real time atomic force microscopy (AFM) has been conducted to capture other six intermediate states with different morphologies on the polymeric film surfaces. Secondly, upon recognizing each intermediate state, the solution of immunoglobulin gamma (IgG) proteins has been deposited on the surface and been rinsed off with buffer solution before the protein-bounded surface is imaged by AFM. It has been found IgG showing a strong adsorption preference on PS over P4VP block. Among all the six intermediate states, the proteins are almost exclusively adsorbed on PS nanodomains regardless the concentration and deposition time. Thirdly, a trinodular shape protein fibrinogen (Fg) is selected for investigating how geometry and surface charge of proteins would interplay with cylindrical nanodomains on a surface developed from Polystyrene -block-Poly-(methyl methacrylate) PS-b-PMMA. Also, Fg adsorptions on chemically homogeneous surfaces are included here to have a better contrast of showing how much difference it can make by using it on a nanoscale surface. Interestingly, higher concentration of protein solution promotes the occurrences of single phase packed Fg on the PS domain. The densely packed network has formed where each Fg keeps its main body in PS domain and leaves its two alpha C chains on nearby PMMA domain. We believe this conformation and orientation would maximize both the hydrophobic and electrostatic interactions between Fg and the underlying surface.

Determination of conformation and orientation of immobilized peptides and proteins at buried interfaces

NASA Astrophysics Data System (ADS)

Shen, Lei; Ulrich, Nathan W.; Mello, Charlene M.; Chen, Zhan

2015-01-01

Surface immobilized peptides/proteins have important applications such as antimicrobial coating and biosensing. We report a study of such peptides/proteins using sum frequency generation vibrational spectroscopy and ATR-FTIR. Immobilization on surfaces via physical adsorption and chemical coupling revealed that structures of chemically immobilized peptides are determined by immobilization sites, chemical environments, and substrate surfaces. In addition, controlling enzyme orientation by engineering the surface immobilization site demonstrated that structures can be well-correlated to measured chemical activity. This research facilitates the development of immobilized peptides/proteins with improved activities by optimizing their surface orientation and structure.

Electro-induced protein deposition on low-fouling surfaces

NASA Astrophysics Data System (ADS)

Cole, M. A.; Voelcker, N. H.; Thissen, H.

2007-12-01

Control over protein adsorption is a key issue for numerous biomedical applications ranging from diagnostic microarrays to tissue-engineered medical devices. Here, we describe a method for creating surfaces that prevent non-specific protein adsorption, which upon application of an external trigger can be transformed into surfaces showing high protein adsorption on demand. Silicon wafers were used as substrate materials upon which thin functional coatings were constructed by the deposition of an allylamine plasma polymer followed by high-density grafting of poly(ethylene oxide) aldehyde, resulting in a low-fouling surface. When the underlying highly doped silicon substrate was used as an electrode, the resulting electrostatic attraction between the electrode and charged proteins in solution induced protein deposition at the low-fouling interface. X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM) were used to characterize the surface modifications. Controlled protein adsorption experiments were carried out using horseradish peroxidase. The amount of protein deposited at the surface was then investigated by means of a colorimetric assay. It is expected that the concept described here will find use in a variety of biotechnological and biomedical applications, particularly in the area of biochips.

TM9/Phg1 and SadA proteins control surface expression and stability of SibA adhesion molecules in Dictyostelium.

PubMed

Froquet, Romain; le Coadic, Marion; Perrin, Jackie; Cherix, Nathalie; Cornillon, Sophie; Cosson, Pierre

2012-02-01

TM9 proteins form a family of conserved proteins with nine transmembrane domains essential for cellular adhesion in many biological systems, but their exact role in this process remains unknown. In this study, we found that genetic inactivation of the TM9 protein Phg1A dramatically decreases the surface levels of the SibA adhesion molecule in Dictyostelium amoebae. This is due to a decrease in sibA mRNA levels, in SibA protein stability, and in SibA targeting to the cell surface. A similar phenotype was observed in cells devoid of SadA, a protein that does not belong to the TM9 family but also exhibits nine transmembrane domains and is essential for cellular adhesion. A contact site A (csA)-SibA chimeric protein comprising only the transmembrane and cytosolic domains of SibA and the extracellular domain of the Dictyostelium surface protein csA also showed reduced stability and relocalization to endocytic compartments in phg1A knockout cells. These results indicate that TM9 proteins participate in cell adhesion by controlling the levels of adhesion proteins present at the cell surface.

Interactions of fat globule surface proteins during concentration of whole milk in a pilot-scale multiple-effect evaporator.

PubMed

Ye, Aiqian; Singh, Harjinder; Taylor, Michael W; Anema, Skelte G

2004-11-01

The changes in milk fat globules and fat globule surface proteins during concentration of whole milk using a pilot-scale multiple-effect evaporator were examined. The effects of heat treatment of milk at 95 degrees C for 20 s, prior to evaporation, on fat globule size and the milk fat globule membrane (MFGM) proteins were also determined. In both non-preheated and preheated whole milk, the size of milk fat globules decreased while the amount of total surface proteins at the fat globules increased as the milk passed through each effect of the evaporator. In non-preheated samples, the amount of caseins at the surface of fat globules increased markedly during evaporation with a relatively small increase in whey proteins. In preheated samples, both caseins and whey proteins were observed at the surface of fat globules and the amounts of these proteins increased during subsequent steps of evaporation. The major original MFGM proteins, xanthine oxidase, butyrophilin, PAS 6 and PAS 7, did not change during evaporation, however, PAS 6 and PAS 7 decreased during preheating. These results indicate that the proteins from the skim milk were adsorbed onto the fat globule surface when the milk fat globules were disrupted during evaporation.

Protein-Protein Interaction Site Predictions with Three-Dimensional Probability Distributions of Interacting Atoms on Protein Surfaces

PubMed Central

Chen, Ching-Tai; Peng, Hung-Pin; Jian, Jhih-Wei; Tsai, Keng-Chang; Chang, Jeng-Yih; Yang, Ei-Wen; Chen, Jun-Bo; Ho, Shinn-Ying; Hsu, Wen-Lian; Yang, An-Suei

2012-01-01

Protein-protein interactions are key to many biological processes. Computational methodologies devised to predict protein-protein interaction (PPI) sites on protein surfaces are important tools in providing insights into the biological functions of proteins and in developing therapeutics targeting the protein-protein interaction sites. One of the general features of PPI sites is that the core regions from the two interacting protein surfaces are complementary to each other, similar to the interior of proteins in packing density and in the physicochemical nature of the amino acid composition. In this work, we simulated the physicochemical complementarities by constructing three-dimensional probability density maps of non-covalent interacting atoms on the protein surfaces. The interacting probabilities were derived from the interior of known structures. Machine learning algorithms were applied to learn the characteristic patterns of the probability density maps specific to the PPI sites. The trained predictors for PPI sites were cross-validated with the training cases (consisting of 432 proteins) and were tested on an independent dataset (consisting of 142 proteins). The residue-based Matthews correlation coefficient for the independent test set was 0.423; the accuracy, precision, sensitivity, specificity were 0.753, 0.519, 0.677, and 0.779 respectively. The benchmark results indicate that the optimized machine learning models are among the best predictors in identifying PPI sites on protein surfaces. In particular, the PPI site prediction accuracy increases with increasing size of the PPI site and with increasing hydrophobicity in amino acid composition of the PPI interface; the core interface regions are more likely to be recognized with high prediction confidence. The results indicate that the physicochemical complementarity patterns on protein surfaces are important determinants in PPIs, and a substantial portion of the PPI sites can be predicted correctly with the physicochemical complementarity features based on the non-covalent interaction data derived from protein interiors. PMID:22701576

Adhesion, invasion and evasion: the many functions of the surface proteins of Staphylococcus aureus

PubMed Central

Foster, Timothy J.; Geoghegan, Joan A.; Ganesh, Vannakambadi K.; Höök, Magnus

2014-01-01

Staphylococcus aureus is an important opportunistic pathogen and persistently colonizes about 20% of the human population. Its surface is ‘decorated’ with proteins that are covalently anchored to the cell wall peptidoglycan. Structural and functional analysis has identified four distinct classes of surface proteins, of which microbial surface component recognizing adhesive matrix molecules (MSCRAMMs) are the largest class. These surface proteins have numerous functions, including adhesion to and invasion of host cells and tissues, evasion of immune responses and biofilm formation. Thus, cell wall-anchored proteins are essential virulence factors for the survival of S. aureus in the commensal state and during invasive infections, and targeting them with vaccines could combat S. aureus infections. PMID:24336184

Extractable Bacterial Surface Proteins in Probiotic–Host Interaction

PubMed Central

do Carmo, Fillipe L. R.; Rabah, Houem; De Oliveira Carvalho, Rodrigo D.; Gaucher, Floriane; Cordeiro, Barbara F.; da Silva, Sara H.; Le Loir, Yves; Azevedo, Vasco; Jan, Gwénaël

2018-01-01

Some Gram-positive bacteria, including probiotic ones, are covered with an external proteinaceous layer called a surface-layer. Described as a paracrystalline layer and formed by the self-assembly of a surface-layer-protein (Slp), this optional structure is peculiar. The surface layer per se is conserved and encountered in many prokaryotes. However, the sequence of the corresponding Slp protein is highly variable among bacterial species, or even among strains of the same species. Other proteins, including surface layer associated proteins (SLAPs), and other non-covalently surface-bound proteins may also be extracted with this surface structure. They can be involved a various functions. In probiotic Gram-positives, they were shown by different authors and experimental approaches to play a role in key interactions with the host. Depending on the species, and sometime on the strain, they can be involved in stress tolerance, in survival within the host digestive tract, in adhesion to host cells or mucus, or in the modulation of intestinal inflammation. Future trends include the valorization of their properties in the formation of nanoparticles, coating and encapsulation, and in the development of new vaccines. PMID:29670603

Conformational transition free energy profiles of an adsorbed, lattice model protein by multicanonical Monte Carlo simulation

NASA Astrophysics Data System (ADS)

Castells, Victoria; Van Tassel, Paul R.

2005-02-01

Proteins often undergo changes in internal conformation upon interacting with a surface. We investigate the thermodynamics of surface induced conformational change in a lattice model protein using a multicanonical Monte Carlo method. The protein is a linear heteropolymer of 27 segments (of types A and B) confined to a cubic lattice. The segmental order and nearest neighbor contact energies are chosen to yield, in the absence of an adsorbing surface, a unique 3×3×3 folded structure. The surface is a plane of sites interacting either equally with A and B segments (equal affinity surface) or more strongly with the A segments (A affinity surface). We use a multicanonical Monte Carlo algorithm, with configuration bias and jump walking moves, featuring an iteratively updated sampling function that converges to the reciprocal of the density of states 1/Ω(E), E being the potential energy. We find inflection points in the configurational entropy, S(E)=klnΩ(E), for all but a strongly adsorbing equal affinity surface, indicating the presence of free energy barriers to transition. When protein-surface interactions are weak, the free energy profiles F(E)=E-TS(E) qualitatively resemble those of a protein in the absence of a surface: a free energy barrier separates a folded, lowest energy state from globular, higher energy states. The surface acts in this case to stabilize the globular states relative to the folded state. When the protein surface interactions are stronger, the situation differs markedly: the folded state no longer occurs at the lowest energy and free energy barriers may be absent altogether.

Evaluating the efficacy of subcellular fractionation of blast cells using live cell labeling and 2D DIGE.

PubMed

Ho, Yin Ying; Penno, Megan; Perugini, Michelle; Lewis, Ian; Hoffmann, Peter

2012-01-01

Labeling of exposed cell surface proteins of live cells using CyDye DIGE fluor minimal dyes is an efficient strategy for cell surface proteome profiling and quantifying differentially expressed proteins in diseases. Here we describe a strategy to evaluate a two-step detergent-based protein fractionation method using live cell labeling followed by visualization of the fluorescently labeled cell surface proteins and fractionated proteins within a single 2D gel.

Mussel adhesion is dictated by time-regulated secretion and molecular conformation of mussel adhesive proteins

NASA Astrophysics Data System (ADS)

Petrone, Luigi; Kumar, Akshita; Sutanto, Clarinda N.; Patil, Navinkumar J.; Kannan, Srinivasaraghavan; Palaniappan, Alagappan; Amini, Shahrouz; Zappone, Bruno; Verma, Chandra; Miserez, Ali

2015-10-01

Interfacial water constitutes a formidable barrier to strong surface bonding, hampering the development of water-resistant synthetic adhesives. Notwithstanding this obstacle, the Asian green mussel Perna viridis attaches firmly to underwater surfaces via a proteinaceous secretion (byssus). Extending beyond the currently known design principles of mussel adhesion, here we elucidate the precise time-regulated secretion of P. viridis mussel adhesive proteins. The vanguard 3,4-dihydroxy-L-phenylalanine (Dopa)-rich protein Pvfp-5 acts as an adhesive primer, overcoming repulsive hydration forces by displacing surface-bound water and generating strong surface adhesion. Using homology modelling and molecular dynamics simulations, we find that all mussel adhesive proteins are largely unordered, with Pvfp-5 adopting a disordered structure and elongated conformation whereby all Dopa residues reside on the protein surface. Time-regulated secretion and structural disorder of mussel adhesive proteins appear essential for optimizing extended nonspecific surface interactions and byssus' assembly. Our findings reveal molecular-scale principles to help the development of wet-resistant adhesives.

Protein covalent immobilization via its scarce thiol versus abundant amine groups: Effect on orientation, cell binding domain exposure and conformational lability.

PubMed

Ba, O M; Hindie, M; Marmey, P; Gallet, O; Anselme, K; Ponche, A; Duncan, A C

2015-10-01

Quantity, orientation, conformation and covalent linkage of naturally cell adhesive proteins adsorbed or covalently linked to a surface, are known to influence the preservation of their subsequent long term cell adhesion properties and bioactivity. In the present work, we explore two different strategies for the covalent linking of plasma fibronectin (pFN) - used as a cell adhesive model protein, onto a polystyrene (PS) surface. One is aimed at tethering the protein to the surface in a semi-oriented fashion (via one of the 4 free thiol reactive groups on the protein) with a heterofunctional coupling agent (SSMPB method). The other aims to immobilize the protein in a more random fashion by reaction between the abundant pendant primary amine bearing amino acids of the pFN and activated carboxylic surface functions obtained after glutaric anhydride surface treatment (GA method). The overall goal will be to verify the hypothesis of a correlation between covalent immobilization of a model cell adhesive protein to a PS surface in a semi-oriented configuration (versus randomly oriented) with promotion of enhanced exposure of the protein's cell binding domain. This in turn would lead to enhanced cell adhesion. Ideally the goal is to elaborate substrates exhibiting a long term stable protein monolayer with preserved cell adhesive properties and bioactivity for biomaterial and/or cell adhesion commercial plate applications. However, the initial restrictive objective of this paper is to first quantitatively and qualitatively investigate the reversibly (merely adsorbed) versus covalently irreversibly bound protein to the surface after the immobilization procedure. Although immobilized surface amounts were similar (close to the monolayer range) for all immobilization approaches, covalent grafting showed improved retention and stronger "tethering" of the pFN protein to the surface (roughly 40%) after SDS rinsing compared to that for mere adsorption (0%) suggesting an added value to the covalent grafting immobilization methods. However no differences in exposure of the cell binding domains were observed (ELISA results) before SDS rinsing, suggesting that pFN protein grafting to the surface is initially kinetically driven be a stochastic random adsorption phenomenon. Covalent grafting acts in the final stage as a process that simply tethers and stabilizes (or freezes) the initial conformation/orientation of the adsorbed protein on the surface. In addition covalent linkage via the SSMPB approach is likely favored by surface-induce exposure of one of the normally hidden free thiol group pair, thus optimizing covalent linkage to the surface. However after SDS rinsing, this "tethering"/"freezing" effect was significantly more prominent for the GA grafting approach (due to greater number of potential covalent links between the protein and the surface) compared to that for the SSMPB approach. This hypothesis was buttressed by the improved resistance to denaturation (smaller conformational lability) for the GA compared to the SMPB approach and improved exposure of the cell binding domain for the former (>50%) even after SDS rinsing. These results are promising in that they suggest covalent tethering of fibronectin to PS substrate in a monolayer range, with significantly improved irreversible protein surface bonding via both approaches (compared to that for mere adsorption). The latter are likely applicable to a wide range of proteins. Copyright © 2015 Elsevier B.V. All rights reserved.

Hidden complexity of free energy surfaces for peptide (protein) folding.

PubMed

Krivov, Sergei V; Karplus, Martin

2004-10-12

An understanding of the thermodynamics and kinetics of protein folding requires a knowledge of the free energy surface governing the motion of the polypeptide chain. Because of the many degrees of freedom involved, surfaces projected on only one or two progress variables are generally used in descriptions of the folding reaction. Such projections result in relatively smooth surfaces, but they could mask the complexity of the unprojected surface. Here we introduce an approach to determine the actual (unprojected) free energy surface and apply it to the second beta-hairpin of protein G, which has been used as a model system for protein folding. The surface is represented by a disconnectivity graph calculated from a long equilibrium folding-unfolding trajectory. The denatured state is found to have multiple low free energy basins. Nevertheless, the peptide shows exponential kinetics in folding to the native basin. Projected surfaces obtained from the present analysis have a simple form in agreement with other studies of the beta-hairpin. The hidden complexity found for the beta-hairpin surface suggests that the standard funnel picture of protein folding should be revisited.

sl(1|2) Super-Toda Fields

NASA Astrophysics Data System (ADS)

Yang, Zhan-Ying; Xue, Pan-Pan; Zhao, Liu; Shi, Kang-Jie

2008-11-01

Explicit exact solution of supersymmetric Toda fields associated with the Lie superalgebra sl(2|1) is constructed. The approach used is a super extension of Leznov Saveliev algebraic analysis, which is based on a pair of chiral and antichiral Drienfeld Sokolov systems. Though such approach is well understood for Toda field theories associated with ordinary Lie algebras, its super analogue was only successful in the super Liouville case with the underlying Lie superalgebra osp(1|2). The problem lies in that a key step in the construction makes use of the tensor product decomposition of the highest weight representations of the underlying Lie superalgebra, which is not clear until recently. So our construction made in this paper presents a first explicit example of Leznov Saveliev analysis for super Toda systems associated with underlying Lie superalgebras of the rank higher than 1.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bauer, R.; Ebersberger, B.; Kupfer, C.

SnAg solder bump is one bump type which is used to replace eutectic SnPb bumps. In this work tests have been done to characterize the reliability properties of this bump type. Electromigration (EM) tests, which were accelerated by high current and high temperature and high temperature storage (HTS) tests were performed. It was found that the reliability properties are sensitive to the material combinations in the interconnect stack. The interconnect stack includes substrate pad, pad finish, bump, underbump metallization (UBM) and the chip pad. Therefore separate test groups for SnAg bumps on Cu substrate pads with organic solderability preservative (OSP)more » finish and the identical bumps on pads with Ni/Au finish were used. In this paper the reliability test results and the corresponding failure analysis are presented. Some explanations about the differences in formation of intermetallic compounds (IMCs) are given.« less

Post-translational processing targets functionally diverse proteins in Mycoplasma hyopneumoniae

PubMed Central

Tacchi, Jessica L.; Raymond, Benjamin B. A.; Haynes, Paul A.; Berry, Iain J.; Widjaja, Michael; Bogema, Daniel R.; Woolley, Lauren K.; Jenkins, Cheryl; Minion, F. Chris; Padula, Matthew P.; Djordjevic, Steven P.

2016-01-01

Mycoplasma hyopneumoniae is a genome-reduced, cell wall-less, bacterial pathogen with a predicted coding capacity of less than 700 proteins and is one of the smallest self-replicating pathogens. The cell surface of M. hyopneumoniae is extensively modified by processing events that target the P97 and P102 adhesin families. Here, we present analyses of the proteome of M. hyopneumoniae-type strain J using protein-centric approaches (one- and two-dimensional GeLC–MS/MS) that enabled us to focus on global processing events in this species. While these approaches only identified 52% of the predicted proteome (347 proteins), our analyses identified 35 surface-associated proteins with widely divergent functions that were targets of unusual endoproteolytic processing events, including cell adhesins, lipoproteins and proteins with canonical functions in the cytosol that moonlight on the cell surface. Affinity chromatography assays that separately used heparin, fibronectin, actin and host epithelial cell surface proteins as bait recovered cleavage products derived from these processed proteins, suggesting these fragments interact directly with the bait proteins and display previously unrecognized adhesive functions. We hypothesize that protein processing is underestimated as a post-translational modification in genome-reduced bacteria and prokaryotes more broadly, and represents an important mechanism for creating cell surface protein diversity. PMID:26865024

Characterizing and modeling protein-surface interactions in lab-on-chip devices

NASA Astrophysics Data System (ADS)

Katira, Parag

Protein adsorption on surfaces determines the response of other biological species present in the surrounding solution. This phenomenon plays a major role in the design of biomedical and biotechnological devices. While specific protein adsorption is essential for device function, non-specific protein adsorption leads to the loss of device function. For example, non-specific protein adsorption on bioimplants triggers foreign body response, in biosensors it leads to reduced signal to noise ratios, and in hybrid bionanodevices it results in the loss of confinement and directionality of molecular shuttles. Novel surface coatings are being developed to reduce or completely prevent the non-specific adsorption of proteins to surfaces. A novel quantification technique for extremely low protein coverage on surfaces has been developed. This technique utilizes measurement of the landing rate of microtubule filaments on kinesin proteins adsorbed on a surface to determine the kinesin density. Ultra-low limits of detection, dynamic range, ease of detection and availability of a ready-made kinesin-microtubule kit makes this technique highly suitable for detecting protein adsorption below the detection limits of standard techniques. Secondly, a random sequential adsorption model is presented for protein adsorption to PEO-coated surfaces. The derived analytical expressions accurately predict the observed experimental results from various research groups, suggesting that PEO chains act as almost perfect steric barriers to protein adsorption. These expressions can be used to predict the performance of a variety of systems towards resisting protein adsorption and can help in the design of better non-fouling surface coatings. Finally, in biosensing systems, target analytes are captured and concentrated on specifically adsorbed proteins for detection. Non-specific adsorption of proteins results in the loss of signal, and an increase in the background. The use of nanoscale transducers as receptors is beneficial from the point of view of signal enhancement, but has a strong mass transport limitation. To overcome this, the use of molecular shuttles has been proposed that can selectively capture analytes and actively transport them to the nanoreceptors. The effect of employing such a two-stage capture process on biosensor sensitivity is theoretically investigated and an optimum design for a kinesin-microtubule-driven hybrid biosensor is proposed.

