Sample records for model protein bovine

  1. Protein Crystal Bovine Insulin

    NASA Technical Reports Server (NTRS)

    1991-01-01

    The comparison of protein crystal, Bovine Insulin space-grown (left) and earth-grown (right). Facilitates the incorporation of glucose into cells. In diabetics, there is either a decrease in or complete lack of insulin, thereby leading to several harmful complications. Principal Investigator is Larry DeLucas.

  2. Original article Effects of bovine colostrum acid protein on bone

    E-print Network

    Boyer, Edmond

    Original article Effects of bovine colostrum acid protein on bone loss and hemobiochemistry indexes that bovine milk and its basic proteins, and bovine colostrums (BC) and their extracts have positive effects hazard on blood lipids of rats under present experimental condition. bovine colostrum / acid protein

  3. NMR structure of the bovine prion protein

    Microsoft Academic Search

    Francisco López García; Ralph Zahn; Roland Riek; Kurt Wüthrich

    2000-01-01

    The NMR structures of the recombinant 217-residue polypeptide chain of the mature bovine prion protein, bPrP(23-230), and a C-terminal fragment, bPrP(121-230), include a globular domain extending from residue 125 to residue 227, a short flexible chain end of residues 228-230, and an N-terminal flexibly disordered \\

  4. Structural analysis of bovine pancreatic thread protein.

    PubMed

    Cai, L; Harris, W R; Marshak, D R; Gross, J; Crabb, J W

    1990-10-01

    Pancreatic thread protein (PTP) forms double helical threads in the neutral pH range after purification, undergoing freely reversible, pH-dependent globule-fibril transformation. The purified bovine PTP consists on SDS gels of two carbohydrate-free polypeptide chains (Gross et al., 1985). Plasma desorption mass spectrometry and amino acid sequence analysis now confirm that bovine PTP contains two disulfide-bonded polypeptides, an A chain of 101 amino acid residues with a molecular weight of 11,073 and a B chain of 35 residues with a molecular weight of 3970. The intact protein exhibits a molecular weight of 15,036, agreeing greater than 99.9% with the molecular weight calculated from the sequence. The B chain sequence was determined by gas-phase Edman degradation of the intact polypeptide. The A chain sequence was determined from overlapping peptides generated by cleavage at lysyl, tryptophanyl, and aspartyl-prolyl residues. Based upon the bovine PTP cDNA structure, the two chains of the protein result from cleavage of a single polypeptide with removal of a dipeptide between the NH2-terminal A chain and COOH-terminal B chain. Comparison of bovine PTP with other proteins reveals significant structural relatedness with the single-chain homologues from human and rat pancreas and with the motif associated with Ca2(+)-dependent carbohydrate recognition domains. The physiological role of PTP has not yet been resolved. The protein is present in very high concentration in pancreatic secretion and it has been detected in brain lesions in Alzheimer's disease and Down syndrome and in regenerating rat pancreatic islets. The present results provide a firm protein base for ongoing molecular, physical-chemical, and structure-function studies of this unusual protein. PMID:2085387

  5. Sequence and structural implications of a bovine corneal keratan sulfate proteoglycan core protein. Protein 37B represents bovine lumican and proteins 37A and 25 are unique

    NASA Technical Reports Server (NTRS)

    Funderburgh, J. L.; Funderburgh, M. L.; Brown, S. J.; Vergnes, J. P.; Hassell, J. R.; Mann, M. M.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

    1993-01-01

    Amino acid sequence from tryptic peptides of three different bovine corneal keratan sulfate proteoglycan (KSPG) core proteins (designated 37A, 37B, and 25) showed similarities to the sequence of a chicken KSPG core protein lumican. Bovine lumican cDNA was isolated from a bovine corneal expression library by screening with chicken lumican cDNA. The bovine cDNA codes for a 342-amino acid protein, M(r) 38,712, containing amino acid sequences identified in the 37B KSPG core protein. The bovine lumican is 68% identical to chicken lumican, with an 83% identity excluding the N-terminal 40 amino acids. Location of 6 cysteine and 4 consensus N-glycosylation sites in the bovine sequence were identical to those in chicken lumican. Bovine lumican had about 50% identity to bovine fibromodulin and 20% identity to bovine decorin and biglycan. About two-thirds of the lumican protein consists of a series of 10 amino acid leucine-rich repeats that occur in regions of calculated high beta-hydrophobic moment, suggesting that the leucine-rich repeats contribute to beta-sheet formation in these proteins. Sequences obtained from 37A and 25 core proteins were absent in bovine lumican, thus predicting a unique primary structure and separate mRNA for each of the three bovine KSPG core proteins.

  6. TRANSLATION OF BOVINE LEUKEMIA VIRUS GENOME INFORMATION IN HETEROLOGOUS PROTEIN SYNTHESIZING SYSTEMS

    E-print Network

    Paris-Sud XI, Université de

    TRANSLATION OF BOVINE LEUKEMIA VIRUS GENOME INFORMATION IN HETEROLOGOUS PROTEIN SYNTHESIZING, probablement situé du côté 3' du génome viral. Introduction. Bovine leukemia is a lymphoproliferative disease whose etiological agent is a retrovirus called Bovine Leukemia Virus (BLV). The cha- racteristics