Targeted Identification of Metastasis-associated Cell-surface Sialoglycoproteins in Prostate Cancer*

PubMed Central

Yang, Lifang; Nyalwidhe, Julius O.; Guo, Siqi; Drake, Richard R.; Semmes, O. John

2011-01-01

Covalent attachment of carbohydrates to proteins is one of the most common post-translational modifications. At the cell surface, sugar moieties of glycoproteins contribute to molecular recognition events involved in cancer metastasis. We have combined glycan metabolic labeling with mass spectrometry analysis to identify and characterize metastasis-associated cell surface sialoglycoproteins. Our model system used syngeneic prostate cancer cell lines derived from PC3 (N2, nonmetastatic, and ML2, highly metastatic). The metabolic incorporation of AC4ManNAz and subsequent specific labeling of cell surface sialylation was confirmed by flow cytometry and confocal microscopy. Affinity isolation of the modified sialic-acid containing cell surface proteins via click chemistry was followed by SDS-PAGE separation and liquid chromatography-tandem MS analysis. We identified 324 proteins from N2 and 372 proteins of ML2. Using conservative annotation, 64 proteins (26%) from N2 and 72 proteins (29%) from ML2 were classified as extracellular or membrane-associated glycoproteins. A selective enrichment of sialoglycoproteins was confirmed. When compared with global proteomic analysis of the same cells, the proportion of identified glycoprotein and cell-surface proteins were on average threefold higher using the selective capture approach. Functional clustering of differentially expressed proteins by Ingenuity Pathway Analysis revealed that the vast majority of glycoproteins overexpressed in the metastatic ML2 subline were involved in cell motility, migration, and invasion. Our approach effectively targeted surface sialoglycoproteins and efficiently identified proteins that underlie the metastatic potential of the ML2 cells. PMID:21447706

Targeted identification of metastasis-associated cell-surface sialoglycoproteins in prostate cancer.

PubMed

Yang, Lifang; Nyalwidhe, Julius O; Guo, Siqi; Drake, Richard R; Semmes, O John

2011-06-01

Covalent attachment of carbohydrates to proteins is one of the most common post-translational modifications. At the cell surface, sugar moieties of glycoproteins contribute to molecular recognition events involved in cancer metastasis. We have combined glycan metabolic labeling with mass spectrometry analysis to identify and characterize metastasis-associated cell surface sialoglycoproteins. Our model system used syngeneic prostate cancer cell lines derived from PC3 (N2, nonmetastatic, and ML2, highly metastatic). The metabolic incorporation of AC(4)ManNAz and subsequent specific labeling of cell surface sialylation was confirmed by flow cytometry and confocal microscopy. Affinity isolation of the modified sialic-acid containing cell surface proteins via click chemistry was followed by SDS-PAGE separation and liquid chromatography-tandem MS analysis. We identified 324 proteins from N2 and 372 proteins of ML2. Using conservative annotation, 64 proteins (26%) from N2 and 72 proteins (29%) from ML2 were classified as extracellular or membrane-associated glycoproteins. A selective enrichment of sialoglycoproteins was confirmed. When compared with global proteomic analysis of the same cells, the proportion of identified glycoprotein and cell-surface proteins were on average threefold higher using the selective capture approach. Functional clustering of differentially expressed proteins by Ingenuity Pathway Analysis revealed that the vast majority of glycoproteins overexpressed in the metastatic ML2 subline were involved in cell motility, migration, and invasion. Our approach effectively targeted surface sialoglycoproteins and efficiently identified proteins that underlie the metastatic potential of the ML2 cells.

Interpreting the adsorption of serum albumin and lactoglobulin onto ZnS nanopaticles: effect of conformational rigidity of the proteins.

PubMed

Saikia, Jiban; Saha, Bedabrata; Das, Gopal

2014-02-15

The work we have undertaken is to investigate the adsorption of two different proteins (BSA and BLG) having near same IEP and differing in their conformational flexibility, onto the surface of ZnS nanoparticles (ZnS NPs). BSA and BLG both have an IEP value around pH~5. BSA is more prone to conformational deformation and considered "soft" while BLG holds the conformational rigidity and considered as "hard" protein. To ascertain the differences in surface coverage and conformation of the protein onto ZnS surface (PZC ~ 3.7), we have evaluated the adsorption profile at pH 7, where the entire surface behaves negatively. An integrated approach was taken by incorporating zeta (ζ) potential, fluorescence and CD for analyzing the adsorption process. In both systems, an increase in protein surface coverage was observed with the increase in free protein concentration in the solution and ζ values approaching that of native protein at high surface coverage. An alteration in the tertiary structure was observed for both BSA and BLG. The CD spectra analysis reveals that the secondary structure of the BSA was more deviated from the native protein structure, accommodating the increased adsorption value. For BLG no such prominent structural alteration was observed. These findings help us to understand better, how adjustment of the protein adsorption amount can be achieved onto the surface of nanoparticles having like charges. Copyright © 2013 Elsevier Inc. All rights reserved.

DIGE compatible labelling of surface proteins on vital cells in vitro and in vivo.

PubMed

Mayrhofer, Corina; Krieger, Sigurd; Allmaier, Günter; Kerjaschki, Dontscho

2006-01-01

Efficient methods for profiling of the cell surface proteome are desirable to get a deeper insight in basic biological processes, to localise proteins and to uncover proteins differentially expressed in diseases. Here we present a strategy to target cell surface exposed proteins via fluorescence labelling using CyDye DIGE fluors. This method has been applied to human cell lines in vitro as well as to a complex biological system in vivo. It allows detection of fluorophore-tagged cell surface proteins and visualisation of the accessible proteome within a single 2-D gel, simplifying subsequent UV MALDI-MS analysis.

Interfacial layers from the protein HFBII hydrophobin: dynamic surface tension, dilatational elasticity and relaxation times.

PubMed

Alexandrov, Nikola A; Marinova, Krastanka G; Gurkov, Theodor D; Danov, Krassimir D; Kralchevsky, Peter A; Stoyanov, Simeon D; Blijdenstein, Theodorus B J; Arnaudov, Luben N; Pelan, Eddie G; Lips, Alex

2012-06-15

The pendant-drop method (with drop-shape analysis) and Langmuir trough are applied to investigate the characteristic relaxation times and elasticity of interfacial layers from the protein HFBII hydrophobin. Such layers undergo a transition from fluid to elastic solid films. The transition is detected as an increase in the error of the fit of the pendant-drop profile by means of the Laplace equation of capillarity. The relaxation of surface tension after interfacial expansion follows an exponential-decay law, which indicates adsorption kinetics under barrier control. The experimental data for the relaxation time suggest that the adsorption rate is determined by the balance of two opposing factors: (i) the barrier to detachment of protein molecules from bulk aggregates and (ii) the attraction of the detached molecules by the adsorption layer due to the hydrophobic surface force. The hydrophobic attraction can explain why a greater surface coverage leads to a faster adsorption. The relaxation of surface tension after interfacial compression follows a different, square-root law. Such behavior can be attributed to surface diffusion of adsorbed protein molecules that are condensing at the periphery of interfacial protein aggregates. The surface dilatational elasticity, E, is determined in experiments on quick expansion or compression of the interfacial protein layers. At lower surface pressures (<11 mN/m) the experiments on expansion, compression and oscillations give close values of E that are increasing with the rise of surface pressure. At higher surface pressures, E exhibits the opposite tendency and the data are scattered. The latter behavior can be explained with a two-dimensional condensation of adsorbed protein molecules at the higher surface pressures. The results could be important for the understanding and control of dynamic processes in foams and emulsions stabilized by hydrophobins, as well as for the modification of solid surfaces by adsorption of such proteins. Copyright © 2012 Elsevier Inc. All rights reserved.

Chemical and physical effects on the adhesion, maturation, and survival of monocytes, macrophages, and foreign body giant cells

NASA Astrophysics Data System (ADS)

Collier, Terry Odell, III

Injury caused by biomedical device implantation initiates inflammatory and wound healing responses. Cells migrate to the site of injury to degrade bacteria and toxins, create new vasculature, and form new and repair injured tissue. Blood-proteins rapidly adsorb onto the implanted material surface and express adhesive ligands which mediate cell adhesion on the material surface. Monocyte-derived macrophages and multi-nucleated foreign body giant cells adhere to the surface and degrade the surface of the material. Due to the role of macrophage and foreign body giant cell on material biocompatibility and biostability, the effects of surface chemistry, surface topography and specific proteins on the maturation and survival of monocytes, macrophages and foreign body giant cells has been investigated. Novel molecularly designed materials were used to elucidate the dynamic interactions which occur between inflammatory cells, proteins and surfaces. The effect of protein and protein adhesion was investigated using adhesive protein depleted serum conditions on RGD-modified and silane modified surfaces. The effects of surface chemistry were investigated using temperature responsive surfaces of poly (N-isopropylacrylamide) and micropatterned surfaces of N-(2 aminoethyl)-3-aminopropyltrimethoxysilane regions on an interpenetrating polymer network of polyacrylamide and poly(ethylene glycol). The physical effects were investigated using polyimide scaffold materials and polyurethane materials with surface modifying end groups. The depletion of immunoglobulin G caused decreased levels of macrophage adhesion, foreign body giant cell formation and increased levels of apoptosis. The temporal nature of macrophage adhesion was observed with changing effectiveness of adherent cell detachment with time, which correlated to increased expression of beta1 integrin receptors on detached macrophages with time. The limited ability of the micropatterned surface, polyimide scaffold and surface modified polyurethane materials to control macrophage adhesion indicates the complexity of macrophage adhesion and protein adsorption onto a surface. These studies have indicated components and adhesive mechanisms which can be utilized to create materials with enhanced resistance to macrophage adhesion and/or degradative abilities.

Distinct Adsorption Configurations and Self-Assembly Characteristics of Fibrinogen on Chemically Uniform and Alternating Surfaces including Block Copolymer Nanodomains

PubMed Central

2015-01-01

Understanding protein–surface interactions is crucial to solid-state biomedical applications whose functionality is directly correlated with the precise control of the adsorption configuration, surface packing, loading density, and bioactivity of protein molecules. Because of the small dimensions and highly amphiphilic nature of proteins, investigation of protein adsorption performed on nanoscale topology can shed light on subprotein-level interaction preferences. In this study, we examine the adsorption and assembly behavior of a highly elongated protein, fibrinogen, on both chemically uniform (as-is and buffered HF-treated SiO2/Si, and homopolymers of polystyrene and poly(methyl methacrylate)) and varying (polystyrene-block-poly(methyl methacrylate)) surfaces. By focusing on high-resolution imaging of individual protein molecules whose configurations are influenced by protein–surface rather than protein–protein interactions, fibrinogen conformations characteristic to each surface are identified and statistically analyzed for structural similarities/differences in key protein domains. By exploiting block copolymer nanodomains whose repeat distance is commensurate with the length of the individual protein, we determine that fibrinogen exhibits a more neutral tendency for interaction with both polystyrene and poly(methyl methacrylate) blocks relative to the case of common globular proteins. Factors affecting fibrinogen–polymer interactions are discussed in terms of hydrophobic and electrostatic interactions. In addition, assembly and packing attributes of fibrinogen are determined at different loading conditions. Primary orientations of fibrinogen and its rearrangements with respect to the underlying diblock nanodomains associated with different surface coverage are explained by pertinent protein interaction mechanisms. On the basis of two-dimensional stacking behavior, a protein assembly model is proposed for the formation of an extended fibrinogen network on the diblock copolymer. PMID:24708538

Protein-Glass Surface Interactions and Ion Desalting in Electrospray Ionization with Submicron Emitters

NASA Astrophysics Data System (ADS)

Xia, Zije; Williams, Evan R.

2018-01-01

Theta glass electrospray emitters can rapidly mix solutions to investigate fast reactions that occur as quickly as 1 μs, but emitters with submicron tips have the unusual properties of desalting protein ions and affecting the observed abundances of some proteins as a result of protein-surface interactions. The role of protein physical properties on ion signal was investigated using 1.7 ± 0.1 μm and 269 ± 7 nm emitters and 100 mM aqueous ammonium acetate or ammonium bicarbonate solutions. Protein ion desalting occurs for both positive and negative ions. The signal of a mixture of proteins with the 269 nm tips is time-dependent and the order in which ions of each protein is observed is related to the expected strengths of the protein-surface interactions. These results indicate that it is not just the high surface-to-volume ratio that plays a role in protein adsorption and reduction or absence of initial ion signal, but the small diffusion distance and extremely low flow rates of the smaller emitters can lead to complete adsorption of some proteins and loss of signal until the adsorption sites are filled and the zeta potential is significantly reduced. After about 30 min, signals for a protein mixture from the two different size capillaries are similar. These results show the advantages of submicron emitters but also indicate that surface effects must be taken into account in experiments using such small tips or that coating the emitter surface to prevent adsorption should be considered. [Figure not available: see fulltext.

Molecular Characteristics and Biological Functions of Surface-Active and Surfactant Proteins.

PubMed

Sunde, Margaret; Pham, Chi L L; Kwan, Ann H

2017-06-20

Many critical biological processes take place at hydrophobic:hydrophilic interfaces, and a wide range of organisms produce surface-active proteins and peptides that reduce surface and interfacial tension and mediate growth and development at these boundaries. Microorganisms produce both small lipid-associated peptides and amphipathic proteins that allow growth across water:air boundaries, attachment to surfaces, predation, and improved bioavailability of hydrophobic substrates. Higher-order organisms produce surface-active proteins with a wide variety of functions, including the provision of protective foam environments for vulnerable reproductive stages, evaporative cooling, and gas exchange across airway membranes. In general, the biological functions supported by these diverse polypeptides require them to have an amphipathic nature, and this is achieved by a diverse range of molecular structures, with some proteins undergoing significant conformational change or intermolecular association to generate the structures that are surface active.

Water-wettable polypropylene fibers by facile surface treatment based on soy proteins.

PubMed

Salas, Carlos; Genzer, Jan; Lucia, Lucian A; Hubbe, Martin A; Rojas, Orlando J

2013-07-24

Modification of the wetting behavior of hydrophobic surfaces is essential in a variety of materials, including textiles and membranes that require control of fluid interactions, adhesion, transport processes, sensing, etc. This investigation examines the enhancement of wettability of an important class of textile materials, viz., polypropylene (PP) fibers, by surface adsorption of different proteins from soybeans, including soy flour, isolate,glycinin, and β-conglycinin. Detailed investigations of soy adsorption from aqueous solution (pH 7.4, 25 °C) on polypropylene thin films is carried out using quartz crystal microbalance (QCM) and surface plasmon resonance (SPR). A significant amount of protein adsorbs onto the PP surfaces primarily due to hydrophobic interactions. We establish that adsorption of a cationic surfactant, dioctadecyldimethylammonium bromide (DODA) onto PP surfaces prior to the protein deposition dramatically enhances its adsorption. The adsorption of proteins from native (PBS buffer, pH 7.4, 25 °C) and denatured conditions (PBS buffer, pH 7.4, 95 °C) onto DODA-treated PP leads to a high coverage of the proteins on the PP surface as confirmed by a significant improvement in water wettability. A shift in the contact angle from 128° to completely wettable surfaces (≈0°) is observed and confirmed by imaging experiments conducted with fluorescence tags. Furthermore, the results from wicking tests indicate that hydrophobic PP nonwovens absorb a significant amount of water after protein treatment, i.e., the PP-modified surfaces become completely hydrophilic.

Recent advances in surface functionalization techniques on polymethacrylate materials for optical biosensor applications.

PubMed

Hosseini, Samira; Ibrahim, Fatimah; Djordjevic, Ivan; Koole, Leo H

2014-06-21

Biosensor chips for immune-based assay systems have been investigated for their application in early diagnostics. The development of such systems strongly depends on the effective protein immobilization on polymer substrates. In order to achieve this complex heterogeneous interaction the polymer surface must be functionalized with chemical groups that are reactive towards proteins in a way that surface functional groups (such as carboxyl, -COOH; amine, -NH2; and hydroxyl, -OH) chemically or physically anchor the proteins to the polymer platform. Since the proteins are very sensitive towards their environment and can easily lose their activity when brought in close proximity to the solid surface, effective surface functionalization and high level of control over surface chemistry present the most important steps in the fabrication of biosensors. This paper reviews recent developments in surface functionalization and preparation of polymethacrylates for protein immobilization. Due to their versatility and cost effectiveness, this particular group of plastic polymers is widely used both in research and in industry.

Influence of the Amino Acid Sequence on Protein-Mineral Interactions in Soil

NASA Astrophysics Data System (ADS)

Chacon, S. S.; Reardon, P. N.; Purvine, S.; Lipton, M. S.; Washton, N.; Kleber, M.

2017-12-01

The intimate associations between protein and mineral surfaces have profound impacts on nutrient cycling in soil. Proteins are an important source of organic C and N, and a subset of proteins, extracellular enzymes (EE), can catalyze the depolymerization of soil organic matter (SOM). Our goal was to determine how variation in the amino acid sequence could influence a protein's susceptibility to become chemically altered by mineral surfaces to infer the fate of adsorbed EE function in soil. We hypothesized that (1) addition of charged amino acids would enhance the adsorption onto oppositely charged mineral surfaces (2) addition of aromatic amino acids would increase adsorption onto zero charged surfaces (3) Increase adsorption of modified proteins would enhance their susceptibility to alterations by redox active minerals. To test these hypotheses, we generated three engineered proxies of a model protein Gb1 (IEP 4.0, 6.2 kDA) by inserting either negatively charged, positively charged or aromatic amino acids in the second loop. These modified proteins were allowed to interact with functionally different mineral surfaces (goethite, montmorillonite, kaolinite and birnessite) at pH 5 and 7. We used LC-MS/MS and solution-state Heteronuclear Single Quantum Coherence Spectroscopy NMR to observe modifications on engineered proteins as a consequence to mineral interactions. Preliminary results indicate that addition of any amino acids to a protein increase its susceptibility to fragmentation and oxidation by redox active mineral surfaces, and alter adsorption to the other mineral surfaces. This suggest that not all mineral surfaces in soil may act as sorbents for EEs and chemical modification of their structure should also be considered as an explanation for decrease in EE activity. Fragmentation of proteins by minerals can bypass the need to produce proteases, but microbial acquisition of other nutrients that require enzymes such as cellulases, ligninases or phosphatases may be hampered by mineral association.

Modulating surface rheology by electrostatic protein/polysaccharide interactions.

PubMed

Ganzevles, Renate A; Zinoviadou, Kyriaki; van Vliet, Ton; Cohen, Martien A; de Jongh, Harmen H

2006-11-21

There is a large interest in mixed protein/polysaccharide layers at air-water and oil-water interfaces because of their ability to stabilize foams and emulsions. Mixed protein/polysaccharide adsorbed layers at air-water interfaces can be prepared either by adsorption of soluble protein/polysaccharide complexes or by sequential adsorption of complexes or polysaccharides to a previously formed protein layer. Even though the final protein and polysaccharide bulk concentrations are the same, the behavior of the adsorbed layers can be very different, depending on the method of preparation. The surface shear modulus of a sequentially formed beta-lactoglobulin/pectin layer can be up to a factor of 6 higher than that of a layer made by simultaneous adsorption. Furthermore, the surface dilatational modulus and surface shear modulus strongly (up to factors of 2 and 7, respectively) depend on the bulk -lactoglobulin/pectin mixing ratio. On the basis of the surface rheological behavior, a mechanistic understanding of how the structure of the adsorbed layers depends on the protein/polysaccharide interaction in bulk solution, mixing ratio, ionic strength, and order of adsorption to the interface (simultaneous or sequential) is derived. Insight into the effect of protein/polysaccharide interactions on the properties of adsorbed layers provides a solid basis to modulate surface rheological behavior.

Functional Cell Surface Display and Controlled Secretion of Diverse Agarolytic Enzymes by Escherichia coli with a Novel Ligation-Independent Cloning Vector Based on the Autotransporter YfaL

PubMed Central

Ko, Hyeok-Jin; Park, Eunhye; Song, Joseph; Yang, Taek Ho; Lee, Hee Jong; Kim, Kyoung Heon

2012-01-01

Autotransporters have been employed as the anchoring scaffold for cell surface display by replacing their passenger domains with heterologous proteins to be displayed. We adopted an autotransporter (YfaL) of Escherichia coli for the cell surface display system. The critical regions in YfaL for surface display were identified for the construction of a ligation-independent cloning (LIC)-based display system. The designed system showed no detrimental effect on either the growth of the host cell or overexpressing heterologous proteins on the cell surface. We functionally displayed monomeric red fluorescent protein (mRFP1) as a reporter protein and diverse agarolytic enzymes from Saccharophagus degradans 2-40, including Aga86C and Aga86E, which previously had failed to be functional expressed. The system could display different sizes of proteins ranging from 25.3 to 143 kDa. We also attempted controlled release of the displayed proteins by incorporating a tobacco etch virus protease cleavage site into the C termini of the displayed proteins. The maximum level of the displayed protein was 6.1 × 104 molecules per a single cell, which corresponds to 5.6% of the entire cell surface of actively growing E. coli. PMID:22344647

Architecture of a Host-Parasite Interface: Complex Targeting Mechanisms Revealed Through Proteomics.

PubMed

Gadelha, Catarina; Zhang, Wenzhu; Chamberlain, James W; Chait, Brian T; Wickstead, Bill; Field, Mark C

2015-07-01

Surface membrane organization and composition is key to cellular function, and membrane proteins serve many essential roles in endocytosis, secretion, and cell recognition. The surface of parasitic organisms, however, is a double-edged sword; this is the primary interface between parasites and their hosts, and those crucial cellular processes must be carried out while avoiding elimination by the host immune defenses. For extracellular African trypanosomes, the surface is partitioned such that all endo- and exocytosis is directed through a specific membrane region, the flagellar pocket, in which it is thought the majority of invariant surface proteins reside. However, very few of these proteins have been identified, severely limiting functional studies, and hampering the development of potential treatments. Here we used an integrated biochemical, proteomic and bioinformatic strategy to identify surface components of the human parasite Trypanosoma brucei. This surface proteome contains previously known flagellar pocket proteins as well as multiple novel components, and is significantly enriched in proteins that are essential for parasite survival. Molecules with receptor-like properties are almost exclusively parasite-specific, whereas transporter-like proteins are conserved in model organisms. Validation shows that the majority of surface proteome constituents are bona fide surface-associated proteins and, as expected, most present at the flagellar pocket. Moreover, the largest systematic analysis of trypanosome surface molecules to date provides evidence that the cell surface is compartmentalized into three distinct domains with free diffusion of molecules in each, but selective, asymmetric traffic between. This work provides a paradigm for the compartmentalization of a cell surface and a resource for its analysis. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Salivary protein adsorption and Streptococccus gordonii adhesion to dental material surfaces.

PubMed

Schweikl, Helmut; Hiller, Karl-Anton; Carl, Ulrich; Schweiger, Rainer; Eidt, Andreas; Ruhl, Stefan; Müller, Rainer; Schmalz, Gottfried

2013-10-01

The initial adhesion of microorganisms to clinically used dental biomaterials is influenced by physico-chemical parameters like hydrophobicity and pre-adsorption of salivary proteins. Here, polymethyl methacrylate (PMMA), polyethylene (PE), polytetrafluoroethylene (PTFE), silicone (Mucopren soft), silorane-based (Filtek Silorane) and methacrylate-based (Tetric EvoCeram) dental composites, a conventional glassionomer cement as well as cobalt-chromium-molybdenum (Co28Cr6Mo) and titanium (Ti6Al4V) were tested for adsorption of salivary proteins and adhesion of Streptococcus gordonii DL1. Wettability of material surfaces precoated with salivary proteins or left in phosphate-buffered saline was determined by the measurement of water contact angles. Amounts of adsorbed proteins were determined directly on material surfaces after biotinylation of amino groups and detection by horseradish peroxidase-conjugated avidin-D. The same technique was used to analyze for the binding of biotinylated bacteria to material surfaces. The highest amount of proteins (0.18μg/cm(2)) adsorbed to hydrophobic PTFE samples, and the lowest amount (0.025μg/cm(2)) was detected on silicone. The highest number of S. gordonii (3.2×10(4)CFU/mm(2)) adhered to the hydrophilic glassionomer cement surface coated with salivary proteins, and the lowest number (4×10(3)CFU/mm(2)) was found on the hydrophobic silorane-based composite. Hydrophobicity of pure material surfaces and the number of attached microorganisms were weakly negatively correlated. No such correlation between hydrophobicity and the number of bacteria was detected when surfaces were coated with salivary proteins. Functional groups added by the adsorption of specific salivary proteins to material surfaces are more relevant for initial bacterial adhesion than hydrophobicity as a physical property. Copyright © 2013 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

The effects of tether placement on antibody stability on surfaces

NASA Astrophysics Data System (ADS)

Grawe, Rebecca W.; Knotts, Thomas A.

2017-06-01

Despite their potential benefits, antibody microarrays have fallen short of performing reliably and have not found widespread use outside of the research setting. Experimental techniques have been unable to determine what is occurring on the surface of an atomic level, so molecular simulation has emerged as the primary method of investigating protein/surface interactions. Simulations of small proteins have indicated that the stability of the protein is a function of the residue on the protein where a tether is placed. The purpose of this research is to see whether these findings also apply to antibodies, with their greater size and complexity. To determine this, 24 tethering locations were selected on the antibody Protein Data Bank (PDB) ID: 1IGT. Replica exchange simulations were run on two different surfaces, one hydrophobic and one hydrophilic, to determine the degree to which these tethering sites stabilize or destabilize the antibody. Results showed that antibodies tethered to hydrophobic surfaces were in general less stable than antibodies tethered to hydrophilic surfaces. Moreover, the stability of the antibody was a function of the tether location on hydrophobic surfaces but not hydrophilic surfaces.

Redesigning of Microbial Cell Surface and Its Application to Whole-Cell Biocatalysis and Biosensors.

PubMed

Han, Lei; Zhao, Yukun; Cui, Shan; Liang, Bo

2018-06-01

Microbial cell surface display technology can redesign cell surfaces with functional proteins and peptides to endow cells some unique features. Foreign peptides or proteins are transported out of cells and immobilized on cell surface by fusing with anchoring proteins, which is an effective solution to avoid substance transfer limitation, enzyme purification, and enzyme instability. As the most frequently used prokaryotic and eukaryotic protein surface display system, bacterial and yeast surface display systems have been widely applied in vaccine, biocatalysis, biosensor, bioadsorption, and polypeptide library screening. In this review of bacterial and yeast surface display systems, different cell surface display mechanisms and their applications in biocatalysis as well as biosensors are described with their strengths and shortcomings. In addition to single enzyme display systems, multi-enzyme co-display systems are presented here. Finally, future developments based on our and other previous reports are discussed.

Equilibrium of adsorption of mixed milk protein/surfactant solutions at the water/air interface.

PubMed

Kotsmar, C; Grigoriev, D O; Xu, F; Aksenenko, E V; Fainerman, V B; Leser, M E; Miller, R

2008-12-16

Ellipsometry and surface profile analysis tensiometry were used to study and compare the adsorption behavior of beta-lactoglobulin (BLG)/C10DMPO, beta-casein (BCS)/C10DMPO and BCS/C12DMPO mixtures at the air/solution interface. The adsorption from protein/surfactant mixed solutions is of competitive nature. The obtained adsorption isotherms suggest a gradual replacement of the protein molecules at the interface with increasing surfactant concentration for all studied mixed systems. The thickness, refractive index, and the adsorbed amount of the respective adsorption layers, determined by ellipsometry, decrease monotonically and reach values close to those for a surface covered only by surfactant molecules, indicating the absence of proteins from a certain surfactant concentration on. These results correlate with the surface tension data. A continuous increase of adsorption layer thickness was observed up to this concentration, caused by the desorption of segments of the protein and transforming the thin surface layer into a rather diffuse and thick one. Replacement and structural changes of the protein molecules are discussed in terms of protein structure and surface activity of surfactant molecules. Theoretical models derived recently were used for the quantitative description of the equilibrium state of the mixed surface layers.

A Multi-technique Characterization of Adsorbed Protein Films: Orientation and Structure by ToF-SIMS, NEXAFS, SFG, and XPS

NASA Astrophysics Data System (ADS)

Baio, Joseph E.

There are many techniques that allow surface scientists to study interfaces. However, few are routinely applied to probe biological surfaces. The work presented here demonstrates how detailed information about the conformation, orientation, chemical state, and molecular structure of biological molecules immobilized onto a surface can be assessed by electron spectroscopy, mass spectrometry, and nonlinear vibrational spectroscopy techniques. This investigation began with the development of simple model systems (small proteins, and peptides) and evolved into a study of more complex --- real world systems. Initially, two model systems based on the chemical and electrostatic immobilization of a small rigid protein (Protein G B1 domain, 6kDa) were built to develop the capabilities of time-of-flight secondary ion mass spectrometry (ToFSIMS), near edge X-ray absorption fine structure spectroscopy (NEXAFS) and sum frequency generation (SFG) spectroscopy as tools to probe the structure of surface immobilized proteins. X-ray photoelectron spectroscopy (XPS) was used to measure the amount of immobilized protein and ToF-SIMS sampled the amino acid composition of the exposed surface of the protein film. Within the ToF-SIMS spectra, an enrichment of secondary ions from amino acids located at opposite ends of the proteins were used to describe protein orientation. SFG spectral peaks characteristic of ordered alpha-helix and beta-sheet elements were observed for both systems and the phase of the peaks indicated a predominantly upright orientation for both the covalent and electrostatic configurations. Polarization dependence of the NEXAFS signal from the N 1s to pi* transition of the peptide bonds that make up the beta-sheets also indicated protein ordering at the surface. Building upon the Protein G B1 studies, the orientation and structure of a surface immobilized antibody (HuLys Fv: variant of humanized anti-lysozyme variable fragment, 26kDa) was characterized across two immobilization schemes. This protein contained both a hexahistidine tag and a cysteine residue, introduced at opposite ends of the HuLys Fv, for immobilization onto nitrilotriacetic acid (NTA) and maleimide oligo- (ethylene glycol) (MEG)-terminated substrates. The thiol group on the cysteine residue selectively binds to the MEG groups, while the his-tag selectively binds to the Ni-loaded NTA groups. XPS was used to monitor protein coverage on both surfaces by following the change in the nitrogen atomic %. The ToF-SIMS data provided a clear differentiation between the two samples due to the intensity differences of secondary ions originating from asymmetrically located amino acids in HuLys Fv. Indicating that the HuLys Fv fragment when adsorbed into the NTA and MEG substrates will be induced into two different orientations. On the NTA substrate the protein's binding site is accessible, while on the MEG substrate the binding site is oriented towards the surface. By taking advantage of the electron pathway through the heme group in cytochrome c (CytoC) electrochemists have built sensors based upon CytoC immobilized onto functionalized metal electrodes. When immobilized onto a charged surface, CytoC, with its distribution of lysine and glutamate residues around its surface, should orient and form a well-ordered protein film. Here a detailed examination of CytoC orientation when electrostatically immobilized onto both amine (NH 3+) and carboxyl (COO-) functionalized gold is presented. Again, protein coverage, on both surfaces, was monitored by the change in the atomic % N, as determined by XPS. ToF-SIMS data demonstrated a clear separation between the two samples based on the intensity differences of secondary ions stemming from amino acids located asymmetrically within CytoC, indicating opposite orientations of the protein on the two different surfaces. Spectral features within the in situ sum frequency generation vibrational spectra, acquired for the protein interacting with positively and negatively charged surfaces, indicates that these electrostatic interactions do induce the protein into a well ordered film.

Prediction of Protein-Protein Interaction Sites Using Electrostatic Desolvation Profiles

PubMed Central

Fiorucci, Sébastien; Zacharias, Martin

2010-01-01

Abstract Protein-protein complex formation involves removal of water from the interface region. Surface regions with a small free energy penalty for water removal or desolvation may correspond to preferred interaction sites. A method to calculate the electrostatic free energy of placing a neutral low-dielectric probe at various protein surface positions has been designed and applied to characterize putative interaction sites. Based on solutions of the finite-difference Poisson equation, this method also includes long-range electrostatic contributions and the protein solvent boundary shape in contrast to accessible-surface-area-based solvation energies. Calculations on a large set of proteins indicate that in many cases (>90%), the known binding site overlaps with one of the six regions of lowest electrostatic desolvation penalty (overlap with the lowest desolvation region for 48% of proteins). Since the onset of electrostatic desolvation occurs even before direct protein-protein contact formation, it may help guide proteins toward the binding region in the final stage of complex formation. It is interesting that the probe desolvation properties associated with residue types were found to depend to some degree on whether the residue was outside of or part of a binding site. The probe desolvation penalty was on average smaller if the residue was part of a binding site compared to other surface locations. Applications to several antigen-antibody complexes demonstrated that the approach might be useful not only to predict protein interaction sites in general but to map potential antigenic epitopes on protein surfaces. PMID:20441756

Regular Nanoscale Protein Patterns via Directed Adsorption through Self-Assembled DNA Origami Masks.

PubMed

Ramakrishnan, Saminathan; Subramaniam, Sivaraman; Stewart, A Francis; Grundmeier, Guido; Keller, Adrian

2016-11-16

DNA origami has become a widely used method for synthesizing well-defined nanostructures with promising applications in various areas of nanotechnology, biophysics, and medicine. Recently, the possibility to transfer the shape of single DNA origami nanostructures into different materials via molecular lithography approaches has received growing interest due to the great structural control provided by the DNA origami technique. Here, we use ordered monolayers of DNA origami nanostructures with internal cavities on mica surfaces as molecular lithography masks for the fabrication of regular protein patterns over large surface areas. Exposure of the masked sample surface to negatively charged proteins results in the directed adsorption of the proteins onto the exposed surface areas in the holes of the mask. By controlling the buffer and adsorption conditions, the protein coverage of the exposed areas can be varied from single proteins to densely packed monolayers. To demonstrate the versatility of this approach, regular nanopatterns of four different proteins are fabricated: the single-strand annealing proteins Redβ and Sak, the iron-storage protein ferritin, and the blood protein bovine serum albumin (BSA). We furthermore demonstrate the desorption of the DNA origami mask after directed protein adsorption, which may enable the fabrication of hierarchical patterns composed of different protein species. Because selectivity in adsorption is achieved by electrostatic interactions between the proteins and the exposed surface areas, this approach may enable also the large-scale patterning of other charged molecular species or even nanoparticles.

Double-chimera proteins to enhance recruitment of endothelial cells and their progenitor cells.

PubMed

Behjati, M; Kazemi, M; Hashemi, M; Zarkesh-Esfahanai, S H; Bahrami, E; Hashemi-Beni, B; Ahmadi, R

2013-08-20

Enhanced attraction of selective vascular reparative cells is of great importance in order to increase vascular patency after endovascular treatments. We aimed to evaluate efficient attachment of endothelial cells and their progenitors on surfaces coated with mixture of specific antibodies, L-selectin and VE-cadherin, with prohibited platelet attachment. The most efficient conditions for coating of L-selectin-Fc chimera and VE-cadherin-Fc chimera proteins were first determined by protein coating on ELISA plates. The whole processes were repeated on titanium substrates, which are commonly used to coat stents. Endothelial progenitor cells (EPCs) and human umbilical vein endothelial cells (HUVECs) were isolated and characterized by flow cytometry. Cell attachment, growth, proliferation, viability and surface cytotoxicity were evaluated using nuclear staining and MTT assay. Platelet and cell attachment were evaluated using scanning electron microscopy. Optimal concentration of each protein for surface coating was 50 ng/ml. The efficacy of protein coating was both heat and pH independent. Calcium ions had significant impact on simultaneous dual-protein coating (P<0.05). Coating stability data revealed more than one year stability for these coated proteins at 4°C. L-selectin and VE-cadherin (ratio of 50:50) coated surface showed highest EPC and HUVEC attachment, viability and proliferation compared to single protein coated and non-coated titanium surfaces (P<0.05). This double coated surface did not show any cytotoxic effect. Surfaces coated with L-selectin and VE-cadherin are friendly surface for EPC and endothelial cell attachment with less platelet attachment. These desirable factors make the L-selectin and VE-cadherin coated surfaces perfect candidate endovascular device. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Modeling and simulation of protein-surface interactions: achievements and challenges.

PubMed

Ozboyaci, Musa; Kokh, Daria B; Corni, Stefano; Wade, Rebecca C

2016-01-01

Understanding protein-inorganic surface interactions is central to the rational design of new tools in biomaterial sciences, nanobiotechnology and nanomedicine. Although a significant amount of experimental research on protein adsorption onto solid substrates has been reported, many aspects of the recognition and interaction mechanisms of biomolecules and inorganic surfaces are still unclear. Theoretical modeling and simulations provide complementary approaches for experimental studies, and they have been applied for exploring protein-surface binding mechanisms, the determinants of binding specificity towards different surfaces, as well as the thermodynamics and kinetics of adsorption. Although the general computational approaches employed to study the dynamics of proteins and materials are similar, the models and force-fields (FFs) used for describing the physical properties and interactions of material surfaces and biological molecules differ. In particular, FF and water models designed for use in biomolecular simulations are often not directly transferable to surface simulations and vice versa. The adsorption events span a wide range of time- and length-scales that vary from nanoseconds to days, and from nanometers to micrometers, respectively, rendering the use of multi-scale approaches unavoidable. Further, changes in the atomic structure of material surfaces that can lead to surface reconstruction, and in the structure of proteins that can result in complete denaturation of the adsorbed molecules, can create many intermediate structural and energetic states that complicate sampling. In this review, we address the challenges posed to theoretical and computational methods in achieving accurate descriptions of the physical, chemical and mechanical properties of protein-surface systems. In this context, we discuss the applicability of different modeling and simulation techniques ranging from quantum mechanics through all-atom molecular mechanics to coarse-grained approaches. We examine uses of different sampling methods, as well as free energy calculations. Furthermore, we review computational studies of protein-surface interactions and discuss the successes and limitations of current approaches.

Probing the Orientation of Surface-Immobilized Protein G B1 Using ToF-SIMS Sum Frequency Generation and NEXAFS Spectroscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

L Baugh; T Weidner; J Baio

2011-12-31

The ability to orient active proteins on surfaces is a critical aspect of many medical technologies. An important related challenge is characterizing protein orientation in these surface films. This study uses a combination of time-of-flight secondary ion mass spectrometry (ToF-SIMS), sum frequency generation (SFG) vibrational spectroscopy, and near-edge X-ray absorption fine structure (NEXAFS) spectroscopy to characterize the orientation of surface-immobilized Protein G B1, a rigid 6 kDa domain that binds the Fc fragment of IgG. Two Protein G B1 variants with a single cysteine introduced at either end were immobilized via the cysteine thiol onto maleimide-oligo(ethylene glycol)-functionalized gold and baremore » gold substrates. X-ray photoelectron spectroscopy was used to measure the amount of immobilized protein, and ToF-SIMS was used to measure the amino acid composition of the exposed surface of the protein films and to confirm covalent attachment of protein thiol to the substrate maleimide groups. SFG and NEXAFS were used to characterize the ordering and orientation of peptide or side chain bonds. On both substrates and for both cysteine positions, ToF-SIMS data showed enrichment of mass peaks from amino acids located at the end of the protein opposite to the cysteine surface position as compared with nonspecifically immobilized protein, indicating end-on protein orientations. Orientation on the maleimide substrate was enhanced by increasing pH (7.0-9.5) and salt concentration (0-1.5 M NaCl). SFG spectral peaks characteristic of ordered {alpha}-helix and {beta}-sheet elements were observed for both variants but not for cysteine-free wild type protein on the maleimide surface. The phase of the {alpha}-helix and {beta}-sheet peaks indicated a predominantly upright orientation for both variants, consistent with an end-on protein binding configuration. Polarization dependence of the NEXAFS signal from the N 1s to {pi}* transition of {beta}-sheet peptide bonds also indicated protein ordering, with an estimated tilt angle of inner {beta}-strands of 40-50{sup o} for both variants (one variant more tilted than the other), consistent with SFG results. The combined results demonstrate the power of using complementary techniques to probe protein orientation on surfaces.« less

Expression of bovine non-classical major histocompatibility complex class I proteins in mouse P815 and human K562 cells.