  7. Models of bovine babesiosis including juvenile cattle.

    PubMed

    Saad-Roy, C M; Shuai, Zhisheng; van den Driessche, P

    2015-03-01

    Bovine Babesiosis in cattle is caused by the transmission of protozoa of Babesia spp. by ticks as vectors. Juvenile cattle (<9 months of age) have resistance to Bovine Babesiosis, rarely show symptoms, and acquire immunity upon recovery. Susceptibility to the disease varies between breeds of cattle. Models of the dynamics of Bovine Babesiosis transmitted by the cattle tick that include these factors are formulated as systems of ordinary differential equations. Basic reproduction numbers are calculated, and it is proved that if these numbers are below the threshold value of one, then Bovine Babesiosis dies out. However, above the threshold number of one, the disease may approach an endemic state. In this case, control measures are suggested by determining target reproduction numbers. The percentage of a particular population (for example, the adult bovine population) needed to be controlled to eradicate the disease is evaluated numerically using Columbia data from the literature. PMID:25715822

  8. Do Bovine Lymphocytes Express a Peculiar Prion Protein?

    PubMed Central

    Mélot, France; Thielen, Caroline; Labiet, Thouraya; Eisher, Sabine; Jolois, Olivier; Heinen, Ernst; Antoine, Nadine

    2002-01-01

    The cellular prion protein (PrPc) is a glycolipid-anchored cell surface protein that usually exhibits three glycosylation states. Its post-translationally modified isoform, PrPsc, is involved in the pathogenesis of various transmissible spongiform encephalopathies (TSEs). In bovine species, BSE infectivity appears to be restricted to the central nervous system; few or no detectable infectivity is found in lymphoid tissues in contrast to scrapie or variant CJD. Since expression of PrPc is a prerequisite for prion replication, we have investigated PrPc expression by bovine immune cells. Lymphocytes from blood and five different lymph organs were isolated from the same animal to assess intra- and interindividual variability of PrPc expression, considering six individuals. As shown by flow cytometry, this expression is absent or weak on granulocytes but is measurable on monocytes, B and T cells from blood and lymph organs. The activation of the bovine cells produces an upregulation of PrPc. The results of our in vitro study of PrPc biosynthesis are consistent with previous studies in other species. Interestingly, western blotting experiments showed only one form of the protein, the diglycosylated band. We propose that the glycosylation state could explain the lack of infectivity of the bovine immune cells. PMID:15144021

  9. Bovine immunoglobulin protein isolates for the nutritional management of enteropathy.

    PubMed

    Petschow, Bryon W; Blikslager, Anthony T; Weaver, Eric M; Campbell, Joy M; Polo, Javier; Shaw, Audrey L; Burnett, Bruce P; Klein, Gerald L; Rhoads, J Marc

    2014-09-01

    The gastrointestinal tract is responsible for a multitude of digestive and immune functions which depend upon the balanced interaction of the intestinal microbiota, diet, gut barrier function, and mucosal immune response. Disruptions in one or more of these factors can lead to intestinal disorders or enteropathies which are characterized by intestinal inflammation, increased gut permeability, and reduced capacity to absorb nutrients. Enteropathy is frequently associated with human immunodeficiency virus (HIV) infection, inflammatory bowel disease, autoimmune enteropathy, radiation enteritis, and irritable bowel syndrome (IBS), where pathologic changes in the intestinal tract lead to abdominal discomfort, bloating, abnormal bowel function (e.g., diarrhea, urgency, constipation and malabsorption). Unfortunately, effective therapies for the management of enteropathy and restoring intestinal health are still not available. An accumulating body of preclinical studies has demonstrated that oral administration of plasma- or serum-derived protein concentrates containing high levels of immunoglobulins can improve weight, normalize gut barrier function, and reduce the severity of enteropathy in animal models. Recent studies in humans, using serum-derived bovine immunoglobulin/protein isolate, demonstrate that such protein preparations are safe and improve symptoms, nutritional status, and various biomarkers associated with enteropathy. Benefits have been shown in patients with HIV infection or diarrhea-predominant IBS. This review summarizes preclinical and clinical studies with plasma/serum protein concentrates and describes the effects on host nutrition, intestinal function, and markers of intestinal inflammation. It supports the concept that immunoglobulin-containing protein preparations may offer a new strategy for restoring functional homeostasis in the intestinal tract of patients with enteropathy. PMID:25206275

  10. Cellular Requirements for Bovine Immunodeficiency Virus Vif-Mediated Inactivation of Bovine APOBEC3 Proteins

    PubMed Central

    Zhang, Wenyan; Wang, Hong; Li, Zhaolong; Liu, Xin; Liu, Guanchen; Harris, Reuben S.