PubMed

Parasar, Parveen; Wilhelm, Amanda; Rutigliano, Heloisa M; Thomas, Aaron J; Teng, Lihong; Shi, Bi; Davis, William C; Suarez, Carlos E; New, Daniel D; White, Kenneth L; Davies, Christopher J

2016-08-01

Major histocompatibility complex class I (MHC-I) proteins can be expressed as cell surface or secreted proteins. To investigate whether bovine non-classical MHC-I proteins are expressed as cell surface or secreted proteins, and to assess the reactivity pattern of monoclonal antibodies with non-classical MHC-I isoforms, we expressed the MHC proteins in murine P815 and human K562 (MHC-I deficient) cells. Following antibiotic selection, stably transfected cell lines were stained with H1A or W6/32 antibodies to detect expression of the MHC-I proteins by flow cytometry. Two non-classical proteins (BoLA-NC1*00501 and BoLA-NC3*00101) were expressed on the cell surface in both cell lines. Surprisingly, the BoLA-NC4*00201 protein was expressed on the cell membrane of human K562 but not mouse P815 cells. Two non-classical proteins (BoLA-NC1*00401, which lacks a transmembrane domain, and BoLA-NC2*00102) did not exhibit cell surface expression. Nevertheless, Western blot analyses demonstrated expression of the MHC-I heavy chain in all transfected cell lines. Ammonium-sulfate precipitation of proteins from culture supernatants showed that BoLA-NC1*00401 was secreted and that all surface expressed proteins where shed from the cell membrane by the transfected cells. Interestingly, the surface expressed MHC-I proteins were present in culture supernatants at a much higher concentration than BoLA-NC1*00401. This comprehensive study shows that bovine non-classical MHC-I proteins BoLA-NC1*00501, BoLA-NC3*00101, and BoLA-NC4*00201 are expressed as surface isoforms with the latter reaching the cell membrane only in K562 cells. Furthermore, it demonstrated that BoLA-NC1*00401 is a secreted isoform and that significant quantities of membrane associated MHC-I proteins can be shed from the cell membrane. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Organophosphonate biofunctionalization of diamond electrodes.

PubMed

Caterino, R; Csiki, R; Wiesinger, M; Sachsenhauser, M; Stutzmann, M; Garrido, J A; Cattani-Scholz, A; Speranza, Giorgio; Janssens, S D; Haenen, K

2014-08-27

The modification of the diamond surface with organic molecules is a crucial aspect to be considered for any bioapplication of this material. There is great interest in broadening the range of linker molecules that can be covalently bound to the diamond surface. In the case of protein immobilization, the hydropathicity of the surface has a major influence on the protein conformation and, thus, on the functionality of proteins immobilized at surfaces. For electrochemical applications, particular attention has to be devoted to avoid that the charge transfer between the electrode and the redox center embedded in the protein is hindered by a thick insulating linker layer. This paper reports on the grafting of 6-phosphonohexanoic acid on OH-terminated diamond surfaces, serving as linkers to tether electroactive proteins onto diamond surfaces. X-ray photoelectron spectroscopy (XPS) confirms the formation of a stable layer on the surface. The charge transfer between electroactive molecules and the substrate is studied by electrochemical characterization of the redox activity of aminomethylferrocene and cytochrome c covalently bound to the substrate through this linker. Our work demonstrates that OH-terminated diamond functionalized with 6-phosphonohexanoic acid is a suitable platform to interface redox-proteins, which are fundamental building blocks for many bioelectronics applications.

Enhanced initial protein adsorption on engineered nanostructured cubic zirconia.

PubMed

Sabirianov, R F; Rubinstein, A; Namavar, F

2011-04-14

Motivated by experimentally-observed biocompatibility enhancement of nanoengineered cubic zirconia (ZrO(2)) coatings to mesenchymal stromal cells, we have carried out computational analysis of the initial immobilization of one known structural fragment of the adhesive protein (fibronectin) on the corresponding surface. We constructed an atomistic model of the ZrO(2) nano-hillock of 3-fold symmetry based on Atom Force Microscopy and Transmission Electron Microscopy images. First principle quantum mechanical calculations show a substantial variation of electrostatic potential at the hillock due to the presence of surface features such as edges and vertexes. Using an implemented Monte Carlo simulated annealing method, we found the orientation of the immobilized protein on the ZrO(2) surface and the contribution of the amino acid residues from the protein sequence to the adsorption energy. Accounting for the variation of the dielectric permittivity at the protein-implant interface, we used a model distance-dependent dielectric function to describe the inter-atom electrostatic interactions in the adsorption potential. We found that the initial immobilization of the rigid protein fragment on the nanostructured pyramidal ZrO(2) surface is achieved with a magnitude of adsorption energy larger than that of the protein on the smooth (atomically flat) surface. The strong attractive electrostatic interactions are a major contributing factor in the enhanced adsorption at the nanostructured surface. In the case of adsorption on the flat, uncharged surface this factor is negligible. We show that the best electrostatic and steric fit of the protein to the inorganic surface corresponds to a minimum of the adsorption energy determined by the non-covalent interactions.

Biologically Complex Planar Cell Plasma Membranes Supported on Polyelectrolyte Cushions Enhance Transmembrane Protein Mobility and Retain Native Orientation.

PubMed

Liu, Han-Yuan; Chen, Wei-Liang; Ober, Christopher K; Daniel, Susan

2018-01-23

Reconstituted supported lipid bilayers (SLB) are widely used as in vitro cell-surface models because they are compatible with a variety of surface-based analytical techniques. However, one of the challenges of using SLBs as a model of the cell surface is the limited complexity in membrane composition, including the incorporation of transmembrane proteins and lipid diversity that may impact the activity of those proteins. Additionally, it is challenging to preserve the transmembrane protein native orientation, function, and mobility in SLBs. Here, we leverage the interaction between cell plasma membrane vesicles and polyelectrolyte brushes to create planar bilayers from cell plasma membrane vesicles that have budded from the cell surface. This approach promotes the direct incorporation of membrane proteins and other species into the planar bilayer without using detergent or reconstitution and preserves membrane constituents. Furthermore, the structure of the polyelectrolyte brush serves as a cushion between the planar bilayer and rigid supporting surface, limiting the interaction of the cytosolic domains of membrane proteins with this surface. Single particle tracking was used to analyze the motion of GPI-linked yellow fluorescent proteins (GPI-YFP) and neon-green fused transmembrane P2X2 receptors (P2X2-neon) and shows that this platform retains over 75% mobility of multipass transmembrane proteins in its native membrane environment. An enzyme accessibility assay confirmed that the protein orientation is preserved and results in the extracellular domain facing toward the bulk phase and the cytosolic side facing the support. Because the platform presented here retains the complexity of the cell plasma membrane and preserves protein orientation and mobility, it is a better representative mimic of native cell surfaces, which may find many applications in biological assays aimed at understanding cell membrane phenomena.

Gold nanoparticles: role of size and surface chemistry on blood protein adsorption

NASA Astrophysics Data System (ADS)

Benetti, F.; Fedel, M.; Minati, L.; Speranza, G.; Migliaresi, C.

2013-06-01

Material interaction with blood proteins is a critical issue, since it could influence the biological processes taking place in the body following implantation/injection. This is particularly important in the case of nanoparticles, where innovative properties, such as size and high surface to volume ratio can lead to a behavioral change with respect to bulk macroscopic materials and could be responsible for a potential risk for human health. The aim of this work was to compare gold nanoparticles (AuNP) and planar surfaces to study the role of surface curvature moving from the macro- to the nano-size in the process of blood protein adsorption. In the course of the study, different protocols were tested to optimize the analysis of protein adsorption on gold nanoparticles. AuNP with different size (10, 60 and 200 nm diameter) and surface coatings (citrate and polyethylene glycol) were carefully characterized. The stabilizing action of blood proteins adsorbed on AuNP was studied measuring the variation of size and solubility of the nanoparticles following incubation with single protein solutions (human serum albumin and fibrinogen) and whole blood plasma. In addition, we developed a method to elute proteins from AuNP to study the propensity of gold materials to adsorb plasma proteins in function of dimensional characteristics and surface chemistry. We showed a different efficacy of the various eluting media tested, proving that even the most aggressive agent cannot provide a complete detachment of the protein corona. Enhanced protein adsorption was evidenced on AuNP if compared to gold laminae (bare and PEGylated) used as macroscopic control, probably due to the superior AuNP surface reactivity.

Fabrication of nanometer- and micrometer-scale protein structures by site-specific immobilization of histidine-tagged proteins to aminosiloxane films with photoremovable protein-resistant protecting groups

DOE PAGES

Xia, Sijing; Cartron, Michael; Morby, James; ...

2016-01-28

The site-specific immobilization of histidine-tagged proteins to patterns formed by far-field and near-field exposure of films of aminosilanes with protein-resistant photolabile protecting groups is demonstrated. After deprotection of the aminosilane, either through a mask or using a scanning near-field optical microscope, the amine terminal groups are derivatized first with glutaraldehyde and then with N-(5-amino-1-carboxypentyl)iminodiacetic acid to yield a nitrilo-triacetic-acid-terminated surface. After complexation with Ni 2+, this surface binds histidine-tagged GFP and CpcA-PEB in a site-specific fashion. The chemistry is simple and reliable and leads to extensive surface functionalization. Bright fluorescence is observed in fluorescence microscopy images of micrometer- and nanometer-scalemore » patterns. X-ray photoelectron spectroscopy is used to study quantitatively the efficiency of photodeprotection and the reactivity of the modified surfaces. The efficiency of the protein binding process is investigated quantitatively by ellipsometry and by fluorescence microscopy. We find that regions of the surface not exposed to UV light bind negligible amounts of His-tagged proteins, indicating that the oligo(ethylene glycol) adduct on the nitrophenyl protecting group confers excellent protein resistance; in contrast, exposed regions bind His-GFP very effectively, yielding strong fluorescence that is almost completely removed on treatment of the surface with imidazole, confirming a degree of site-specific binding in excess of 90%. As a result, this simple strategy offers a versatile generic route to the spatially selective site-specific immobilization of proteins at surfaces.« less

Fabrication of Nanometer- and Micrometer-Scale Protein Structures by Site-Specific Immobilization of Histidine-Tagged Proteins to Aminosiloxane Films with Photoremovable Protein-Resistant Protecting Groups

PubMed Central

2016-01-01

The site-specific immobilization of histidine-tagged proteins to patterns formed by far-field and near-field exposure of films of aminosilanes with protein-resistant photolabile protecting groups is demonstrated. After deprotection of the aminosilane, either through a mask or using a scanning near-field optical microscope, the amine terminal groups are derivatized first with glutaraldehyde and then with N-(5-amino-1-carboxypentyl)iminodiacetic acid to yield a nitrilo-triacetic-acid-terminated surface. After complexation with Ni2+, this surface binds histidine-tagged GFP and CpcA-PEB in a site-specific fashion. The chemistry is simple and reliable and leads to extensive surface functionalization. Bright fluorescence is observed in fluorescence microscopy images of micrometer- and nanometer-scale patterns. X-ray photoelectron spectroscopy is used to study quantitatively the efficiency of photodeprotection and the reactivity of the modified surfaces. The efficiency of the protein binding process is investigated quantitatively by ellipsometry and by fluorescence microscopy. We find that regions of the surface not exposed to UV light bind negligible amounts of His-tagged proteins, indicating that the oligo(ethylene glycol) adduct on the nitrophenyl protecting group confers excellent protein resistance; in contrast, exposed regions bind His-GFP very effectively, yielding strong fluorescence that is almost completely removed on treatment of the surface with imidazole, confirming a degree of site-specific binding in excess of 90%. This simple strategy offers a versatile generic route to the spatially selective site-specific immobilization of proteins at surfaces. PMID:26820378

Molecular dynamics and Monte Carlo simulations resolve apparent diffusion rate differences for proteins confined in nanochannels

DOE PAGES

Tringe, J. W.; Ileri, N.; Levie, H. W.; ...

2015-08-01

We use Molecular Dynamics and Monte Carlo simulations to examine molecular transport phenomena in nanochannels, explaining four orders of magnitude difference in wheat germ agglutinin (WGA) protein diffusion rates observed by fluorescence correlation spectroscopy (FCS) and by direct imaging of fluorescently-labeled proteins. We first use the ESPResSo Molecular Dynamics code to estimate the surface transport distance for neutral and charged proteins. We then employ a Monte Carlo model to calculate the paths of protein molecules on surfaces and in the bulk liquid transport medium. Our results show that the transport characteristics depend strongly on the degree of molecular surface coverage.more » Atomic force microscope characterization of surfaces exposed to WGA proteins for 1000 s show large protein aggregates consistent with the predicted coverage. These calculations and experiments provide useful insight into the details of molecular motion in confined geometries.« less

Versatile multi-functionalization of protein nanofibrils for biosensor applications

NASA Astrophysics Data System (ADS)

Sasso, L.; Suei, S.; Domigan, L.; Healy, J.; Nock, V.; Williams, M. A. K.; Gerrard, J. A.

2014-01-01

Protein nanofibrils offer advantages over other nanostructures due to the ease in their self-assembly and the versatility of surface chemistry available. Yet, an efficient and general methodology for their post-assembly functionalization remains a significant challenge. We introduce a generic approach, based on biotinylation and thiolation, for the multi-functionalization of protein nanofibrils self-assembled from whey proteins. Biochemical characterization shows the effects of the functionalization onto the nanofibrils' surface, giving insights into the changes in surface chemistry of the nanostructures. We show how these methods can be used to decorate whey protein nanofibrils with several components such as fluorescent quantum dots, enzymes, and metal nanoparticles. A multi-functionalization approach is used, as a proof of principle, for the development of a glucose biosensor platform, where the protein nanofibrils act as nanoscaffolds for glucose oxidase. Biotinylation is used for enzyme attachment and thiolation for nanoscaffold anchoring onto a gold electrode surface. Characterization via cyclic voltammetry shows an increase in glucose-oxidase mediated current response due to thiol-metal interactions with the gold electrode. The presented approach for protein nanofibril multi-functionalization is novel and has the potential of being applied to other protein nanostructures with similar surface chemistry.Protein nanofibrils offer advantages over other nanostructures due to the ease in their self-assembly and the versatility of surface chemistry available. Yet, an efficient and general methodology for their post-assembly functionalization remains a significant challenge. We introduce a generic approach, based on biotinylation and thiolation, for the multi-functionalization of protein nanofibrils self-assembled from whey proteins. Biochemical characterization shows the effects of the functionalization onto the nanofibrils' surface, giving insights into the changes in surface chemistry of the nanostructures. We show how these methods can be used to decorate whey protein nanofibrils with several components such as fluorescent quantum dots, enzymes, and metal nanoparticles. A multi-functionalization approach is used, as a proof of principle, for the development of a glucose biosensor platform, where the protein nanofibrils act as nanoscaffolds for glucose oxidase. Biotinylation is used for enzyme attachment and thiolation for nanoscaffold anchoring onto a gold electrode surface. Characterization via cyclic voltammetry shows an increase in glucose-oxidase mediated current response due to thiol-metal interactions with the gold electrode. The presented approach for protein nanofibril multi-functionalization is novel and has the potential of being applied to other protein nanostructures with similar surface chemistry. Electronic supplementary information (ESI) available: Cyclic voltammetry characterization of biosensor platforms including bare Au electrodes (Fig. S1), biosensor response to various glucose concentrations (Fig. S2), and AFM roughness measurements due to WPNF modifications (Fig. S3). See DOI: 10.1039/c3nr05752f

A Parametric Rosetta Energy Function Analysis with LK Peptides on SAM Surfaces.

PubMed

Lubin, Joseph H; Pacella, Michael S; Gray, Jeffrey J

2018-05-08

Although structures have been determined for many soluble proteins and an increasing number of membrane proteins, experimental structure determination methods are limited for complexes of proteins and solid surfaces. An economical alternative or complement to experimental structure determination is molecular simulation. Rosetta is one software suite that models protein-surface interactions, but Rosetta is normally benchmarked on soluble proteins. For surface interactions, the validity of the energy function is uncertain because it is a combination of independent parameters from energy functions developed separately for solution proteins and mineral surfaces. Here, we assess the performance of the RosettaSurface algorithm and test the accuracy of its energy function by modeling the adsorption of leucine/lysine (LK)-repeat peptides on methyl- and carboxy-terminated self-assembled monolayers (SAMs). We investigated how RosettaSurface predictions for this system compare with the experimental results, which showed that on both surfaces, LK-α peptides folded into helices and LK-β peptides held extended structures. Utilizing this model system, we performed a parametric analysis of Rosetta's Talaris energy function and determined that adjusting solvation parameters offered improved predictive accuracy. Simultaneously increasing lysine carbon hydrophilicity and the hydrophobicity of the surface methyl head groups yielded computational predictions most closely matching the experimental results. De novo models still should be interpreted skeptically unless bolstered in an integrative approach with experimental data.

Local functional descriptors for surface comparison based binding prediction

PubMed Central

2012-01-01

Background Molecular recognition in proteins occurs due to appropriate arrangements of physical, chemical, and geometric properties of an atomic surface. Similar surface regions should create similar binding interfaces. Effective methods for comparing surface regions can be used in identifying similar regions, and to predict interactions without regard to the underlying structural scaffold that creates the surface. Results We present a new descriptor for protein functional surfaces and algorithms for using these descriptors to compare protein surface regions to identify ligand binding interfaces. Our approach uses descriptors of local regions of the surface, and assembles collections of matches to compare larger regions. Our approach uses a variety of physical, chemical, and geometric properties, adaptively weighting these properties as appropriate for different regions of the interface. Our approach builds a classifier based on a training corpus of examples of binding sites of the target ligand. The constructed classifiers can be applied to a query protein providing a probability for each position on the protein that the position is part of a binding interface. We demonstrate the effectiveness of the approach on a number of benchmarks, demonstrating performance that is comparable to the state-of-the-art, with an approach with more generality than these prior methods. Conclusions Local functional descriptors offer a new method for protein surface comparison that is sufficiently flexible to serve in a variety of applications. PMID:23176080

Major surfome and secretome profile of Streptococcus agalactiae from Nile tilapia (Oreochromis niloticus): Insight into vaccine development.

PubMed

Li, Wei; Wang, Hai-Qing; He, Run-Zhen; Li, Yan-Wei; Su, You-Lu; Li, An-Xing

2016-08-01

Streptococcus agalactiae is a major piscine pathogen that is responsible for huge economic losses to the aquaculture industry. Safe recombinant vaccines, based on a small number of antigenic proteins, are emerging as the most attractive, cost-effective solution against S. agalactiae. The proteins of S. agalactiae exposed to the environment, including surface proteins and secretory proteins, are important targets for the immune system and they are likely to be good vaccine candidates. To obtain a precise profile of its surface proteins, S. agalactiae strain THN0901, which was isolated from tilapia (Oreochromis niloticus), was treated with proteinase K to cleave surface-exposed proteins, which were identified by liquid chromatography-tandem spectrometry (LC-MS/MS). Forty surface-associated proteins were identified, including ten proteins containing cell wall-anchoring motifs, eight lipoproteins, eleven membrane proteins, seven secretory proteins, three cytoplasmic proteins, and one unknown protein. In addition, culture supernatant proteins of S. agalactiae were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and all of the Coomassie-stained bands were subsequently identified by LC-MS/MS. A total of twenty-six extracellular proteins were identified, including eleven secretory proteins, seven cell wall proteins, three membrane proteins, two cytoplasmic proteins and three unknown proteins. Of these, six highly expressed surface-associated and secretory proteins are putative to be vaccine candidate of piscine S. agalactiae. Moreover, immunogenic secreted protein, a highly expressed protein screened from the secretome in the present study, was demonstrated to induce high antibody titer in tilapia, and it conferred protection against S. agalactiae, as evidenced by the relative percent survival (RPS) 48.61± 8.45%. The data reported here narrow the scope of screening protective antigens, and provide guidance in the development of a novel vaccine against piscine S. agalactiae. Copyright © 2016 Elsevier Ltd. All rights reserved.

Entropic (de)stabilization of surface-bound peptides conjugated with polymers

NASA Astrophysics Data System (ADS)

Carmichael, Scott P.; Shell, M. Scott

2015-12-01

In many emerging biotechnologies, functional proteins must maintain their native structures on or near interfaces (e.g., tethered peptide arrays, protein coated nanoparticles, and amphiphilic peptide micelles). Because the presence of a surface is known to dramatically alter the thermostability of tethered proteins, strategies to stabilize surface-bound proteins are highly sought. Here, we show that polymer conjugation allows for significant control over the secondary structure and thermostability of a model surface-tethered peptide. We use molecular dynamics simulations to examine the folding behavior of a coarse-grained helical peptide that is conjugated to polymers of various lengths and at various conjugation sites. These polymer variations reveal surprisingly diverse behavior, with some stabilizing and some destabilizing the native helical fold. We show that ideal-chain polymer entropies explain these varied effects and can quantitatively predict shifts in folding temperature. We then develop a generic theoretical model, based on ideal-chain entropies, that predicts critical lengths for conjugated polymers to effect changes in the folding of a surface-bound protein. These results may inform new design strategies for the stabilization of surface-associated proteins important for a range technological applications.

Entropic (de)stabilization of surface-bound peptides conjugated with polymers.

PubMed

Carmichael, Scott P; Shell, M Scott

2015-12-28

In many emerging biotechnologies, functional proteins must maintain their native structures on or near interfaces (e.g., tethered peptide arrays, protein coated nanoparticles, and amphiphilic peptide micelles). Because the presence of a surface is known to dramatically alter the thermostability of tethered proteins, strategies to stabilize surface-bound proteins are highly sought. Here, we show that polymer conjugation allows for significant control over the secondary structure and thermostability of a model surface-tethered peptide. We use molecular dynamics simulations to examine the folding behavior of a coarse-grained helical peptide that is conjugated to polymers of various lengths and at various conjugation sites. These polymer variations reveal surprisingly diverse behavior, with some stabilizing and some destabilizing the native helical fold. We show that ideal-chain polymer entropies explain these varied effects and can quantitatively predict shifts in folding temperature. We then develop a generic theoretical model, based on ideal-chain entropies, that predicts critical lengths for conjugated polymers to effect changes in the folding of a surface-bound protein. These results may inform new design strategies for the stabilization of surface-associated proteins important for a range technological applications.

Extreme Evolutionary Conservation of Functionally Important Regions in H1N1 Influenza Proteome

PubMed Central

Warren, Samantha; Wan, Xiu-Feng; Conant, Gavin; Korkin, Dmitry

2013-01-01

The H1N1 subtype of influenza A virus has caused two of the four documented pandemics and is responsible for seasonal epidemic outbreaks, presenting a continuous threat to public health. Co-circulating antigenically divergent influenza strains significantly complicates vaccine development and use. Here, by combining evolutionary, structural, functional, and population information about the H1N1 proteome, we seek to answer two questions: (1) do residues on the protein surfaces evolve faster than the protein core residues consistently across all proteins that constitute the influenza proteome? and (2) in spite of the rapid evolution of surface residues in influenza proteins, are there any protein regions on the protein surface that do not evolve? To answer these questions, we first built phylogenetically-aware models of the patterns of surface and interior substitutions. Employing these models, we found a single coherent pattern of faster evolution on the protein surfaces that characterizes all influenza proteins. The pattern is consistent with the events of inter-species reassortment, the worldwide introduction of the flu vaccine in the early 80’s, as well as the differences caused by the geographic origins of the virus. Next, we developed an automated computational pipeline to comprehensively detect regions of the protein surface residues that were 100% conserved over multiple years and in multiple host species. We identified conserved regions on the surface of 10 influenza proteins spread across all avian, swine, and human strains; with the exception of a small group of isolated strains that affected the conservation of three proteins. Surprisingly, these regions were also unaffected by genetic variation in the pandemic 2009 H1N1 viral population data obtained from deep sequencing experiments. Finally, the conserved regions were intrinsically related to the intra-viral macromolecular interaction interfaces. Our study may provide further insights towards the identification of novel protein targets for influenza antivirals. PMID:24282564

Crystallisation via novel 3D nanotemplates as a tool for protein purification and bio-separation

NASA Astrophysics Data System (ADS)

Shah, Umang V.; Jahn, Niklas H.; Huang, Shanshan; Yang, Zhongqiang; Williams, Daryl R.; Heng, Jerry Y. Y.

2017-07-01

This study reports an experimental validation of the surface preferential nucleation of proteins on the basis of a relationship between nucleant pore diameter and protein hydrodynamic diameter. The validated correlation was employed for the selection of nucleant pore diameter to crystallise a target protein from binary, equivolume protein mixture. We report proof-of-concept preliminary experimental evidence for the rational approach for crystallisation of a target protein from a binary protein mixture on the surface of 3D nanotemplates with controlled surface porosity and narrow pore-size distribution selected on the basis of a relationship between the nucleant pore diameter and protein hydrodynamic diameter. The outcome of this study opens up an exciting opportunity for exploring protein crystallisation as a potential route for protein purification and bio-separation in both technical and pharmaceutical applications.

Saliva and Serum Protein Exchange at the Tooth Enamel Surface

PubMed Central

Heller, D.; Helmerhorst, E.J.; Oppenheim, F.G.

2016-01-01

The acquired enamel pellicle is an oral, fluid-derived protein layer that forms on the tooth surface. It is a biologically and clinically important integument that protects teeth against enamel demineralization, and abrasion. Tooth surfaces are exposed to different proteinaceous microenvironments depending on the enamel location. For instance, tooth surfaces close to the gingival sulcus contact serum proteins that emanate via this sulcus, which may impact pellicle composition locally. The aims of this study were to define the major salivary and serum components that adsorb to hydroxyapatite, to study competition among them, and to obtain preliminary evidence in an in vivo saliva/serum pellicle model. Hydroxyapatite powder was incubated with saliva and serum, and the proteins that adsorbed were identified by mass spectrometry. To study competition, saliva and serum proteins were labeled with CyDyes, mixed in various proportions, and incubated with hydroxyapatite. In vivo competition was assessed using a split-mouth design, with half the buccal tooth surfaces coated with serum and the other half with saliva. After exposure to the oral environment for 0 min, 30 min and 2 h, the pellicles were analyzed by SDS-PAGE. In pure saliva- or serum-derived pellicles, 82 and 84 proteins were identified, respectively. When present concomitantly, salivary protein adsorbers effectively competed with serum protein adsorbers for the hydroxyapatite surface. Specifically, acidic proline-rich protein, cystatin, statherin and protein S100-A9 proteins competed off apolipoproteins, complement C4-A, haptoglobin, transthyretin and serotransferrin. In vivo evidence further supported the replacement of serum proteins by salivary proteins. In conclusion, although significant numbers of serum proteins emanate from the gingival sulcus, their ability to participate in dental pellicle formation is likely reduced in the presence of strong salivary protein adsorbers. The functional properties of the acquired enamel pellicle will therefore be mostly dictated by the salivary component. PMID:27879420

Membrane-Protein Binding Measured with Solution-Phase Plasmonic Nanocube Sensors

PubMed Central

Wu, Hung-Jen; Henzie, Joel; Lin, Wan-Chen; Rhodes, Christopher; Li, Zhu; Sartorel, Elodie; Thorner, Jeremy; Yang, Peidong; Groves, Jay. T.

2013-01-01

We describe a solution-phase sensor of lipid-protein binding based on localized surface plasmon resonance (LSPR) of silver nanocubes. When silica-coated nanocubes are mixed into a suspension of lipid vesicles, supported membranes spontaneously assemble on their surfaces. Using a standard laboratory spectrophotometer, we calibrate the LSPR peak shift due to protein binding to the membrane surface and then characterize the lipid-binding specificity of a pleckstrin-homology domain protein. PMID:23085614

Probing the surface of a sweet protein: NMR study of MNEI with a paramagnetic probe

PubMed Central

Niccolai, Neri; Spadaccini, Roberta; Scarselli, Maria; Bernini, Andrea; Crescenzi, Orlando; Spiga, Ottavia; Ciutti, Arianna; Di Maro, Daniela; Bracci, Luisa; Dalvit, Claudio; Temussi, Piero A.

2001-01-01

The design of safe sweeteners is very important for people who are affected by diabetes, hyperlipemia, and caries and other diseases that are linked to the consumption of sugars. Sweet proteins, which are found in several tropical plants, are many times sweeter than sucrose on a molar basis. A good understanding of their structure–function relationship can complement traditional SAR studies on small molecular weight sweeteners and thus help in the design of safe sweeteners. However, there is virtually no sequence homology and very little structural similarity among known sweet proteins. Studies on mutants of monellin, the best characterized of sweet proteins, proved not decisive in the localization of the main interaction points of monellin with its receptor. Accordingly, we resorted to an unbiased approach to restrict the search of likely areas of interaction on the surface of a typical sweet protein. It has been recently shown that an accurate survey of the surface of proteins by appropriate paramagnetic probes may locate interaction points on protein surface. Here we report the survey of the surface of MNEI, a single chain monellin, by means of a paramagnetic probe, and a direct assessment of bound water based on an application of ePHOGSY, an NMR experiment that is ideally suited to detect interactions of small ligands to a protein. Detailed surface mapping reveals the presence, on the surface of MNEI, of interaction points that include residues previously predicted by ELISA tests and by mutagenesis. PMID:11468346

Proteomic Plasma Membrane Profiling Reveals an Essential Role for gp96 in the Cell Surface Expression of LDLR Family Members, Including the LDL Receptor and LRP6

PubMed Central

2012-01-01

The endoplasmic reticulum chaperone gp96 is required for the cell surface expression of a narrow range of proteins, including toll-like receptors (TLRs) and integrins. To identify a more comprehensive repertoire of proteins whose cell surface expression is dependent on gp96, we developed plasma membrane profiling (PMP), a technique that combines SILAC labeling with selective cell surface aminooxy-biotinylation. This approach allowed us to compare the relative abundance of plasma membrane (PM) proteins on gp96-deficient versus gp96-reconstituted murine pre-B cells. Analysis of unfractionated tryptic peptides initially identified 113 PM proteins, which extended to 706 PM proteins using peptide prefractionation. We confirmed a requirement for gp96 in the cell surface expression of certain TLRs and integrins and found a marked decrease in cell surface expression of four members of the extended LDL receptor family (LDLR, LRP6, Sorl1 and LRP8) in the absence of gp96. Other novel gp96 client proteins included CD180/Ly86, important in the B-cell response to lipopolysaccharide. We highlight common structural motifs in these client proteins that may be recognized by gp96, including the beta-propeller and leucine-rich repeat. This study therefore identifies the extended LDL receptor family as an important new family of proteins whose cell surface expression is regulated by gp96. PMID:22292497

Proteomic plasma membrane profiling reveals an essential role for gp96 in the cell surface expression of LDLR family members, including the LDL receptor and LRP6.

PubMed

Weekes, Michael P; Antrobus, Robin; Talbot, Suzanne; Hör, Simon; Simecek, Nikol; Smith, Duncan L; Bloor, Stuart; Randow, Felix; Lehner, Paul J

2012-03-02

The endoplasmic reticulum chaperone gp96 is required for the cell surface expression of a narrow range of proteins, including toll-like receptors (TLRs) and integrins. To identify a more comprehensive repertoire of proteins whose cell surface expression is dependent on gp96, we developed plasma membrane profiling (PMP), a technique that combines SILAC labeling with selective cell surface aminooxy-biotinylation. This approach allowed us to compare the relative abundance of plasma membrane (PM) proteins on gp96-deficient versus gp96-reconstituted murine pre-B cells. Analysis of unfractionated tryptic peptides initially identified 113 PM proteins, which extended to 706 PM proteins using peptide prefractionation. We confirmed a requirement for gp96 in the cell surface expression of certain TLRs and integrins and found a marked decrease in cell surface expression of four members of the extended LDL receptor family (LDLR, LRP6, Sorl1 and LRP8) in the absence of gp96. Other novel gp96 client proteins included CD180/Ly86, important in the B-cell response to lipopolysaccharide. We highlight common structural motifs in these client proteins that may be recognized by gp96, including the beta-propeller and leucine-rich repeat. This study therefore identifies the extended LDL receptor family as an important new family of proteins whose cell surface expression is regulated by gp96.

The RSV F and G glycoproteins interact to form a complex on the surface of infected cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Low, Kit-Wei; Tan, Timothy; Ng, Ken

2008-02-08

In this study, the interaction between the respiratory syncytial virus (RSV) fusion (F) protein, attachment (G) protein, and small hydrophobic (SH) proteins was examined. Immunoprecipitation analysis suggested that the F and G proteins exist as a protein complex on the surface of RSV-infected cells, and this conclusion was supported by ultracentrifugation analysis that demonstrated co-migration of surface-expressed F and G proteins. Although our analysis provided evidence for an interaction between the G and SH proteins, no evidence was obtained for a single protein complex involving all three of the virus proteins. These data suggest the existence of multiple virus glycoproteinmore » complexes within the RSV envelope. Although the stimulus that drives RSV-mediated membrane fusion is unknown, the association between the G and F proteins suggest an indirect role for the G protein in this process.« less

Membrane tension controls the assembly of curvature-generating proteins

PubMed Central

Simunovic, Mijo; Voth, Gregory A.

2015-01-01

Proteins containing a Bin/Amphiphysin/Rvs (BAR) domain regulate membrane curvature in the cell. Recent simulations have revealed that BAR proteins assemble into linear aggregates, strongly affecting membrane curvature and its in-plane stress profile. Here, we explore the opposite question: do mechanical properties of the membrane impact protein association? By using coarse-grained molecular dynamics simulations, we show that increased surface tension significantly impacts the dynamics of protein assembly. While tensionless membranes promote a rapid formation of long-living linear aggregates of N-BAR proteins, increase in tension alters the geometry of protein association. At high tension, protein interactions are strongly inhibited. Increasing surface density of proteins leads to a wider range of protein association geometries, promoting the formation of meshes, which can be broken apart with membrane tension. Our work indicates that surface tension may play a key role in recruiting proteins to membrane-remodelling sites in the cell. PMID:26008710

Steric Shielding of Surface Epitopes and Impaired Immune Recognition Induced by the Ebola Virus Glycoprotein

PubMed Central

Francica, Joseph R.; Varela-Rohena, Angel; Medvec, Andrew; Plesa, Gabriela; Riley, James L.; Bates, Paul

2010-01-01

Many viruses alter expression of proteins on the surface of infected cells including molecules important for immune recognition, such as the major histocompatibility complex (MHC) class I and II molecules. Virus-induced downregulation of surface proteins has been observed to occur by a variety of mechanisms including impaired transcription, blocks to synthesis, and increased turnover. Viral infection or transient expression of the Ebola virus (EBOV) glycoprotein (GP) was previously shown to result in loss of staining of various host cell surface proteins including MHC1 and β1 integrin; however, the mechanism responsible for this effect has not been delineated. In the present study we demonstrate that EBOV GP does not decrease surface levels of β1 integrin or MHC1, but rather impedes recognition by steric occlusion of these proteins on the cell surface. Furthermore, steric occlusion also occurs for epitopes on the EBOV glycoprotein itself. The occluded epitopes in host proteins and EBOV GP can be revealed by removal of the surface subunit of GP or by removal of surface N- and O- linked glycans, resulting in increased surface staining by flow cytometry. Importantly, expression of EBOV GP impairs CD8 T-cell recognition of MHC1 on antigen presenting cells. Glycan-mediated steric shielding of host cell surface proteins by EBOV GP represents a novel mechanism for a virus to affect host cell function, thereby escaping immune detection. PMID:20844579

Insulin adsorption on crystalline SiO2: Comparison between polar and nonpolar surfaces using accelerated molecular-dynamics simulations

NASA Astrophysics Data System (ADS)

Nejad, Marjan A.; Mücksch, Christian; Urbassek, Herbert M.

2017-02-01

Adsorption of insulin on polar and nonpolar surfaces of crystalline SiO2 (cristobalite and α -quartz) is studied using molecular dynamics simulation. Acceleration techniques are used in order to sample adsorption phase space efficiently and to identify realistic adsorption conformations. We find major differences between the polar and nonpolar surfaces. Electrostatic interactions govern the adsorption on polar surfaces and can be described by the alignment of the protein dipole with the surface dipole; hence spreading of the protein on the surface is irrelevant. On nonpolar surfaces, on the other hand, van-der-Waals interaction dominates, inducing surface spreading of the protein.

Imparting albumin-binding affinity to a human protein by mimicking the contact surface of a bacterial binding protein.

PubMed

Oshiro, Satoshi; Honda, Shinya

2014-04-18

Attachment of a bacterial albumin-binding protein module is an attractive strategy for extending the plasma residence time of protein therapeutics. However, a protein fused with such a bacterial module could induce unfavorable immune reactions. To address this, we designed an alternative binding protein by imparting albumin-binding affinity to a human protein using molecular surface grafting. The result was a series of human-derived 6 helix-bundle proteins, one of which specifically binds to human serum albumin (HSA) with adequate affinity (KD = 100 nM). The proteins were designed by transferring key binding residues of a bacterial albumin-binding module, Finegoldia magna protein G-related albumin-binding domain (GA) module, onto the human protein scaffold. Despite 13-15 mutations, the designed proteins maintain the original secondary structure by virtue of careful grafting based on structural informatics. Competitive binding assays and thermodynamic analyses of the best binders show that the binding mode resembles that of the GA module, suggesting that the contacting surface of the GA module is mimicked well on the designed protein. These results indicate that the designed protein may act as an alternative low-risk binding module to HSA. Furthermore, molecular surface grafting in combination with structural informatics is an effective approach for avoiding deleterious mutations on a target protein and for imparting the binding function of one protein onto another.

Prediction of the interaction site on the surface of an isolated protein structure by analysis of side chain energy scores.

PubMed

Liang, Shide; Zhang, Jian; Zhang, Shicui; Guo, Huarong

2004-11-15

We show that residues at the interfaces of protein-protein complexes have higher side-chain energy than other surface residues. Eight different sets of protein complexes were analyzed. For each protein pair, the complex structure was used to identify the interface residues in the unbound monomer structures. Side-chain energy was calculated for each surface residue in the unbound monomer using our previously developed scoring function.1 The mean energy was calculated for the interface residues and the other surface residues. In 15 of the 16 monomers, the mean energy of the interface residues was higher than that of other surface residues. By decomposing the scoring function, we found that the energy term of the buried surface area of non-hydrogen-bonded hydrophilic atoms is the most important factor contributing to the high energy of the interface regions. In spite of lacking hydrophilic residues, the interface regions were found to be rich in buried non-hydrogen-bonded hydrophilic atoms. Although the calculation results could be affected by the inaccuracy of the scoring function, patch analysis of side-chain energy on the surface of an isolated protein may be helpful in identifying the possible protein-protein interface. A patch was defined as 20 residues surrounding the central residue on the protein surface, and patch energy was calculated as the mean value of the side-chain energy of all residues in the patch. In 12 of the studied monomers, the patch with the highest energy overlaps with the observed interface. The results are more remarkable when only three residues with the highest energy in a patch are averaged to derive the patch energy. All three highest-energy residues of the top energy patch belong to interfacial residues in four of the eight small protomers. We also found that the residue with the highest energy score on the surface of a small protomer is very possibly the key interaction residue. (c) 2004 Wiley-Liss, Inc.

Interaction of β-sheet folds with a gold surface.

PubMed

Hoefling, Martin; Monti, Susanna; Corni, Stefano; Gottschalk, Kay Eberhard

2011-01-01

The adsorption of proteins on inorganic surfaces is of fundamental biological importance. Further, biomedical and nanotechnological applications increasingly use interfaces between inorganic material and polypeptides. Yet, the underlying adsorption mechanism of polypeptides on surfaces is not well understood and experimentally difficult to analyze. Therefore, we investigate here the interactions of polypeptides with a gold(111) surface using computational molecular dynamics (MD) simulations with a polarizable gold model in explicit water. Our focus in this paper is the investigation of the interaction of polypeptides with β-sheet folds. First, we concentrate on a β-sheet forming model peptide. Second, we investigate the interactions of two domains with high β-sheet content of the biologically important extracellular matrix protein fibronectin (FN). We find that adsorption occurs in a stepwise mechanism both for the model peptide and the protein. The positively charged amino acid Arg facilitates the initial contact formation between protein and gold surface. Our results suggest that an effective gold-binding surface patch is overall uncharged, but contains Arg for contact initiation. The polypeptides do not unfold on the gold surface within the simulation time. However, for the two FN domains, the relative domain-domain orientation changes. The observation of a very fast and strong adsorption indicates that in a biological matrix, no bare gold surfaces will be present. Hence, the bioactivity of gold surfaces (like bare gold nanoparticles) will critically depend on the history of particle administration and the proteins present during initial contact between gold and biological material. Further, gold particles may act as seeds for protein aggregation. Structural re-organization and protein aggregation are potentially of immunological importance.

Nano-functionalization of protein microspheres

NASA Astrophysics Data System (ADS)

Yoon, Sungkwon; Nichols, William T.

2014-08-01

Protein microspheres are promising building blocks for the assembly of complex functional materials. Here we demonstrate a set of three techniques that add functionality to the surface of protein microspheres. In the first technique, a positive surface charge on the protein spheres is deposited by electrostatic adsorption. Negatively charged silica and gold nanoparticle colloids can then electrostatically bind reversibly to the microsphere surface. In the second technique, nanoparticles are covalently anchored to the protein shell using a simple one-pot process. The strong covalent bond between sulfur groups in cysteine in the protein shell irreversibly binds to the gold nanoparticles. In the third technique, surface morphology of the protein microsphere is tuned through hydrodynamic instability at the water-oil interface. This is accomplished through the degree of solubility of the oil phase in water. Taken together these three techniques form a platform to create nano-functionalized protein microspheres, which can then be used as building blocks for the assembly of more complex macroscopic materials.

Patch Finder Plus (PFplus): a web server for extracting and displaying positive electrostatic patches on protein surfaces.

PubMed

Shazman, Shula; Celniker, Gershon; Haber, Omer; Glaser, Fabian; Mandel-Gutfreund, Yael

2007-07-01

Positively charged electrostatic patches on protein surfaces are usually indicative of nucleic acid binding interfaces. Interestingly, many proteins which are not involved in nucleic acid binding possess large positive patches on their surface as well. In some cases, the positive patches on the protein are related to other functional properties of the protein family. PatchFinderPlus (PFplus) http://pfp.technion.ac.il is a web-based tool for extracting and displaying continuous electrostatic positive patches on protein surfaces. The input required for PFplus is either a four letter PDB code or a protein coordinate file in PDB format, provided by the user. PFplus computes the continuum electrostatics potential and extracts the largest positive patch for each protein chain in the PDB file. The server provides an output file in PDB format including a list of the patch residues. In addition, the largest positive patch is displayed on the server by a graphical viewer (Jmol), using a simple color coding.