    2014-01-01

    ABSTRACT Human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) viral infectivity factor (Vif) form a CRL5 E3 ubiquitin ligase complex to suppress virus restriction by host APOBEC3 (A3) proteins. The primate lentiviral Vif complex is composed of the unique cofactor core binding factor ? (CBF-?) and canonical ligase components Cullin 5 (CUL5), Elongin B/C (ELOB/C), and RBX2. However, the mechanism by which the Vif protein of the related lentivirus bovine immunodeficiency virus (BIV) overcomes its host A3 proteins is less clear. In this study, we show that BIV Vif interacts with Cullin 2 (CUL2), ELOB/C, and RBX1, but not with CBF-? or CUL5, to form a CRL2 E3 ubiquitin ligase and degrade the restrictive bovine A3 proteins (A3Z2Z3 and A3Z3). RNA interference-mediated knockdown of ELOB or CUL2 inhibited BIV Vif-mediated degradation of these A3 proteins, whereas knockdown of CUL5 or CBF-? did not. BIV Vif with mutations in the BC box (Vif SLQ-AAA) or putative VHL box (Vif YI-AA), which cannot interact with ELOB/C or CUL2, respectively, lost the ability to counteract bovine A3 proteins. Moreover, CUL2 and UBE2M dominant negative mutants competitively inhibited the BIV Vif-mediated degradation mechanism. Thus, although the general strategy for inhibiting A3 proteins is conserved between HIV-1/SIV and BIV, the precise mechanisms can differ substantially, with only the HIV-1/SIV Vif proteins requiring CBF-? as a cofactor, HIV-1/SIV Vif using CUL5-RBX2, and BIV Vif using CUL2-RBX1. IMPORTANCE Primate lentivirus HIV-1 and SIV Vif proteins form a ubiquitin ligase complex to target host antiviral APOBEC3 proteins for degradation. However, the mechanism by which the nonprimate lentivirus BIV Vif inhibits bovine APOBEC3 proteins is unclear. In the present study, we determined the mechanism for BIV Vif-mediated degradation of bovine APOBEC3 proteins and found that it differs from the mechanism of HIV-1/SIV Vif by being CBF-? independent and requiring different ubiquitin ligase scaffolding proteins (CUL2-RBX1 instead of CUL5-RBX2). BIV Vif is the only known retroviral protein that can interact with CUL2. This information broadens our understanding of the distinct mechanisms by which the Vif proteins of different lentiviruses facilitate viral infection. This novel mechanism for assembly of the BIV Vif-APOBEC3 ubiquitin ligase complex advances our understanding of viral hijacking of host E3 ubiquitin ligases and illustrates the evolutionary flexibility of lentiviruses. PMID:25142583

  11. Bovine Immunoglobulin/Protein Isolate Binds Pro-Inflammatory Bacterial Compounds and Prevents Immune Activation in an Intestinal Co-Culture Model

    PubMed Central

    Detzel, Christopher J.; Horgan, Alan; Henderson, Abigail L.; Petschow, Bryon W.; Warner, Christopher D.; Maas, Kenneth J.; Weaver, Eric M.

    2015-01-01

    Intestinal barrier dysfunction is associated with chronic gastrointestinal tract inflammation and diseases such as IBD and IBS. Serum-derived bovine immunoglobulin/protein isolate (SBI) is a specially formulated protein preparation (>90%) for oral administration. The composition of SBI is greater than 60% immunoglobulin including contributions from IgG, IgA, and IgM. Immunoglobulin within the lumen of the gut has been recognized to have anti-inflammatory properties and is involved in maintaining gut homeostasis. The binding of common intestinal antigens (LPS and Lipid A) and the ligand Pam3CSK4, by IgG, IgA, and IgM in SBI was shown using a modified ELISA technique. Each of these antigens stimulated IL-8 and TNF-? cytokine production by THP-1 monocytes. Immune exclusion occurred as SBI (?50 mg/mL) bound free antigen in a dose dependent manner that inhibited cytokine production by THP-1 monocytes in response to 10 ng/mL LPS or 200 ng/mL Lipid A. Conversely, Pam3CSK4 stimulation of THP-1 monocytes was unaffected by SBI/antigen binding. A co-culture model of the intestinal epithelium consisted of a C2BBe1 monolayer separating an apical compartment from a basal compartment containing THP-1 monocytes. The C2BBe1 monolayer was permeabilized with dimethyl palmitoyl ammonio propanesulfonate (PPS) to simulate a damaged epithelial barrier. Results indicate that Pam3CSK4 was able to translocate across the PPS-damaged C2BBe1 monolayer. However, binding of Pam3CSK4 by immunoglobulins in SBI prevented Pam3CSK4 translocation across the damaged C2BBe1 barrier. These results demonstrated steric exclusion of antigen by SBI which prevented apical to basal translocation of antigen due to changes in the physical properties of Pam3CSK4, most likely as a result of immunoglobulin binding. This study demonstrates that immunoglobulins in SBI can reduce antigen-associated inflammation through immune and steric exclusion mechanisms and furthers the mechanistic understanding of how SBI might improve immune status and reduce inflammation in various intestinal disease states. PMID:25830826

  12. Crystallization of Proteins from Crude Bovine Rod Outer Segments?

    PubMed Central

    Baker, Bo Y.; Gulati, Sahil; Shi, Wuxian; Wang, Benlian; Stewart, Phoebe L.; Palczewski, Krzysztof

    2015-01-01

    Obtaining protein crystals suitable for X-ray diffraction studies comprises the greatest challenge in the determination of protein crystal structures, especially for membrane proteins and protein complexes. Although high purity has been broadly accepted as one of the most significant requirements for protein crystallization, a recent study of the Escherichia coli proteome showed that many proteins have an inherent propensity to crystallize and do not require a highly homogeneous sample (Totir et al., 2012). As exemplified by RPE65 (Kiser, Golczak, Lodowski, Chance, & Palczewski, 2009), there also are cases of mammalian proteins crystallized from less purified samples. To test whether this phenomenon can be applied more broadly to the study of proteins from higher organisms, we investigated the protein crystallization profile of bovine rod outer segment (ROS) crude extracts. Interestingly, multiple protein crystals readily formed from such extracts, some of them diffracting to high resolution that allowed structural determination. A total of seven proteins were crystallized, one of which was a membrane protein. Successful crystallization of proteins from heterogeneous ROS extracts demonstrates that many mammalian proteins also have an intrinsic propensity to crystallize from complex biological mixtures. By providing an alternative approach to heterologous expression to achieve crystallization, this strategy could be useful for proteins and complexes that are difficult to purify or obtain by recombinant techniques. PMID:25950977

  13. Fetal bovine bone cells synthesize bone-specific matrix proteins

    Microsoft Academic Search

    S. WILLIAM WHITSON; WILBUR HARRISON; MARY K. DUNLAP; DANIEL E. BOWERS; LARRY W. FISHER; PAMELA GEHRON ROBEY; JOHN D. TERMINE

    1984-01-01

    We isolatedcellsfrom both calvariaand the outercorticesoflong bones from 3- to 5-mo bovine fetuses. The cellswere identifiedas functionalosteoblastsby indirect immunofluorescence using antibodiesagainstthree bone-specific,noncollagenous matrix proteins(osteonectin,the bone proteoglycan,and the bone sialoprotein)and againsttype I Collagen.Inseparateexperiments,confluentculturesofthecellswere radiolabeledand shown to synthesizeand secreteosteonectin,the bone proteoglycanand the bone sialoproteinby imunoprecipitationand fluorographyofSIDSpolyacrylamidegels.Analysisofthe radiolabeled collagenssynthesizedby theculturesshowed thatthey produced predominantly (-94%) type I collagen,with smallamounts of types IIIand V collagens.