Patch Finder Plus (PFplus): A web server for extracting and displaying positive electrostatic patches on protein surfaces

PubMed Central

Shazman, Shula; Celniker, Gershon; Haber, Omer; Glaser, Fabian; Mandel-Gutfreund, Yael

2007-01-01

Positively charged electrostatic patches on protein surfaces are usually indicative of nucleic acid binding interfaces. Interestingly, many proteins which are not involved in nucleic acid binding possess large positive patches on their surface as well. In some cases, the positive patches on the protein are related to other functional properties of the protein family. PatchFinderPlus (PFplus) http://pfp.technion.ac.il is a web-based tool for extracting and displaying continuous electrostatic positive patches on protein surfaces. The input required for PFplus is either a four letter PDB code or a protein coordinate file in PDB format, provided by the user. PFplus computes the continuum electrostatics potential and extracts the largest positive patch for each protein chain in the PDB file. The server provides an output file in PDB format including a list of the patch residues. In addition, the largest positive patch is displayed on the server by a graphical viewer (Jmol), using a simple color coding. PMID:17537808

Surface Density of the Hendra G Protein Modulates Hendra F Protein-Promoted Membrane Fusion: Role for Hendra G Protein Trafficking and Degradation

PubMed Central

Whitman, Shannon D.; Dutch, Rebecca Ellis

2007-01-01

Hendra virus, like most paramyxoviruses, requires both a fusion (F) and attachment (G) protein for promotion of cell-cell fusion. Recent studies determined that Hendra F is proteolytically processed by the cellular protease cathepsin L after endocytosis. This unique cathepsin L processing results in a small percentage of Hendra F on the cell surface. To determine how the surface densities of the two Hendra glycoproteins affect fusion promotion, we performed experiments that varied the levels of glycoproteins expressed in transfected cells. Using two different fusion assays, we found a marked increase in fusion when expression of the Hendra G protein was increased, with a 1:1 molar ratio of Hendra F:G on the cell surface resulting in optimal membrane fusion. Our results also showed that Hendra G protein levels are modulated by both more rapid protein turnover and slower protein trafficking than is seen for Hendra F. PMID:17328935

Miscibility of binary monolayers at the air-water interface and interaction of protein with immobilized monolayers by surface plasmon resonance technique.

PubMed

Wang, Yuchun; Du, Xuezhong

2006-07-04

The miscibility and stability of the binary monolayers of zwitterionic dipalmitoylphosphatidylcholine (DPPC) and cationic dioctadecyldimethylammonium bromide (DOMA) at the air-water interface and the interaction of ferritin with the immobilized monolayers have been studied in detail using surface pressure-area isotherms and surface plasmon resonance technique, respectively. The surface pressure-area isotherms indicated that the binary monolayers of DPPC and DOMA at the air-water interface were miscible and more stable than the monolayers of the two individual components. The surface plasmon resonance studies indicated that ferritin binding to the immobilized monolayers was primarily driven by the electrostatic interaction and that the amount of adsorbed protein at saturation was closely related not only to the number of positive charges in the monolayers but also to the pattern of positive charges at a given mole fraction of DOMA. The protein adsorption kinetics was determined by the properties of the monolayers (i.e., the protein-monolayer interaction) and the structure of preadsorbed protein molecules (i.e., the protein-protein interaction).

IMMUNOBLOT ANALYSIS OF PROTEINS ASSOCIATED WITH SELF-ASSEMBLED MONOLAYER SURFACES OF DEFINED CHEMISTRIES

PubMed Central

Cornelius, Rena M.; Shankar, Sucharita P.; Brash, John L.; Babensee, Julia E.

2011-01-01

Intact and fragmented proteins, eluted from self assembled monolayer (SAM) surfaces of alkanethiols of different chemistries (-CH3, -OH, -COOH, -NH2 ), following exposure to human plasma (HP) or human serum (HS), were examined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting techniques. The SAM surfaces were incubated for 1 hour with 10% (v/v) sterile-filtered heat-inactivated (h.i.) HS or 1% (v/v) sterile-filtered h.i. HP preparations [both in phosphate buffered saline (PBS)]. Adsorbed proteins were eluted using 10% SDS/2.3% dithioerythritol for characterization of protein profiles. The type of incubating medium may be an important determinant of adsorbed protein profiles, since some variations were observed in eluates from filtered versus control unfiltered h.i. 10% HS or 1% HP. Albumin and apolipoprotein A1 were consistently detected in both filtered h.i 10% HS and 1% HP eluates from all SAM surfaces and from control tissue culture-treated polystyrene (TCPS). Interestingly, Factor H and Factor I, antithrombin, prothrombin, high molecular weight kininogen (HMWK) and IgG were present in eluates from OH, COOH and NH2 SAM surfaces and in eluates from TCPS, but not in eluates from CH3 SAM surfaces, following exposure to filtered h.i. 10% HS. These results suggest that CH3 SAM surfaces were the least pro-inflammatory of all SAM surfaces. Overall, similar trends were observed in the profiles of proteins eluted from surfaces exposed to filtered 10% HS or 1% HP. However the unique profiles of adsorbed proteins on different SAM surface chemistries may be related to their differential interactions with cells, including immune/inflammatory cells. PMID:21509932

Effects of tethering a multistate folding protein to a surface

NASA Astrophysics Data System (ADS)

Wei, Shuai; Knotts, Thomas A.

2011-05-01

Protein/surface interactions are important in a variety of fields and devices, yet fundamental understanding of the relevant phenomena remains fragmented due to resolution limitations of experimental techniques. Molecular simulation has provided useful answers, but such studies have focused on proteins that fold through a two-state process. This study uses simulation to show how surfaces can affect proteins which fold through a multistate process by investigating the folding mechanism of lysozyme (PDB ID: 7LZM). The results demonstrate that in the bulk 7LZM folds through a process with four stable states: the folded state, the unfolded state, and two stable intermediates. The folding mechanism remains the same when the protein is tethered to a surface at most residues; however, in one case the folding mechanism changes in such a way as to eliminate one of the intermediates. An analysis of the molecular configurations shows that tethering at this site is advantageous for protein arrays because the active site is both presented to the bulk phase and stabilized. Taken as a whole, the results offer hope that rational design of protein arrays is possible once the behavior of the protein on the surface is ascertained.

Chemical Patterning by Mechanical Removal of Aqueous Polymers

NASA Astrophysics Data System (ADS)

Barnett, Katherine; Knoebel, Jodi; Davis, Robert C.

2006-10-01

We are developing a new method for micro and nanoscale patterning of lipids and proteins on solid surfaces. A layer of polyethylene glycol (PEG) teminated polyallyl amine (PAA) was initially applied to a mica surface. The PEG surface is a low adhesion surface for proteins. Following polymer deposition an Atomic Force Microscope (AFM) tip was used to remove the polymer layer in desired regions. AFM imaging of the surface after mechanical polymer removal shows squares of exposed MICA surrounded by the PEG surface. The clean mica regions are now available for specific adsorption of lipid or protein layers.

Discriminatory bio-adhesion over nano-patterned polymer brushes

NASA Astrophysics Data System (ADS)

Gon, Saugata

Surfaces functionalized with bio-molecular targeting agents are conventionally used for highly-specific protein and cell adhesion. This thesis explores an alternative approach: Small non-biological adhesive elements are placed on a surface randomly, with the rest of the surface rendered repulsive towards biomolecules and cells. While the adhesive elements themselves, for instance in solution, typically exhibit no selectivity for various compounds within an analyte suspension, selective adhesion of targeted objects or molecules results from their placement on the repulsive surface. The mechanism of selectivity relies on recognition of length scales of the surface distribution of adhesive elements relative to species in the analyte solution, along with the competition between attractions and repulsions between various species in the suspension and different parts of the collecting surface. The resulting binding selectivity can be exquisitely sharp; however, complex mixtures generally require the use of multiple surfaces to isolate the various species: Different components will be adhered, sharply, with changes in collector composition. The key feature of these surface designs is their lack of reliance on biomolecular fragments for specificity, focusing entirely on physicochemical principles at the lengthscales from 1 - 100 nm. This, along with a lack of formal patterning, provides the advantages of simplicity and cost effectiveness. This PhD thesis demonstrates these principles using a system in which cationic poly-L-lysine (PLL) patches (10 nm) are deposited randomly on a silica substrate and the remaining surface is passivated with a bio-compatible PEG brush. TIRF microscopy revealed that the patches were randomly arranged, not clustered. By precisely controlling the number of patches per unit area, the interfaces provide sharp selectivity for adhesion of proteins and bacterial cells. For instance, it was found that a critical density of patches (on the order of 1000/mum 2) was required for fibrinogen adsorption while a greater density comprised the adhesion threshold for albumin. Surface compositions between these two thresholds discriminated binding of the two proteins. The binding behavior of the two proteins from a mixture was well anticipated by the single- protein binding behaviors of the individual proteins. The mechanism for protein capture was shown to be multivalent: protein adhesion always occurred for averages spacings of the adhesive patches smaller than the dimensions of the protein of interest. For some backfill brush architectures, the spacing between the patches at the threshold for protein capture clearly corresponded to the major dimension of the target protein. For more dense PEG brush backfills however, larger adhesion thresholds were observed, corresponding to greater numbers of patches involved with the adhesion of each protein molecule. . The thesis demonstrates the tuning of the position of the adhesion thresholds, using fibrinogen as a model protein, using variations in brush properties and ionic strength. The directions of the trends indicate that the brushes do indeed exert steric repulsions toward the proteins while the attractions are electrostatic in nature. The surfaces also demonstrated sharp adhesion thresholds for S. Aureus bacteria, at smaller concentrations of adhesive surfaces elements than those needed for the protein capture. The results suggest that bacteria may be captured while proteins are rejected from these surfaces, and there may be potential to discriminate different bacterial types. Such discrimination from protein-containing bacterial suspensions was investigated briefly in this thesis using S. Aureus and fibrinogen as a model mixture. However, due to binding of fibrinogen to the bacterial surface, the separation did not succeed. It is still expected, however, that these surfaces could be used to selectively capture bacteria in the presence of non-interacting proteins. The interaction of these brushes with two different cationic species PLL and lysozyme were studied. The thesis documents rapid and complete brush displacement by PLL, highlighting a major limitation of using such brushes in some applications. Also unanticipated, lysozyme, a small cationic protein, was found to adhere to the brushes in increasing amounts with the PEG content of the brush. This finding contradicts current understanding of protein-brush interactions that suggests increases in interfacial PEG content increase biocompatibility.

An Investigation of the Effects of Self-Assembled Monolayers on Protein Crystallisation

PubMed Central

Zhang, Chen-Yan; Shen, He-Fang; Wang, Qian-Jin; Guo, Yun-Zhu; He, Jin; Cao, Hui-Ling; Liu, Yong-Ming; Shang, Peng; Yin, Da-Chuan

2013-01-01

Most protein crystallisation begins from heterogeneous nucleation; in practice, crystallisation typically occurs in the presence of a solid surface in the solution. The solid surface provides a nucleation site such that the energy barrier for nucleation is lower on the surface than in the bulk solution. Different types of solid surfaces exhibit different surface energies, and the nucleation barriers depend on the characteristics of the solid surfaces. Therefore, treatment of the solid surface may alter the surface properties to increase the chance to obtain protein crystals. In this paper, we propose a method to modify the glass cover slip using a self-assembled monolayer (SAM) of functional groups (methyl, sulfydryl and amino), and we investigated the effect of each SAM on protein crystallisation. The results indicated that both crystallisation success rate in a reproducibility study, and crystallisation hits in a crystallisation screening study, were increased using the SAMs, among which, the methyl-modified SAM demonstrated the most significant improvement. These results illustrated that directly modifying the crystallisation plates or glass cover slips to create surfaces that favour heterogeneous nucleation can be potentially useful in practical protein crystallisation, and the utilisation of a SAM containing a functional group can be considered a promising technique for the treatment of the surfaces that will directly contact the crystallisation solution. PMID:23749116

Protein interactions with layers of TiO2 nanotube and nanopore arrays: Morphology and surface charge influence.

PubMed

Kulkarni, Mukta; Mazare, Anca; Park, Jung; Gongadze, Ekaterina; Killian, Manuela Sonja; Kralj, Slavko; von der Mark, Klaus; Iglič, Aleš; Schmuki, Patrik

2016-11-01

In the present work we investigate the key factors involved in the interaction of small-sized charged proteins with TiO 2 nanostructures, i.e. albumin (negatively charged), histone (positively charged). We examine anodic nanotubes with specific morphology (simultaneous control over diameter and length, e.g. diameter - 15, 50 or 100nm, length - 250nm up to 10μm) and nanopores. The nanostructures surface area has a direct influence on the amount of bound protein, nonetheless the protein physical properties as electric charge and size (in relation to nanotopography and biomaterial's electric charge) are crucial too. The highest quantity of adsorbed protein is registered for histone, for 100nm diameter nanotubes (10μm length) while higher values are registered for 15nm diameter nanotubes when normalizing protein adsorption to nanostructures' surface unit area (evaluated from dye desorption measurements) - consistent with theoretical considerations. The proteins presence on the nanostructures is evaluated by XPS and ToF-SIMS; additionally, we qualitatively assess their presence along the nanostructures length by ToF-SIMS depth profiles, with decreasing concentration towards the bottom. Surface nanostructuring of titanium biomedical devices with TiO 2 nanotubes was shown to significantly influence the adhesion, proliferation and differentiation of mesenchymal stem cells (and other cells too). A high level of control over the nanoscale topography and over the surface area of such 1D nanostructures enables a direct influence on protein adhesion. Herein, we investigate and show how the nanostructure morphology (nanotube diameter and length) influences the interactions with small-sized charged proteins, using as model proteins bovine serum albumin (negatively charged) and histone (positively charged). We show that the protein charge strongly influences their adhesion to the TiO 2 nanostructures. Protein adhesion is quantified by ELISA measurements and determination of the nanostructures' total surface area. We use a quantitative surface charge model to describe charge interactions and obtain an increased magnitude of the surface charge density at the top edges of the nanotubes. In addition, we track the proteins presence on and inside the nanostructures. We believe that these aspects are crucial for applications where the incorporation of active molecules such as proteins, drugs, growth factors, etc., into nanotubes is desired. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Bacterial adhesion to protein-coated surfaces: An AFM and QCM-D study

NASA Astrophysics Data System (ADS)

Strauss, Joshua; Liu, Yatao; Camesano, Terri A.

2009-09-01

Bacterial adhesion to biomaterials, mineral surfaces, or other industrial surfaces is strongly controlled by the way bacteria interact with protein layers or organic matter and other biomolecules that coat the materials. Despite this knowledge, many studies of bacterial adhesion are performed under clean conditions, instead of in the presence of proteins or organic molecules. We chose fetal bovine serum (FBS) as a model protein, and prepared FBS films on quartz crystals. The thickness of the FBS layer was characterized using atomic force microscopy (AFM) imaging under liquid and quartz crystal microbalance with dissipation (QCM-D). Next, we characterized how the model biomaterial surface would interact with the nocosomial pathogen Staphylococcus epidermidis. An AFM probe was coated with S. epidermidis cells and used to probe a gold slide that had been coated with FBS or another protein, fibronectin (FN). These experiments show that AFM and QCM-D can be used in complementary ways to study the complex interactions between bacteria, proteins, and surfaces.

Boosting protein stability with the computational design of β-sheet surfaces.

PubMed

Kim, Doo Nam; Jacobs, Timothy M; Kuhlman, Brian

2016-03-01

β-sheets often have one face packed against the core of the protein and the other facing solvent. Mutational studies have indicated that the solvent-facing residues can contribute significantly to protein stability, and that the preferred amino acid at each sequence position is dependent on the precise structure of the protein backbone and the identity of the neighboring amino acids. This suggests that the most advantageous methods for designing β-sheet surfaces will be approaches that take into account the multiple energetic factors at play including side chain rotamer preferences, van der Waals forces, electrostatics, and desolvation effects. Here, we show that the protein design software Rosetta, which models these energetic factors, can be used to dramatically increase protein stability by optimizing interactions on the surfaces of small β-sheet proteins. Two design variants of the β-sandwich protein from tenascin were made with 7 and 14 mutations respectively on its β-sheet surfaces. These changes raised the thermal midpoint for unfolding from 45°C to 64°C and 74°C. Additionally, we tested an empirical approach based on increasing the number of potential salt bridges on the surfaces of the β-sheets. This was not a robust strategy for increasing stability, as three of the four variants tested were unfolded. © 2016 The Protein Society.

Interaction of gold and silver nanoparticles with human plasma: Analysis of protein corona reveals specific binding patterns.

PubMed

Lai, Wenjia; Wang, Qingsong; Li, Lumeng; Hu, Zhiyuan; Chen, Jiankui; Fang, Qiaojun

2017-04-01

Determining how nanomaterials interact with plasma will assist in understanding their effects on the biological system. This work presents a systematic study of the protein corona formed from human plasma on 20nm silver and gold nanoparticles with three different surface modifications, including positive and negative surface charges. The results show that all nanoparticles, even those with positive surface modifications, acquire negative charges after interacting with plasma. Approximately 300 proteins are identified on the coronas, while 99 are commonly found on each nanomaterial. The 20 most abundant proteins account for over 80% of the total proteins abundance. Remarkably, the surface charge and core of the nanoparticles, as well as the isoelectric point of the plasma proteins, are found to play significant roles in determining the nanoparticle coronas. Albumin and globulins are present at levels of less than 2% on these nanoparticle coronas. Fibrinogen, which presents in the plasma but not in the serum, preferably binds to negatively charged gold nanoparticles. These observations demonstrate the specific plasma protein binding pattern of silver and gold nanoparticles, as well as the importance of the surface charge and core in determining the protein corona compositions. The potential downstream biological impacts of the corona proteins were also investigated. Copyright © 2017 Elsevier B.V. All rights reserved.

Expression of bovine non-classical major histocompatibility complex class 1 proteins in mouse P815 and human K562 cells

USDA-ARS?s Scientific Manuscript database

Major histocompatibility complex class I (MHC-I) proteins can be expressed as cell surface or secreted proteins. To investigate whether bovine non-classical MHC-I proteins are expressed as cell surface or secreted proteins, and to assess the reactivity pattern of monoclonal antibodies with non-class...

Characterization of the bionano interface and mapping extrinsic interactions of the corona of nanomaterials

NASA Astrophysics Data System (ADS)

O'Connell, D. J.; Bombelli, F. Baldelli; Pitek, A. S.; Monopoli, M. P.; Cahill, D. J.; Dawson, K. A.

2015-09-01

Nanoparticles in physiological environments are known to selectively adsorb proteins and other biomolecules forming a tightly bound biomolecular `corona' on their surface. Where the exchange times of the proteins are sufficiently long, it is believed that the protein corona constitutes the particle identity in biological milieu. Here we show that proteins in the corona retain their functional characteristics and can specifically bind to cognate proteins on arrays of thousands of immobilised human proteins. The biological identity of the nanomaterial is seen to be specific to the blood plasma concentration in which they are exposed. We show that the resulting in situ nanoparticle interactome is dependent on the protein concentration in plasma, with the emergence of a small number of dominant protein-protein interactions. These interactions are those driven by proteins that are adsorbed onto the particle surface and whose binding epitopes are subsequently expressed or presented suitably on the particle surface. We suggest that, since specific tailored protein arrays for target systems and organs can be designed, their use may be an important element in an overall study of the biomolecular corona.Nanoparticles in physiological environments are known to selectively adsorb proteins and other biomolecules forming a tightly bound biomolecular `corona' on their surface. Where the exchange times of the proteins are sufficiently long, it is believed that the protein corona constitutes the particle identity in biological milieu. Here we show that proteins in the corona retain their functional characteristics and can specifically bind to cognate proteins on arrays of thousands of immobilised human proteins. The biological identity of the nanomaterial is seen to be specific to the blood plasma concentration in which they are exposed. We show that the resulting in situ nanoparticle interactome is dependent on the protein concentration in plasma, with the emergence of a small number of dominant protein-protein interactions. These interactions are those driven by proteins that are adsorbed onto the particle surface and whose binding epitopes are subsequently expressed or presented suitably on the particle surface. We suggest that, since specific tailored protein arrays for target systems and organs can be designed, their use may be an important element in an overall study of the biomolecular corona. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr01970b

Quantitative study of protein-protein interactions by quartz nanopipettes

NASA Astrophysics Data System (ADS)

Tiwari, Purushottam Babu; Astudillo, Luisana; Miksovska, Jaroslava; Wang, Xuewen; Li, Wenzhi; Darici, Yesim; He, Jin

2014-08-01

In this report, protein-modified quartz nanopipettes were used to quantitatively study protein-protein interactions in attoliter sensing volumes. As shown by numerical simulations, the ionic current through the conical-shaped nanopipette is very sensitive to the surface charge variation near the pore mouth. With the appropriate modification of negatively charged human neuroglobin (hNgb) onto the inner surface of a nanopipette, we were able to detect concentration-dependent current change when the hNgb-modified nanopipette tip was exposed to positively charged cytochrome c (Cyt c) with a series of concentrations in the bath solution. Such current change is due to the adsorption of Cyt c to the inner surface of the nanopipette through specific interactions with hNgb. In contrast, a smaller current change with weak concentration dependence was observed when Cyt c was replaced with lysozyme, which does not specifically bind to hNgb. The equilibrium dissociation constant (KD) for the Cyt c-hNgb complex formation was derived and the value matched very well with the result from surface plasmon resonance measurement. This is the first quantitative study of protein-protein interactions by a conical-shaped nanopore based on charge sensing. Our results demonstrate that nanopipettes can potentially be used as a label-free analytical tool to quantitatively characterize protein-protein interactions.In this report, protein-modified quartz nanopipettes were used to quantitatively study protein-protein interactions in attoliter sensing volumes. As shown by numerical simulations, the ionic current through the conical-shaped nanopipette is very sensitive to the surface charge variation near the pore mouth. With the appropriate modification of negatively charged human neuroglobin (hNgb) onto the inner surface of a nanopipette, we were able to detect concentration-dependent current change when the hNgb-modified nanopipette tip was exposed to positively charged cytochrome c (Cyt c) with a series of concentrations in the bath solution. Such current change is due to the adsorption of Cyt c to the inner surface of the nanopipette through specific interactions with hNgb. In contrast, a smaller current change with weak concentration dependence was observed when Cyt c was replaced with lysozyme, which does not specifically bind to hNgb. The equilibrium dissociation constant (KD) for the Cyt c-hNgb complex formation was derived and the value matched very well with the result from surface plasmon resonance measurement. This is the first quantitative study of protein-protein interactions by a conical-shaped nanopore based on charge sensing. Our results demonstrate that nanopipettes can potentially be used as a label-free analytical tool to quantitatively characterize protein-protein interactions. Electronic supplementary information (ESI) available: Determination of nanopipette diameter; surface modification scheme; numerical simulation; noise analysis; SPR experiments. See DOI: 10.1039/c4nr02964j

Generation and Surface Localization of Intact M Protein in Streptococcus pyogenes Are Dependent on sagA

PubMed Central

Biswas, Indranil; Germon, Pierre; McDade, Kathleen; Scott, June R.

2001-01-01

The M protein is an important surface-located virulence factor of Streptococcus pyogenes, the group A streptococcus (GAS). Expression of M protein is primarily controlled by Mga, a transcriptional activator protein. A recent report suggested that the sag locus, which includes nine genes necessary and sufficient for production of streptolysin S, another GAS virulence factor, is also needed for transcription of emm, encoding the M protein (Z. Li, D. D. Sledjeski, B. Kreikemeyer, A. Podbielski, and M. D. Boyle, J. Bacteriol. 181:6019–6027, 1999). To investigate this in more detail, we constructed an insertion-deletion mutation in sagA, the first gene in the sag locus, in the M6 strain JRS4. The resulting strain, JRS470, produced no detectable streptolysin S and showed a drastic reduction in cell surface-associated M protein, as measured by cell aggregation and Western blot analysis. However, transcription of the emm gene was unaffected by the sagA mutation. Detailed analysis with monoclonal antibodies and an antipeptide antibody showed that the M protein in the sagA mutant strain was truncated so that it lacks the C-repeat region and the C-terminal domain required for anchoring it to the cell surface. This truncated M protein was largely found, as expected, in the culture supernatant. Lack of surface-located M protein made the sagA mutant strain susceptible to phagocytosis. Thus, although sagA does not affect transcription of the M6 protein gene, it is needed for the surface localization of this important virulence factor. PMID:11598078

Molecular dynamics simulations of the adsorption of bone morphogenetic protein-2 on surfaces with medical relevance.

PubMed

Utesch, Tillmann; Daminelli, Grazia; Mroginski, Maria Andrea

2011-11-01

Bone morphogenetic protein-2 (BMP-2) plays a crucial role in osteoblast differentiation and proliferation. Its effective therapeutic use for ectopic bone and cartilage regeneration depends, among other factors, on the interaction with the carrier at the implant site. In this study, we used classical molecular dynamics (MD) and a hybrid approach of steered molecular dynamics (SMD) combined with MD simulations to investigate the initial stages of the adsorption of BMP-2 when approaching two implant surfaces, hydrophobic graphite and hydrophilic titanium dioxide rutile. Surface adsorption was evaluated for six different orientations of the protein, two end-on and four side-on, in explicit water environment. On graphite, we observed a weak but stable adsorption. Depending on the initial orientation, hydrophobic patches as well as flexible loops of the protein were involved in the interaction with graphite. On the contrary, BMP-2 adsorbed only loosely to hydrophilic titanium dioxide. Despite a favorable interaction energy between protein and the TiO(2) surface, the rapid formation of a two-layer water structure prevented the direct interaction between protein and titanium dioxide. The first water adlayer had a strong repulsive effect on the protein, while the second attracted the protein toward the surface. For both surfaces, hydrophobic graphite and hydrophilic titanium dioxide, denaturation of BMP-2 induced by adsorption was not observed on the nanosecond time scale.

Interplay between grain structure and protein adsorption on functional response of osteoblasts: ultrafine-grained versus coarse-grained substrates.

PubMed

Misra, R D K; Nune, C; Pesacreta, T C; Somani, M C; Karjalainen, L P

2013-01-01

The rapid adsorption of proteins is the starting and primary biological response that occurs when a biomedical device is implanted in the physiological system. The biological response, however, depends on the surface characteristics of the device. Considering the significant interest in nano-/ultrafine surfaces and nanostructured coatings, we describe here, the interplay between grain structure and protein adsorption (bovine serum albumin: BSA) on osteoblasts functions by comparing nanograined/ultrafine-grained (NG/UFG) and coarse-grained (CG: grain size in the micrometer range) substrates by investigating cell-substrate interactions. The protein adsorption on NG/UFG surface was beneficial in favorably modulating biological functions including cell attachment, proliferation, and viability, whereas the effect was less pronounced on protein adsorbed CG surface. Additionally, immunofluorescence studies demonstrated stronger vinculin signals associated with actin stress fibers in the outer regions of the cells and cellular extensions on protein adsorbed NG/UFG surface. The functional response followed the sequence: NG/UFG(BSA) > NG/UFG > CG(BSA) > CG. The differences in the cellular response on bare and protein adsorbed NG/UFG and CG surfaces are attributed to cumulative contribution of grain structure and degree of hydrophilicity. The study underscores the potential advantages of protein adsorption on artificial biomedical devices to enhance the bioactivity and regulate biological functions. Copyright © 2012 Wiley Periodicals, Inc.

Prediction of protein orientation upon immobilization on biological and nonbiological surfaces

NASA Astrophysics Data System (ADS)

Talasaz, Amirali H.; Nemat-Gorgani, Mohsen; Liu, Yang; Ståhl, Patrik; Dutton, Robert W.; Ronaghi, Mostafa; Davis, Ronald W.

2006-10-01

We report on a rapid simulation method for predicting protein orientation on a surface based on electrostatic interactions. New methods for predicting protein immobilization are needed because of the increasing use of biosensors and protein microarrays, two technologies that use protein immobilization onto a solid support, and because the orientation of an immobilized protein is important for its function. The proposed simulation model is based on the premise that the protein interacts with the electric field generated by the surface, and this interaction defines the orientation of attachment. Results of this model are in agreement with experimental observations of immobilization of mitochondrial creatine kinase and type I hexokinase on biological membranes. The advantages of our method are that it can be applied to any protein with a known structure; it does not require modeling of the surface at atomic resolution and can be run relatively quickly on readily available computing resources. Finally, we also propose an orientation of membrane-bound cytochrome c, a protein for which the membrane orientation has not been unequivocally determined. electric double layer | electrostatic simulations | orientation flexibility

The establishment of Saccharomyces boulardii surface display system using a single expression vector.

PubMed

Wang, Tiantian; Sun, Hui; Zhang, Jie; Liu, Qing; Wang, Longjiang; Chen, Peipei; Wang, Fangkun; Li, Hongmei; Xiao, Yihong; Zhao, Xiaomin

2014-03-01

In the present study, an a-agglutinin-based Saccharomyces boulardii surface display system was successfully established using a single expression vector. Based on the two protein co-expression vector pSP-G1 built by Partow et al., a S. boulardii surface display vector-pSDSb containing all the display elements was constructed. The display results of heterologous proteins were confirmed by successfully displaying enhanced green fluorescent protein (EGFP) and chicken Eimeria tenella Microneme-2 proteins (EtMic2) on the S. boulardii cell surface. The DNA sequence of AGA1 gene from S. boulardii (SbAGA1) was determined and used as the cell wall anchor partner. This is the first time heterologous proteins have been displayed on the cell surface of S. boulardii. Because S. boulardii is probiotic and eukaryotic, its surface display system would be very valuable, particularly in the development of a live vaccine against various pathogenic organisms especially eukaryotic pathogens such as protistan parasites. Copyright © 2013 Elsevier Inc. All rights reserved.

Listeria monocytogenes biofilm-associated protein (BapL) may contribute to surface attachment of L. monocytogenes but is absent from many field isolates.

PubMed

Jordan, Suzanne J; Perni, Stefano; Glenn, Sarah; Fernandes, Isabel; Barbosa, Manuela; Sol, Manuela; Tenreiro, Rogerio P; Chambel, Lelia; Barata, Belarmino; Zilhao, Isabel; Aldsworth, Timothy G; Adriao, Andreia; Faleiro, M Leonor; Shama, Gilbert; Andrew, Peter W

2008-09-01

Listeria monocytogenes is a food-borne pathogen capable of adhering to a range of surfaces utilized within the food industry, including stainless steel. The factors required for the attachment of this ubiquitous organism to abiotic surfaces are still relatively unknown. In silico analysis of the L. monocytogenes EGD genome identified a putative cell wall-anchored protein (Lmo0435 [BapL]), which had similarity to proteins involved in biofilm formation by staphylococci. An insertion mutation was constructed in L. monocytogenes to determine the influence of this protein on attachment to abiotic surfaces. The results show that the protein may contribute to the surface adherence of strains that possess BapL, but it is not an essential requirement for all L. monocytogenes strains. Several BapL-negative field isolates demonstrated an ability to adhere to abiotic surfaces equivalent to that of BapL-positive strains. BapL is not required for the virulence of L. monocytogenes in mice.

Prediction of protein-protein interaction sites using electrostatic desolvation profiles.

PubMed

Fiorucci, Sébastien; Zacharias, Martin

2010-05-19

Protein-protein complex formation involves removal of water from the interface region. Surface regions with a small free energy penalty for water removal or desolvation may correspond to preferred interaction sites. A method to calculate the electrostatic free energy of placing a neutral low-dielectric probe at various protein surface positions has been designed and applied to characterize putative interaction sites. Based on solutions of the finite-difference Poisson equation, this method also includes long-range electrostatic contributions and the protein solvent boundary shape in contrast to accessible-surface-area-based solvation energies. Calculations on a large set of proteins indicate that in many cases (>90%), the known binding site overlaps with one of the six regions of lowest electrostatic desolvation penalty (overlap with the lowest desolvation region for 48% of proteins). Since the onset of electrostatic desolvation occurs even before direct protein-protein contact formation, it may help guide proteins toward the binding region in the final stage of complex formation. It is interesting that the probe desolvation properties associated with residue types were found to depend to some degree on whether the residue was outside of or part of a binding site. The probe desolvation penalty was on average smaller if the residue was part of a binding site compared to other surface locations. Applications to several antigen-antibody complexes demonstrated that the approach might be useful not only to predict protein interaction sites in general but to map potential antigenic epitopes on protein surfaces. Copyright (c) 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.

InterProSurf: a web server for predicting interacting sites on protein surfaces

PubMed Central

Negi, Surendra S.; Schein, Catherine H.; Oezguen, Numan; Power, Trevor D.; Braun, Werner

2009-01-01

Summary A new web server, InterProSurf, predicts interacting amino acid residues in proteins that are most likely to interact with other proteins, given the 3D structures of subunits of a protein complex. The prediction method is based on solvent accessible surface area of residues in the isolated subunits, a propensity scale for interface residues and a clustering algorithm to identify surface regions with residues of high interface propensities. Here we illustrate the application of InterProSurf to determine which areas of Bacillus anthracis toxins and measles virus hemagglutinin protein interact with their respective cell surface receptors. The computationally predicted regions overlap with those regions previously identified as interface regions by sequence analysis and mutagenesis experiments. PMID:17933856

Surface Sites for Engineering Allosteric Control in Proteins

PubMed Central

Lee, Jeeyeon; Natarajan, Madhusudan; Nashine, Vishal C.; Socolich, Michael; Vo, Tina; Russ, William P.; Benkovic, Stephen J.; Ranganathan, Rama

2010-01-01

Statistical analyses of protein families reveal networks of coevolving amino acids that functionally link distantly positioned functional surfaces. Such linkages suggest a concept for engineering allosteric control into proteins: The intramolecular networks of two proteins could be joined across their surface sites such that the activity of one protein might control the activity of the other. We tested this idea by creating PAS-DHFR, a designed chimeric protein that connects a light-sensing signaling domain from a plant member of the Per/Arnt/Sim (PAS) family of proteins with Escherichia coli dihydrofolate reductase (DHFR). With no optimization, PAS-DHFR exhibited light-dependent catalytic activity that depended on the site of connection and on known signaling mechanisms in both proteins. PAS-DHFR serves as a proof of concept for engineering regulatory activities into proteins through interface design at conserved allosteric sites. PMID:18927392

Identification of mycobacterial surface proteins released into subcellular compartments of infected macrophages.

PubMed

Beatty, W L; Russell, D G

2000-12-01

Considerable effort has focused on the identification of proteins secreted from Mycobacterium spp. that contribute to the development of protective immunity. Little is known, however, about the release of mycobacterial proteins from the bacterial phagosome and the potential role of these molecules in chronically infected macrophages. In the present study, the release of mycobacterial surface proteins from the bacterial phagosome into subcellular compartments of infected macrophages was analyzed. Mycobacterium bovis BCG was surface labeled with fluorescein-tagged succinimidyl ester, an amine-reactive probe. The fluorescein tag was then used as a marker for the release of bacterial proteins in infected macrophages. Fractionation studies revealed bacterial proteins within subcellular compartments distinct from mycobacteria and mycobacterial phagosomes. To identify these proteins, subcellular fractions free of bacteria were probed with mycobacterium-specific antibodies. The fibronectin attachment protein and proteins of the antigen 85-kDa complex were identified among the mycobacterial proteins released from the bacterial phagosome.

Inferring the microscopic surface energy of protein-protein interfaces from mutation data.

PubMed

Moal, Iain H; Dapkūnas, Justas; Fernández-Recio, Juan

2015-04-01

Mutations at protein-protein recognition sites alter binding strength by altering the chemical nature of the interacting surfaces. We present a simple surface energy model, parameterized with empirical ΔΔG values, yielding mean energies of -48 cal mol(-1) Å(-2) for interactions between hydrophobic surfaces, -51 to -80 cal mol(-1) Å(-2) for surfaces of complementary charge, and 66-83 cal mol(-1) Å(-2) for electrostatically repelling surfaces, relative to the aqueous phase. This places the mean energy of hydrophobic surface burial at -24 cal mol(-1) Å(-2) . Despite neglecting configurational entropy and intramolecular changes, the model correlates with empirical binding free energies of a functionally diverse set of rigid-body interactions (r = 0.66). When used to rerank docking poses, it can place near-native solutions in the top 10 for 37% of the complexes evaluated, and 82% in the top 100. The method shows that hydrophobic burial is the driving force for protein association, accounting for 50-95% of the cohesive energy. The model is available open-source from http://life.bsc.es/pid/web/surface_energy/ and via the CCharpPPI web server http://life.bsc.es/pid/ccharppi/. © 2015 Wiley Periodicals, Inc.

Fluorogenic Green-Inside Red-Outside (GIRO) Labeling Approach Reveals Adenylyl Cyclase-Dependent Control of BKα Surface Expression

PubMed Central

2015-01-01

The regulation of surface levels of protein is critical for proper cell function and influences properties including cell adhesion, ion channel contributions to current flux, and the sensitivity of surface receptors to ligands. Here we demonstrate a two-color labeling system in live cells using a single fluorogen activating peptide (FAP) based fusion tag, which enables the rapid and simultaneous quantification of surface and internal proteins. In the nervous system, BK channels can regulate neural excitability and neurotransmitter release, and the surface trafficking of BK channels can be modulated by signaling cascades and assembly with accessory proteins. Using this labeling approach, we examine the dynamics of BK channel surface expression in HEK293 cells. Surface pools of the pore-forming BKα subunit were stable, exhibiting a plasma membrane half-life of >10 h. Long-term activation of adenylyl cyclase by forskolin reduced BKα surface levels by 30%, an effect that could not be attributed to increased bulk endocytosis of plasma membrane proteins. This labeling approach is compatible with microscopic imaging and flow cytometry, providing a solid platform for examining protein trafficking in living cells. PMID:26301573

Electrostatic and dispersion interactions during protein adsorption on topographic nanostructures.

PubMed

Elter, Patrick; Lange, Regina; Beck, Ulrich

2011-07-19

Recently, biomaterials research has focused on developing functional implant surfaces with well-defined topographic nanostructures in order to influence protein adsorption and cellular behavior. To enhance our understanding of how proteins interact with such surfaces, we analyze the adsorption of lysozyme on an oppositely charged nanostructure using a computer simulation. We present an algorithm that combines simulated Brownian dynamics with numerical field calculation methods to predict the preferred adsorption sites for arbitrarily shaped substrates. Either proteins can be immobilized at their initial adsorption sites or surface diffusion can be considered. Interactions are analyzed on the basis of Derjaguin-Landau-Verway-Overbeek (DLVO) theory, including electrostatic and London dispersion forces, and numerical solutions are derived using the Poisson-Boltzmann and Hamaker equations. Our calculations show that for a grooved nanostructure (i.e., groove and plateau width 8 nm, height 4 nm), proteins first contact the substrate primarily near convex edges because of better geometric accessibility and increased electric field strengths. Subsequently, molecules migrate by surface diffusion into grooves and concave corners, where short-range dispersion interactions are maximized. In equilibrium, this mechanism leads to an increased surface protein concentration in the grooves, demonstrating that the total amount of protein per surface area can be increased if substrates have concave nanostructures.

Mechanisms of protein stabilization and prevention of protein aggregation by glycerol.

PubMed

Vagenende, Vincent; Yap, Miranda G S; Trout, Bernhardt L

2009-11-24

The stability of proteins in aqueous solution is routinely enhanced by cosolvents such as glycerol. Glycerol is known to shift the native protein ensemble to more compact states. Glycerol also inhibits protein aggregation during the refolding of many proteins. However, mechanistic insight into protein stabilization and prevention of protein aggregation by glycerol is still lacking. In this study, we derive mechanisms of glycerol-induced protein stabilization by combining the thermodynamic framework of preferential interactions with molecular-level insight into solvent-protein interactions gained from molecular simulations. Contrary to the common conception that preferential hydration of proteins in polyol/water mixtures is determined by the molecular size of the polyol and the surface area of the protein, we present evidence that preferential hydration of proteins in glycerol/water mixtures mainly originates from electrostatic interactions that induce orientations of glycerol molecules at the protein surface such that glycerol is further excluded. These interactions shift the native protein toward more compact conformations. Moreover, glycerol preferentially interacts with large patches of contiguous hydrophobicity where glycerol acts as an amphiphilic interface between the hydrophobic surface and the polar solvent. Accordingly, we propose that glycerol prevents protein aggregation by inhibiting protein unfolding and by stabilizing aggregation-prone intermediates through preferential interactions with hydrophobic surface regions that favor amphiphilic interface orientations of glycerol. These mechanisms agree well with experimental data available in the literature, and we discuss the extent to which these mechanisms apply to other cosolvents, including polyols, arginine, and urea.

The influence of surface treatment on hydrophobicity, protein adsorption and microbial colonisation of silicone hydrogel contact lenses.

PubMed

Santos, Lívia; Rodrigues, Diana; Lira, Madalena; Oliveira, M Elisabete C D Real; Oliveira, Rosário; Vilar, Eva Yebra-Pimentel; Azeredo, Joana

2007-07-01

To evaluate the influence of surface treatment of silicone-hydrogel CL on lens hydrophobicity, protein adsorption and microbial colonisation by studying several silicone hydrogel contact lenses (CL) with and without surface treatment. The lenses used in this study were Balafilcon A, Lotrafilcon A, Lotrafilcon B and Galyfilcon A. A conventional hydrogel CL (Etafilcon A) was also tested. Hydrophobicity was determined through contact angle measurement using the advancing type technique on air. The type and quantity of proteins adsorbed were assessed through SDS-PAGE and fluorescence spectroscopy, respectively. Microbial colonisation was studied by removing the microbes from the lenses through sonication, and counting the colony-forming units on agar plates. Regarding hydrophobicity, both surface and non-surface-treated silicone hydrogel CL were found to be hydrophobic, and the conventional hydrogel CL was found to be hydrophilic. Concerning protein adsorption, different protein profiles were observed on the several lenses tested. Nevertheless, the presence of proteins with the same molecular weight as lysozyme and lactoferrin was common to all lenses, which is probably related to their abundance in tears. In terms of total protein adsorption, silicone hydrogel CL did not exhibit any differences between themselves. However, the conventional hydrogel Etafilcon A adsorbed a larger amount of proteins. Regarding microbial colonisation, Balafilcon A exhibited the greatest amount of colonising microbes, which can be due to its superior hydrophobicity and higher electron acceptor capacity. This study suggests that silicone hydrogel lenses adsorb a lower amount of proteins than the conventional hydrogel lenses and that this phenomenon is independent of the presence of surface treatment. Concerning microbial colonisation, the surface treated Balafilcon A, exhibited a greater propensity, a fact that may compromise the lens wearer's ocular health.

Method for estimating protein binding capacity of polymeric systems.