  14. Envelope proteins of bovine herpesvirus 1: immunological and biochemical studies

    SciTech Connect

    Rodriguez Roque, L.L.

    1986-01-01

    The authors studied immunological and biochemical properties of the bovid herpesvirus 1 (BHV-1) envelope proteins in order to understand the pathogenesis of BHV-1 infection and to provide basic information for the production of effective subunit vaccines against BHV-1. Ten glycoproteins MW 180, 150, 130, 115, 97, 77, 74, 64, 55, and 45 kilodaltons (K), and a single non-glycosylated 108 K protein were quantitatively removed from purified BHV-1 virions by detergent treatment. These glycoproteins were present on the virion envelope and on the surface of BHV-1 infected cells. The quantitative removal from virions by treatment with nonionic detergents and their presence on the surface of infected cells indicate that 180/97, 150/77, and 130/74/55 K are major components of the BHV-1 envelope and are also the targets of virus neutralizing humoral immune response. Envelope glycoproteins of herpes simplex type 1 (HSV-1) bind immunoglobulin by the Fc end and it is suggested this may increase pathogenicity of this virus. They searched for a similar function in BVH-1 by measuring the ability of BHV-1 infected cells and viral envelope proteins to bind radiolabelled rabbit and bovine IgG. Binding activity for rabbit IgG or bovine IgG-Fc could not be demonstrated by BHV-1 infected MDBK cells, whereas, MDBK cells infected with HSV-1 bound rabbit IgG and bovine IgG-Fc. None of the three major envelope proteins of BHV-1 bound to rabbit or bovine IgG. The results of this study indicate that BHV-1, unlike some other herpesviruses, lack Fc binding activity.

  15. Structural characterization of human and bovine lung surfactant protein D.

    PubMed Central

    Leth-Larsen, R; Holmskov, U; Højrup, P

    1999-01-01

    Human and bovine surfactant proteins D (SP-D) were purified from late amniotic fluid and bronchioalveolar lavage on the basis of its Ca(2+)-dependent affinity for maltose. The molecular mass of a trimeric subunit was determined by matrix-assisted laser desorption ionization MS to lie in the range 115-125 kDa for human SP-D and 110-123 kDa for bovine SP-D. A single polypeptide chain was determined at 37-41 and 36-40 kDa for the human and bovine species respectively. The major parts of the primary structures of both SP-D molecules were determined by a combination of MS and Edman degradation. The heterogeneity in SP-D was caused mainly by a high number of post-translational modifications in the collagen-like region. Proline and lysine residues were partly hydroxylated and lysine residues were further O-glycosylated with the disaccharide galactose-glucose. A partly occupied N-linked glycosylation site was characterized in human SP-D. The carbohydrate was determined as a complex type bi-antennary structure, with a small content of mono-antennary and tri-antennary structures. No sialic acid residues were present on the glycan, but some had an attached fucose and/or an N-acetylglucosamine residue linked to the core. Bovine SP-D was determined as having a similar structure. PMID:10527944

  16. Structural and functional characteristics of bovine milk protein glycosylation.

    PubMed

    O'Riordan, Noelle; Kane, Marian; Joshi, Lokesh; Hickey, Rita M

    2014-03-01

    Most secreted and cell membrane proteins in mammals are glycosylated. Many of these glycoproteins are also prevalent in milk and play key roles in the biomodulatory properties of milk and ultimately in determining milk's nutritional quality. Although a significant amount of information exists on the types and roles of free oligosaccharides in milk, very little is known about the glycans associated with milk glycoproteins, in particular, the biological properties that are linked to their presence. The main glycoproteins found in bovine milk are lactoferrin, the immunoglobulins, glycomacropeptide, a glycopeptide derived from ?-casein, and the glycoproteins of the milk fat globule membrane. Here, we review the glycoproteins present in bovine milk, the information currently available on their glycosylation and the biological significance of their oligosaccharide chains. PMID:24398766

  17. Fatty acid-binding protein in bovine skeletal muscle 

    E-print Network

    Moore, Kimberly Kirby

    1989-01-01

    the quality grade of carcasses, so it is important economically in terms of the marketing of beef. To select for increased intramuscular fat in beef cattle is thus a major target for producers. Bovine breeds are known to vary significantly in the amount... was eluted in each column. Five-ml fractions were collected at a flow rate of 0. 4 ml/min, and protein concentrations were determined by a biuret method (26). Selected fractions obtained from the G-100 column were transferred, for dialysis, to Spectra...

  18. Epitope mapping of bovine viral diarrhea virus nonstructural protein 3.

    PubMed

    Mahmoodi, Pezhman; Shapouri, Masoud Reza Seyfi Abad; Ghorbanpour, Masoud; Ekhtelat, Maryam; Hajikolaei, Mohammad Rahim Haji; Lotfi, Mohsen; Boroujeni, Mahdi Pourmahdi; Daghari, Maryam

    2014-10-15

    Six consecutive overlapped coding regions (F1-F6) of whole NS3 molecule of bovine viral diarrhea virus (BVDV) were cloned into pMAL-c2X plasmid vector and expressed in Escherichia coli cells (BL21 strain). The recombinant proteins were then purified by amylose resin to determine the most immunogenic domain(s) of the NS3 molecule. Evaluation of the recombinant proteins was carried out by indirect ELISAs using several bovine sera (previously characterized by virus neutralization test, a commercial ELISA kit, and a newly developed NS3-ELISA) and 6 monoclonal antibodies. The experiments showed that the most immunogenic domain of the NS3 protein was the fourth designed fragment (F4), a 122 amino-acid (AA) region of about 13.5 kDa (nucleotide 1003-1368; residue 335-456). Purified recombinant F4 was also evaluated as single ELISA antigen (F4-ELISA) for the detection of anti-BVDV antibodies in sera of infected cattle. Although this small recombinant fragment of NS3 protein was almost completely soluble and expressed more efficient respect to whole NS3 molecule, it did not show enough sensitivity and specificity to be a proper substitute for NS3 as ELISA antigen to detect specific antibodies against BVDV. However, statistical analyses showed a medium correlation between the results of the developed F4-ELISA and virus neutralization test (kappa coefficient=0.63, P<0.001), with the relative sensitivity and specificity of 78.05% and 84.91%, respectively, suggesting the potential use of this fragment as an ELISA antigen along with other antigens or monoclonal antibody(s) in a competitive ELISA. PMID:25205011

  19. Crystal structure of bovine coronavirus spike protein lectin domain.

    PubMed

    Peng, Guiqing; Xu, Liqing; Lin, Yi-Lun; Chen, Lang; Pasquarella, Joseph R; Holmes, Kathryn V; Li, Fang

    2012-12-01

    The spike protein N-terminal domains (NTDs) of bovine coronavirus (BCoV) and mouse hepatitis coronavirus (MHV) recognize sugar and protein receptors, respectively, despite their significant sequence homology. We recently determined the crystal structure of MHV NTD complexed with its protein receptor murine carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), which surprisingly revealed a human galectin (galactose-binding lectin) fold in MHV NTD. Here, we have determined at 1.55 ? resolution the crystal structure of BCoV NTD, which also has the human galectin fold. Using mutagenesis, we have located the sugar-binding site in BCoV NTD, which overlaps with the galactose-binding site in human galectins. Using a glycan array screen, we have identified 5-N-acetyl-9-O-acetylneuraminic acid as the preferred sugar substrate for BCoV NTD. Subtle structural differences between BCoV and MHV NTDs, primarily involving different conformations of receptor-binding loops, explain why BCoV NTD does not bind CEACAM1 and why MHV NTD does not bind sugar. These results suggest a successful viral evolution strategy in which coronaviruses stole a galectin from hosts, incorporated it into their spike protein, and evolved it into viral receptor-binding domains with altered sugar specificity in contemporary BCoV or novel protein specificity in contemporary MHV. PMID:23091051

  20. A COMPARISON OF HEAT-INDUCED GEL STRENGTHS OF BOVINE PLASMA AND EGG ALBUMEN PROTEINS 1'2

    Microsoft Academic Search

    D. W. Hickson; C. W. Dill; R. G. Morgan

    Summary The heat-induced gel strength in suspensions of two types of plasma protein isolates and egg albumen were compared to investigate the use of bovine blood plasma in food systems as a replacement for egg albumen and other proteins. A viscosity index, based on a counter-flow back-extrusion model, was used to measure gel strength. The optimum pH for gel formation

  1. Survivin protein expression in bovine follicular oocytes and their in vitro developmental competence

    Microsoft Academic Search

    Kilsoo Jeon; Eun Young Kim; Jin Cheol Tae; Chang Hyun Lee; Keum Sil Lee; Yeon Ok Kim; Dong Kee Jeong; Somi K. Cho; Jae Hoon Kim; Hyo Yeon Lee; Key Zung Riu; Ssang Goo Cho; Se Pill Park

    2008-01-01

    This study examined the relationship between survivin expression and the stage of development of in vitro cultured bovine oocytes and embryos; and whether survivin expression is affected by the quality of cumulus–oocyte complexes (COCS) or the quality of pre-implantation embryos. A polyclonal antibody was prepared using recombinant bovine survivin protein. Expression of survivin mRNA and protein was analyzed by real-time

  2. Mycobacterium avium Subspecies paratuberculosis Recombinant Proteins Modulate Antimycobacterial Functions of Bovine Macrophages

    PubMed Central

    Bannantine, John P.; Stabel, Judith R.; Laws, Elizabeth; D. Cardieri, Maria Clara; Souza, Cleverson D.