PubMed

Sharma, Vaibhav; Blackwood, Keith A; Haddow, David; Hook, Lilian; Mason, Chris; Dye, Julian F; García-Gareta, Elena

2015-01-01

Composite biomaterials made from synthetic and protein-based polymers are extensively researched in tissue engineering. To successfully fabricate a protein-polymer composite, it is critical to understand how strongly the protein binds to the synthetic polymer, which occurs through protein adsorption. Currently, there is no cost-effective and simple method for characterizing this interfacial binding. To characterize this interfacial binding, we introduce a simple three-step method that involves: 1) synthetic polymer surface characterisation, 2) a quick, inexpensive and robust novel immuno-based assay that uses protein extraction compounds to characterize protein binding strength followed by 3) an in vitro 2D model of cell culture to confirm the results of the immuno-based assay. Fibrinogen, precursor of fibrin, was adsorbed (test protein) on three different polymeric surfaces: silicone, poly(acrylic acid)-coated silicone and poly(allylamine)-coated silicone. Polystyrene surface was used as a reference. Characterisation of the different surfaces revealed different chemistry and roughness. The novel immuno-based assay showed significantly stronger binding of fibrinogen to both poly(acrylic acid) and poly(allylamine) coated silicone. Finally, cell studies showed that the strength of the interaction between the protein and the polymer had an effect on cell growth. This novel immuno-based assay is a valuable tool in developing composite biomaterials of synthetic and protein-based polymers with the potential to be applied in other fields of research where protein adsorption onto surfaces plays an important role.

Polymer-Based Protein Engineering: Synthesis and Characterization of Armored, High Graft Density Polymer-Protein Conjugates.

PubMed

Carmali, Sheiliza; Murata, Hironobu; Cummings, Chad; Matyjaszewski, Krzysztof; Russell, Alan J

2017-01-01

Atom transfer radical polymerization (ATRP) from the surface of a protein can generate remarkably dense polymer shells that serve as armor and rationally tune protein function. Using straightforward chemistry, it is possible to covalently couple or display multiple small molecule initiators onto a protein surface. The chemistry is fine-tuned to be sequence specific (if one desires a single targeted site) at controlled density. Once the initiator is anchored on the protein surface, ATRP is used to grow polymers on protein surface, in situ. The technique is so powerful that a single-protein polymer conjugate molecule can contain more than 90% polymer coating by weight. If desired, stimuli-responsive polymers can be "grown" from the initiated sites to prepare enzyme conjugates that respond to external triggers such as temperature or pH, while still maintaining enzyme activity and stability. Herein, we focus mainly on the synthesis of chymotrypsin-polymer conjugates. Control of the number of covalently coupled initiator sites by changing the stoichiometric ratio between enzyme and the initiator during the synthesis of protein-initiator complexes allowed fine-tuning of the grafting density. For example, very high grafting density chymotrypsin conjugates were prepared from protein-initiator complexes to grow the temperature-responsive polymers, poly(N-isopropylacrylamide), and poly[N,N'-dimethyl(methacryloyloxyethyl) ammonium propane sulfonate]. Controlled growth of polymers from protein surfaces enables one to predictably manipulate enzyme kinetics and stability without the need for molecular biology-dependent mutagenesis. © 2017 Elsevier Inc. All rights reserved.

Proteinaceous determinants of surface colonization in bacteria: bacterial adhesion and biofilm formation from a protein secretion perspective

PubMed Central

Chagnot, Caroline; Zorgani, Mohamed A.; Astruc, Thierry; Desvaux, Mickaël

2013-01-01

Bacterial colonization of biotic or abiotic surfaces results from two quite distinct physiological processes, namely bacterial adhesion and biofilm formation. Broadly speaking, a biofilm is defined as the sessile development of microbial cells. Biofilm formation arises following bacterial adhesion but not all single bacterial cells adhering reversibly or irreversibly engage inexorably into a sessile mode of growth. Among molecular determinants promoting bacterial colonization, surface proteins are the most functionally diverse active components. To be present on the bacterial cell surface, though, a protein must be secreted in the first place. Considering the close association of secreted proteins with their cognate secretion systems, the secretome (which refers both to the secretion systems and their protein substrates) is a key concept to apprehend the protein secretion and related physiological functions. The protein secretion systems are here considered in light of the differences in the cell-envelope architecture between diderm-LPS (archetypal Gram-negative), monoderm (archetypal Gram-positive) and diderm-mycolate (archetypal acid-fast) bacteria. Besides, their cognate secreted proteins engaged in the bacterial colonization process are regarded from single protein to supramolecular protein structure as well as the non-classical protein secretion. This state-of-the-art on the complement of the secretome (the secretion systems and their cognate effectors) involved in the surface colonization process in diderm-LPS and monoderm bacteria paves the way for future research directions in the field. PMID:24133488

Influence of dynamic flow conditions on adsorbed plasma protein corona and surface-induced thrombus generation on antifouling brushes.

PubMed

Yu, Kai; Andruschak, Paula; Yeh, Han Hung; Grecov, Dana; Kizhakkedathu, Jayachandran N

2018-06-01

The information regarding the nature of protein corona (and its changes) and cell binding on biomaterial surface under dynamic conditions is critical to dissect the mechanism of surface-induced thrombosis. In this manuscript, we investigated the nature of protein corona and blood cell binding in heparinized recalcified human plasma, platelet rich plasma and whole blood on three highly hydrophilic antifouling polymer brushes, (poly(N, N-dimethylacrylamide) (PDMA), poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC) and poly[N-(2-hydroxypropyl) methacrylamide] (PHPMA) using an in vitro blood loop model at comparable arterial and venous flow, and static conditions. A fluid dynamics model was used initially to better understand the resulting flow patterns in a vertical channel containing the substrates to arrive at the placement of the substrates within the blood loop. The protein binding on the brush modified substrates was determined using ellipsometry, fluorescence microscopy and the nature of the protein corona was investigated using mass spectrometry based proteomics. The flow elevated fouling on brush coated surface from blood. The extent of plasma protein adsorption and platelet adhesion onto PDMA brush was lower than other surfaces in both static and flow conditions. The profiles of adsorbed protein corona showed strong dependence on the test conditions (static vs. flow), and the chemistry of the polymer brushes. Specially, the PDMA brush under flow conditions was more enriched with coagulation proteins, complement proteins, vitronectin and fibronectin but was less enriched with serum albumin. Apolipoprotein B-100 and complement proteins were the most abundant proteins seen on PMPC and PHPMA surfaces under both flow and static conditions, respectively. Unlike PDMA brush, the flow conditions did not affect the composition of protein corona on PMPC and PHPMA brushes. The nature of the protein corona formed in flow conditions influenced the platelet and red blood cell binding. The dependence of shear stress on platelet adhesion from platelet rich plasma and whole blood highlights the contribution of red blood cells in enhancing platelet adhesion on the surface under high shear condition. Copyright © 2018 Elsevier Ltd. All rights reserved.

Adsorption induced enzyme denaturation: the role of protein surface in adsorption induced protein denaturation on allyl glycidyl ether (AGE)-ethylene glycol dimethacrylate (EGDM) copolymers.

PubMed

Thudi, Lahari; Jasti, Lakshmi S; Swarnalatha, Y; Fadnavis, Nitin W; Mulani, Khudbudin; Deokar, Sarika; Ponrathnam, Surendra

2012-02-01

The effects of protein size on adsorption and adsorption-induced denaturation of proteins on copolymers of allyl glycidyl ether (AGE)-ethylene glycol dimethacrylate (EGDM) have been studied. Different responses were observed for the amount of protein adsorbed and denatured on the polymer surface for different proteins (trypsin, alchol dehydrogenase from baker's yeast (YADH), glucose dehydrogenase (GDH) from Gluconobacter cerinus, and alkaline phosphates from calf intestinal mucosa (CIAP). Protein adsorption on the copolymer with 25% crosslink density (AGE-25) was dependent not only on the size of the protein but also on the presence of glycoside residues on the protein surface. Adsorption and denaturation of proteins follows the order YADH>trypsin>GDH>CIAP although the molecular weights of the proteins follow the order YADH>CIAP>GDH>trypsin. The lack of correlation between amount of adsorbed protein and its molecular weight was due to the presence of glycoside residues on CIAP and GDH which protect the enzyme surface from denaturation. Enzyme stabilities in aqueous solutions of 1-cyclohexyl-2-pyrrolidinone (CHP) correlate well with the trend in denaturation by the copolymer, strongly suggesting that hydrophobic interactions play a major role in protein binding and the mechanism of protein denaturation is similar to that for water-miscible organic solvents. Copyright © 2011 Elsevier B.V. All rights reserved.

Suppression of protein adsorption on a charged phospholipid polymer interface.

PubMed

Xu, Yan; Takai, Madoka; Ishihara, Kazuhiko

2009-02-09

High capability of a charged interface to suppress adsorption of both anionic and cationic proteins was reported. The interface was covalently constructed on quartz by modifying with an anionic phospholipid copolymer, poly(2-methacryloyloxyethyl phosphorylcholine (MPC)-co-n-butyl methacrylate (BMA)-co-potassium 3-methacryloyloxypropyl sulfonate (PMPS)-co-3-methacryloxypropyl trimethoxysilane (MPTMSi)) (PMBSSi). The PMBSSi interfaces were very hydrophilic and homogeneous and could function effectively for a long time even under long-term fluidic working conditions. The PMBSSi density on the interface, which was controllable by adjusting the PMBSSi concentration of the modification solution, affected the surface properties, including the surface contact angle, the surface roughness, and the surface zeta-potential. When a PMBSSi modification was applied, the adsorption of various proteins (isoelectric point varying from 1.0 to 11.0) on quartz was reduced to at least 87% in amount, despite the various electrical natures these proteins have. The protein adsorption behavior on the PMBSSi interface depended more on the PMBSSi density than on the surface charge. The PMBSSi modification had a stable impact on the surface, not only at the physiologic ionic strength, but also over a range of the ionic strength, suggesting that electrostatic interactions do not dominate the behavior of protein adsorption to the PMBSSi surface.

Model simulations of the adsorption of statherin to solid surfaces: Effects of surface charge and hydrophobicity

NASA Astrophysics Data System (ADS)

Skepö, M.

2008-11-01

The structural properties of the salivary protein statherin upon adsorption have been examined using a coarse-grained model and Monte Carlo simulation. A simple model system with focus on electrostatic interactions and short-ranged attractions among the uncharged amino acids has been used. To mimic hydrophobically modified surfaces, an extra short-ranged interaction was implemented between the amino acids and the surface. It has been shown that the adsorption and the thickness of the adsorbed layer are determined by (i) the affinity for the surface, i.e., denser layer with an extrashort-ranged potential, and (ii) the distribution of the charges along the chain. If all the amino acids have a high affinity for the surface, the protein adsorbs in a train conformation, if the surface is negatively charged the protein adsorbs in a tail-train conformation, whereas if the surface is positively charged the protein adsorbs in a loop conformation. The latter gives rise to a more confined adsorbed layer.

Mapping hydration dynamics and coupled water-protein fluctuations around a protein surface

NASA Astrophysics Data System (ADS)

Zhang, Luyuan; Wang, Lijuan; Kao, Ya-Ting; Qiu, Weihong; Yang, Yi; Okobiah, Oghaghare; Zhong, Dongping

2009-03-01

Elucidation of the molecular mechanism of water-protein interactions is critical to understanding many fundamental aspects of protein science, such as protein folding and misfolding and enzyme catalysis. We recently carried out a global mapping of protein-surface hydration dynamics around a globular α-helical protein apomyoglobin. The intrinsic optical probe tryptophan was employed to scan the protein surface one at a time by site-specific mutagenesis. With femtosecond resolution, we mapped out the dynamics of water-protein interactions with more than 20 mutants and for two states, native and molten globular. A robust bimodal distribution of time scales was observed, representing two types of water motions: local relaxation and protein-coupled fluctuations. The time scales show a strong correlation with the local protein structural rigidity and chemical identity. We also resolved two distinct contributions to the overall Stokes-shifts from the two time scales. These results are significant to understanding the role of hydration water on protein structural stability, dynamics and function.

Protein adsorption at charged surfaces: the role of electrostatic interactions and interfacial charge regulation.

PubMed

Hartvig, Rune A; van de Weert, Marco; Østergaard, Jesper; Jorgensen, Lene; Jensen, Henrik

2011-03-15

The understanding of protein adsorption at charged surfaces is important for a wide range of scientific disciplines including surface engineering, separation sciences and pharmaceutical sciences. Compared to chemical entities having a permanent charge, the adsorption of small ampholytes and proteins is more complicated as the pH near a charged surface can be significantly different from the value in bulk solution. In this work, we have developed a phenomenological adsorption model which takes into account the combined role of interfacial ion distribution, interfacial charge regulation of amino acids in the proximity of the surface, electroneutrality, and mass balance. The model is straightforward to apply to a given set of experimental conditions as most model parameters are obtained from bulk properties and therefore easy to estimate or are directly measurable. The model provides a detailed understanding of the importance of surface charge on adsorption and in particular of how changes in surface charge, concentration, and surface area may affect adsorption behavior. The model is successfully used to explain the experimental adsorption behavior of the two model proteins lysozyme and α-lactalbumin. It is demonstrated that it is possible to predict the pH and surface charge dependent adsorption behavior from experimental or theoretical estimates of a preferred orientation of a protein at a solid charged interface.

Generativity in Elderly Oblate Sisters of Providence

PubMed Central

Black, Helen K.; Hannum, Susan M.; Rubinstein, Robert L.; de Medeiros, Kate

2016-01-01

Purpose of the Study: We explored how generativity and well-being merged in a group of childless older women: African and Hispanic Roman Catholic Religious Sisters, linking two minority identity characteristics. Design and Methods: We qualitatively interviewed 8 Oblate Sisters of Providence (OSP), by providing a framework for examining the range of the women’s generativity—cultural spheres in which generativity is rooted and outlets for generativity. Results: Early negative experiences, such as fleeing despotism in Haiti and Cuba and racism within the Catholic Church, occurred alongside positive experiences—families who stressed education, and Caucasian Religious who taught children of color. This became a foundation for the Sister’s generative commitment. Implications: Findings highlight that research gains from a phenomenological understanding of how religious faith promotes generative cognitions and emotions. Findings also reveal that the experiences of a subculture in society—African-American elderly women religious—add to theories and definitions of generativity. PMID:25352535

Next Generation Launch Technology Program Lessons Learned

NASA Technical Reports Server (NTRS)

Cook, Stephen; Tyson, Richard

2005-01-01

In November 2002, NASA revised its Integrated Space Transportation Plan (ISTP) to evolve the Space Launch Initiative (SLI) to serve as a theme for two emerging programs. The first of these, the Orbital Space Plane (OSP), was intended to provide crew-escape and crew-transfer functions for the ISS. The second, the NGLT Program, developed technologies needed for safe, routine space access for scientific exploration, commerce, and national defense. The NGLT Program was comprised of 12 projects, ranging from fundamental high-temperature materials research to full-scale engine system developments (turbine and rocket) to scramjet flight test. The Program included technology advancement activities with a broad range of objectives, ultimate applications/timeframes, and technology maturity levels. An over-arching Systems Engineering and Analysis (SE&A) approach was employed to focus technology advancements according to a common set of requirements. Investments were categorized into three segments of technology maturation: propulsion technologies, launch systems technologies, and SE&A.

Molecular Typing of Borrelia burgdorferi

PubMed Central

Wang, Guiqing; Liveris, Dionysios; Mukherjee, Priyanka; Jungnick, Sabrina; Margos, Gabriele; Schwartz, Ira

2015-01-01

Borrelia burgdorferi sensu lato is a group of spirochetes belonging to the genus Borrelia in the family of Spirochaetaceae. The spirochete is transmitted between reservoirs and hosts by ticks of the family Ixodidae. Infection with B. burgdorferi in humans causes Lyme disease or Lyme borreliosis. Currently, 20 Lyme disease-associated Borrelia species and more than 20 relapsing fever-associated Borrelia species have been described. Identification and differentiation of different Borrelia species and strains is largely dependent on analyses of their genetic characteristics. A variety of molecular techniques have been described for Borrelia isolate speciation, molecular epidemiology, and pathogenicity studies. In this unit, we focus on three basic protocols, PCR-RFLP-based typing of the rrs-rrlA and rrfA-rrlB ribosomal spacer, ospC typing, and MLST. These protocols can be employed alone or in combination for characterization of B. burgdorferi isolates or directly on uncultivated organisms in ticks, mammalian host reservoirs, and human clinical specimens. PMID:25082003

The First Fundamental Theorem of Invariant Theory for the Orthosymplectic Supergroup

NASA Astrophysics Data System (ADS)

Lehrer, G. I.; Zhang, R. B.

2017-01-01

We give an elementary and explicit proof of the first fundamental theorem of invariant theory for the orthosymplectic supergroup by generalising the geometric method of Atiyah, Bott and Patodi to the supergroup context. We use methods from super-algebraic geometry to convert invariants of the orthosymplectic supergroup into invariants of the corresponding general linear supergroup on a different space. In this way, super Schur-Weyl-Brauer duality is established between the orthosymplectic supergroup of superdimension ( m|2 n) and the Brauer algebra with parameter m - 2 n. The result may be interpreted either in terms of the group scheme OSp( V) over C, where V is a finite dimensional super space, or as a statement about the orthosymplectic Lie supergroup over the infinite dimensional Grassmann algebra {Λ}. We take the latter point of view here, and also state a corresponding theorem for the orthosymplectic Lie superalgebra, which involves an extra invariant generator, the super-Pfaffian.

A Molecular Simulation Study of Antibody-Antigen Interactions on Surfaces for the Rational Design of Next-Generation Antibody Microarrays

NASA Astrophysics Data System (ADS)

Bush, Derek B.

Antibody microarrays constitute a next-generation sensing platform that has the potential to revolutionize the way that molecular detection is conducted in many scientific fields. Unfortunately, current technologies have not found mainstream use because of reliability problems that undermine trust in their results. Although several factors are involved, it is believed that undesirable protein interactions with the array surface are a fundamental source of problems where little detail about the molecular-level biophysics are known. A better understanding of antibody stability and antibody-antigen binding on the array surface is needed to improve microarray technology. Despite the availability of many laboratory methods for studying protein stability and binding, these methods either do not work when the protein is attached to a surface or they do not provide the atomistic structural information that is needed to better understand protein behavior on the surface. As a result, molecular simulation has emerged as the primary method for studying proteins on surfaces because it can provide metrics and views of atomistic structures and molecular motion. Using an advanced, coarse-grain, protein-surface model this study investigated how antibodies react to and function on different types of surfaces. Three topics were addressed: (1) the stability of individual antibodies on surfaces, (2) antibody binding to small antigens while on a surface, and (3) antibody binding to large antigens while on a surface. The results indicate that immobilizing antibodies or antibody fragments in an upright orientation on a hydrophilic surface can provide the molecules with thermal stability similar to their native aqueous stability, enhance antigen binding strength, and minimize the entropic cost of binding. Furthermore, the results indicate that it is more difficult for large antigens to approach the surface than small antigens, that multiple binding sites can aid antigen binding, and that antigen flexiblity simultaneously helps and hinders the binding process as it approaches the surface. The results provide hope that next-generation microarrays and other devices decorated with proteins can be improved through rational design.

Photo-assisted generation of phospholipid polymer substrates for regiospecific protein conjugation and control of cell adhesion.

PubMed

Tanaka, Masako; Iwasaki, Yasuhiko

2016-08-01

Novel photo-reactive phospholipid polymers were synthesized for use in the preparation of nonfouling surfaces with protein conjugation capacity. Poly[2-methacryloyloxyethyl phosphorylcholine (MPC)-ran-N-methacryloyl-(l)-tyrosinemethylester (MAT)] (P(MPC/MAT)) was synthesized by conventional radical polymerization, with the MAT units capable of being oxidized by 254nm UV irradiation. Because of this photo-oxidation, active species such as catechol and quinone were alternately generated in the copolymer. A silicon wafer was subjected to surface modification through spin coating of P(MPC/MAT) from an aqueous solution for use as a model substrate. The surface was then irradiated several times with UV light. The thickness of the polymer layers formed on the Si wafers was influenced by various parameters such as polymer concentration, UV irradiation time, and composition of the MAT units in P(MPC/MAT). Oxidized MAT units were advantageous not only for polymer adhesion to a solid surface but also for protein conjugation with the adhered polymers. The amount of protein immobilized on UV-irradiated P(MPC/MAT) was dependent on the composition of the MAT units in the polymer. Furthermore, it was confirmed that protein immobilization on the polymer occurred through the oxidized MAT units because the protein adsorption was significantly reduced upon blocking these units through pretreatment with glycine. Conjugation of regiospecific protein could also be achieved through the use of a photomask. In addition, nonspecific protein adsorption was reduced on the non-irradiated regions whose surface was covered with physisorbed P(MPC/MAT). Therefore, P(MPC/MAT) can be used in the preparation of nonfouling substrates, which enable micrometer-sized manipulation of proteins through photo-irradiation. Function of proteins immobilized on MPC copolymers was also confirmed by cell adhesion test. As such, photo-reactive MPC copolymers are suitable for performing controlled protein conjugation and preparing polymer-protein hybrid platforms for use in biomedical and diagnostic devices. Novel photo-reactive phospholipid polymers have been synthesized for immobilization on solid surfaces and regiospecific protein conjugation. Tyrosine residues embedded in 2-methacryloyloxyethyl phosphorylcholine (MPC) copolymers could be photo-oxidized, resulting in polymers able to form layers on a solid surface and conjugate with proteins. Moreover, nonspecific biofouling on the surface significantly reduced when the oxidized tyrosine units in the polymer layers were blocked. Upon UV irradiation through a photomask, the UV-exposed tyrosine units were selectively oxidized, forming the only specific regions in which protein conjugation could occur. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.