    2015-01-01

    It has been shown that Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) activates the Mitogen Activated Protein Kinase (MAPK) p38 pathway, yet it is unclear which components of M. paratuberculosis are involved in the process. Therefore, a set of 42 M. paratuberculosis recombinant proteins expressed from coding sequences annotated as lipoproteins were screened for their ability to induce IL-10 expression, an indicator of MAPKp38 activation, in bovine monocyte-derived macrophages. A recombinant lipoprotein, designated as MAP3837c, was among a group of 6 proteins that strongly induced IL-10 gene transcription in bovine macrophages, averaging a 3.1-fold increase compared to non-stimulated macrophages. However, a parallel increase in expression of IL-12 and TNF-? was only observed in macrophages exposed to a subset of these 6 proteins. Selected recombinant proteins were further analyzed for their ability to enhance survival of M. avium within bovine macrophages as measured by recovered viable bacteria and nitrite production. All 6 IL-10 inducing MAP recombinant proteins along with M. paratuberculosis cells significantly enhanced phosphorylation of MAPK-p38 in bovine macrophages. Although these proteins are likely not post translationally lipidated in E. coli and thus is a limitation in this study, these results form the foundation of how the protein component of the lipoprotein interacts with the immune system. Collectively, these data reveal M. paratuberculosis proteins that might play a role in MAPK-p38 pathway activation and hence in survival of this organism within bovine macrophages. PMID:26076028

  3. Proteomic Analysis of Bovine Pregnancy-specific Serum Proteins by 2D Fluorescence Difference Gel Electrophoresis

    PubMed Central

    Lee, Jae Eun; Lee, Jae Young; Kim, Hong Rye; Shin, Hyun Young; Lin, Tao; Jin, Dong Il

    2015-01-01

    Two dimensional-fluorescence difference gel electrophoresis (2D DIGE) is an emerging technique for comparative proteomics, which improves the reproducibility and reliability of differential protein expression analysis between samples. The purpose of this study was to investigate bovine pregnancy-specific proteins in the proteome between bovine pregnant and non-pregnant serum using DIGE technique. Serums of 2 pregnant Holstein dairy cattle at day 21 after artificial insemination and those of 2 non-pregnant were used in this study. The pre-electrophoretic labeling of pregnant and non-pregnant serum proteins were mixed with Cy3 and Cy5 fluorescent dyes, respectively, and an internal standard was labeled with Cy2. Labeled proteins with Cy2, Cy3, and Cy5 were separated together in a single gel, and then were detected by fluorescence image analyzer. The 2D DIGE method using fluorescence CyDye DIGE flour had higher sensitivity than conventional 2D gel electrophoresis, and showed reproducible results. Approximately 1,500 protein spots were detected by 2D DIGE. Several proteins showed a more than 1.5-fold up and down regulation between non-pregnant and pregnant serum proteins. The differentially expressed proteins were identified by MALDI-TOF mass spectrometer. A total 16 protein spots were detected to regulate differentially in the pregnant serum, among which 7 spots were up-regulated proteins such as conglutinin precursor, modified bovine fibrinogen and IgG1, and 6 spots were down-regulated proteins such as hemoglobin, complement component 3, bovine fibrinogen and IgG2a three spots were not identified. The identified proteins demonstrate that early pregnant bovine serum may have several pregnancy-specific proteins, and these could be a valuable information for the development of pregnancy-diagnostic markers in early pregnancy bovine serum. PMID:25925056

  4. Fatty acids bound to vitamin D-binding protein (DBP) from human and bovine sera.

    PubMed

    Ena, J M; Esteban, C; Pérez, M D; Uriel, J; Calvo, M

    1989-07-01

    Human and bovine vitamin D-binding protein (DBP) have been isolated from serum by a method that does not involve denaturing steps. This method includes Cibacron Blue-Sepharose chromatography, gel filtration, DEAE-Sephadex chromatography and albumin immunoadsorption. Analysis of fatty acids bound to the isolated human and bovine DBP showed molar ratios of fatty acid to protein of 0.4 and 1.3 respectively meanwhile human and bovine albumin have bound 1.8 and 1.5 moles per mol respectively. Most of fatty acids bound to human and bovine DBP are monounsaturated and saturated, mainly oleic and palmitic acids, which together account for 50% of the total of fatty acids in both species. By contrast, polyunsaturated fatty acids represented a minor component, less than 5%. PMID:2673244

  5. Production and Functional Analysis of Recombinant Bovine Morphogenic Protein 15 

    E-print Network

    Burns, Gregory Willis

    2013-11-21

    After 40 years of research, in vitro systems for mammalian embryo production produce lower quality embryos than those derived from in vivo sources. Recent reports have demonstrated that in vitro bovine oocyte maturation systems benefit from...

  6. Comparison of the principal proteins in bovine, caprine, buffalo, equine and camel milk.

    PubMed

    Hinz, Katharina; O'Connor, Paula M; Huppertz, Thom; Ross, R Paul; Kelly, Alan L

    2012-05-01

    Proteomic analysis of bovine, caprine, buffalo, equine and camel milk highlighted significant interspecies differences. Camel milk was found to be devoid of ?-lactoglobulin, whereas ?-lactoglobulin was the major whey protein in bovine, buffalo, caprine, and equine milk. Five different isoforms of ?-casein were found in camel milk, analogous to the micro-heterogeneity observed for bovine ?-casein. Several spots observed in 2D-electrophoretograms of milk of all species could tentatively be identified as polypeptides arising from the enzymatic hydrolysis of caseins. The understanding gained from the proteomic comparison of these milks may be of relevance both in terms of identifying sources of hypoallergenic alternatives to bovine milk and detection of adulteration of milk samples and products. PMID:22365180

  7. A protein with amino acid sequence homology to bovine insulin is present in the legume Vigna unguiculata (cowpea)

    Microsoft Academic Search

    T. M. Venâncio; A. E. A. Oliveira; L. B. Silva; O. L. T. Machado; K. V. S. Fernandes; J. Xavier-Filho

    2003-01-01

    Since the discovery of bovine insulin in plants, much effort has been devoted to the characterization of these proteins and elucidation of their functions. We report here the isolation of a protein with similar molecular mass and same amino acid sequence to bovine insulin from developing fruits of cowpea (Vigna unguiculata) genotype Epace 10. Insulin was measured by ELISA using

  8. Insulin regulates milk protein synthesis at multiple levels in the bovine mammary gland

    Microsoft Academic Search

    Karensa K. Menzies; Christophe Lefèvre; Keith L. Macmillan; Kevin R. Nicholas

    2009-01-01

    The role of insulin in milk protein synthesis is unresolved in the bovine mammary gland. This study examined the potential\\u000a role of insulin in the presence of two lactogenic hormones, hydrocortisone and prolactin, in milk protein synthesis. Insulin\\u000a was shown to stimulate milk protein gene expression, casein synthesis and 14C-lysine uptake in mammary explants from late pregnant cows. A global

  9. Is double C2 protein (DOC2) expressed in bovine adrenal medulla? A commercial anti-DOC2 monoclonal antibody recognizes a major bovine mitochondrial antigen.

    PubMed Central

    Duncan, R R; Apps, D K; Learmonth, M P; Shipston, M J; Chow, R H

    2000-01-01

    We have examined the expression in bovine adrenal medulla of double C2 protein (DOC2), a vesicular protein which associates with intracellular phospholipid and Ca(2+) and is implicated in the modulation of regulated exocytosis. Extensive reverse transcription-PCR, Northern blot analyses and in vitro translation reactions have been combined with immunological studies to provide data to suggest that neither DOC2alpha nor DOC2beta is expressed at detectable levels in bovine adrenal chromaffin cells, and that a widely used monoclonal antibody directed against DOC2 also recognizes mitochondrial complex III core protein 2. PMID:10998344

  10. Solubilization of proteins from bovine brain coated vesicles by protein perturbants and Triton X-100

    PubMed Central

    1985-01-01

    To identify integral and peripheral membrane proteins, highly purified coated vesicles from bovine brain were exposed to solutions of various pH, ionic strength, and concentrations of the nonionic detergent Triton X-100. At pH 10.0 or above most major proteins were liberated, but four minor polypeptides sedimented with the vesicles. From quantitative analysis of phospholipids in the pellet and extract, we determined that at a pH of up to 12 all phospholipids could be recovered in the pellet. Electron microscopic examination of coated vesicles at pH 12.0 showed all vesicles devoid of coat structures. Treatment with high ionic strength solutions (0-1.0 M KCl) at pH 6.5-8.5 also liberated all major proteins, except tubulin, which remained sedimentable. The addition of Triton X-100 to coated vesicles or to stripped vesicles from which 90% of the clathrin had been removed resulted in the release of four distinct polypeptides of approximate Mr 38,000, 29,000, 24,000 and 10,000. The 38,000-D polypeptide (pK approximately 5.0), which represents approximately 50% of the protein liberated by Triton X-100, appears to be a glycoprotein on the basis of its reaction with periodic acid-Schiff reagent. Extraction of 90% of the clathrin followed by extraction of 90% of the phospholipids with Triton X-100 produced a protein residue that remained sedimentable and consisted of structures that appeared to be shrunken stripped vesicles. Together our data indicate that most of the major polypeptides of brain coated vesicles behave as peripheral membrane proteins and at least four polypeptides behave as integral membrane proteins. By use of a monoclonal antibody, we have identified one of these polypeptides (38,000 mol wt) as a marker for a subpopulation of calf brain coated vesicles. PMID:2861205

  11. Protection of calves against cryptosporidiosis with immune bovine colostrum induced by a Cryptosporidium parvum recombinant protein

    Microsoft Academic Search

    Lance E Perryman; Sushila J Kapil; Michael L Jones; Elaine L Hunt

    1999-01-01

    The purpose of the study was to determine if immunization with a recombinant protein (rC7) of Cryptosporidium parvum would induce immune bovine colostrum that protected calves against cryptosporidiosis following oral challenge with C. parvum oocysts. Late gestation Holstein cows with low titers of antibody to the p23 antigen of C. parvum were immunized three times with 300 ?g affinity purified

  12. DETECTION OF A PRECURSOR OF BOVINE LEUKEMIA VIRUS STRUCTURAL PROTEINS IN PURIFIED VIRIONS

    E-print Network

    Boyer, Edmond

    DETECTION OF A PRECURSOR OF BOVINE LEUKEMIA VIRUS STRUCTURAL PROTEINS IN PURIFIED VIRIONS P. GUPTA J.F. FERRER Section of Viral Oncology, Comparative Leukemia Studies Unit, School of Veterinary similar to the C-type leukemia viruses of other species (Ferrer etal., 1971 ; Stock and Ferrer, 1972

  13. ~ristalsof bovine chyrnotrypsin. Enzymes are proteins specialized to catalyze biological reac-

    E-print Network

    Vallino, Joseph J.

    Figure 8-1 ~ristalsof bovine chyrnotrypsin. 1 Enzymes are proteins specialized to catalyze of biochemistry is the history of en- zyme research. The name enzyme ("in yeast") was not used until 1877- furic acid. Although Louis Pasteur recognized that fermentation is catalyzed by enzymes, he postulated

  14. The effect of bovine whey protein on ectopic bone formation in young growing rats

    Microsoft Academic Search

    Owen Kelly; Siobhan Cusack; Kevin D. Cashman

    2003-01-01

    The beneficial effect of bovine whey protein (WP) on bone metabolism has been shown in adult human subjects and ovariectomised rats. However, its effect on bone formation in earlier life, particularly during periods of bone mineral accrual, has not been investigated. Twenty-one male rats (4 weeks old, Wistar strain) were randomised by weight into three groups of seven rats each

  15. Localization of viral proteins in cells infected with bovine viral diarrhoea virus

    Microsoft Academic Search

    B. Grummer; M. Beer; E. Liebler-Tenorio; I. Greiser-Wilke

    Bovine viral diarrhoea virus (BVDV) is a member of the genus Pestivirus within the family Flaviviridae. In this report, protein localization studies were performed to assess the mechanism for the release of mature virus particles from infected cells. Since BVDV is an enveloped virus, budding from either intra- or extracellular membranes is feasible. A prerequisite for the latter mechanism is

  16. S-100 protein subunits in bovine oviduct epithelium: In situ distribution and changes during primary cell culture

    Microsoft Academic Search

    I. Walter; I. Miller

    1996-01-01

    Summary  Cultures of bovine oviduct epithelial cells are widely used in co-culture systems to improve the results ofin vitro fertilization. The aim of the present study was to evaluate the suitability of S-100 protein as a differentiation marker\\u000a for bovine oviduct epithelial cellsin vitro. The distribution of S-100? and S-100? was examined immunohistochemically in bovine oviduct epitheliumin situ and in primary

  17. Computational modelling of bovine ovarian follicle development

    PubMed Central

    2013-01-01

    Background The development of ovarian follicles hinges on the timely exposure to the appropriate combination of hormones. Follicle stimulating hormone (FSH) and luteinizing hormone (LH) are both produced in the pituitary gland and are transported via the blood circulation to the thecal layer surrounding the follicle. From there both hormones are transported into the follicle by diffusion. FSH-receptors are expressed mainly in the granulosa while LH-receptors are expressed in a gradient with highest expression in the theca. How this spatial organization is achieved is not known. Equally it is not understood whether LH and FSH trigger distinct signalling programs or whether the distinct spatial localization of their G-protein coupled receptors is sufficient to convey their distinct biological function. Results We have developed a data-based computational model of the spatio-temporal signalling processes within the follicle and (i) predict that FSH and LH form a gradient inside the follicle, (ii) show that the spatial distribution of FSH- and LH-receptors can arise from the well known regulatory interactions, and (iii) find that the differential activity of FSH and LH may well result from the distinct spatial localisation of their receptors, even when both receptors respond with the same intracellular signalling cascade to their ligand. Conclusion The model integrates the large amount of published data into a consistent framework that can now be used to better understand how observed defects translate into failed follicle maturation. PMID:23856357

  18. Immunologic comparison of the proteins of pseudorabies (Aujeszky's disease) virus and bovine herpesvirus-1.

    PubMed

    Bush, C E; Pritchett, R F

    1986-08-01

    The immunologic relationship between bovine herpesvirus-1 and pseudorabies virus was examined by 80% serum cross-neutralization test, enzyme-linked immunosorbent assays, and Western immunoblotting procedures. Immunogenic cross reactivity between the 2 viruses was observed with both the serum-neutralization test and the enzyme-linked immunosorbent assay. A probing of viral Western immunoblots with rabbit hyperimmune antiserum showed that there were a number of viral-specific cross-reactive proteins between bovine herpesvirus-1 and pseudorabies virus. PMID:3019188

  19. Analysis of bovine whey proteins in soybean dairy-like products by capillary electrophoresis

    Microsoft Academic Search

    C Garc??a-Ruiz; M Torre; M. L Marina

    1999-01-01

    The simultaneous separation of bovine whey proteins [?-lactalbumin and ?-lactoglobulin (A+B)] and soybean proteins was performed, for the first time, by capillary electrophoresis. Different experimental conditions were tested. The most suitable consisted of 0.050 M phosphate buffer (pH 8) with 1 M urea and 1.2 mg\\/ml methylhydroxyethylcellulose, UV detection at 280 nm, 15 kV applied voltage, and 30°C temperature. Quantitation

  20. Bovine Parvovirus DNA-binding Proteins: Identification by a Combined DNA Hybridization and Immunodetection Assay

    Microsoft Academic Search

    MURIEL LEDERMAN; BRUCE C. SHULL; ERNEST R. STOUT; ROBERT C. BATES

    1987-01-01

    SUMMARY We have investigated the interaction between bovine parvovirus (BPV) capsid and non-capsid proteins and restriction fragments of the BPV genome by a combined DNA hybridization and immunodetection assay. 3zp-labelled DNA was bound to nitrocellulose membranes bearing lysates of mock-infected and virus-infected cells whose proteins had been separated by SDS-polyacrylamide gel electrophoresis. The position of bound DNA was determined by

  1. A bovine model for evaluating efficacy of Pasteurella haemolytica biologics 

    E-print Network

    Harris, James Robert

    1989-01-01

    of MASTER OF SCIENCE December 1989 Major Subject: Veterinary Microbiology A BOVINE MODEL FOR EVALUATING EFFICACY OF PASTEURELLA HAEMOLYTICA BIOLOGICS A Thesis by James Robert Harris Approved as to style and content by: Russell B. Simpson (Chair... and clinical score TABLE 2. Clinical scoring system for calf pneumonia TABLE 3. Pasteurella ~haemol tl a mouse thalleage Page 16 17 20 TABLE 4. Hean clinical score and mean antibody response of calves 23 TABLE 5. 'Serum turbidity test 25 LIST...

  2. N- and o-glycosylation of a commercial bovine whey protein product.

    PubMed

    van Leeuwen, Sander S; Schoemaker, Ruud J W; Timmer, Christel J A M; Kamerling, Johannis P; Dijkhuizen, Lubbert

    2012-12-26

    Bovine whey protein products are used as a base ingredient in infant formulas to optimize the amino acid pattern to a more human-like composition. Although the protein composition of bovine milk has been studied in detail, glycosylation details of commercial whey protein products are missing. To this end, both the N- and O-glycans of such a protein concentrate were sequentially released, the N-glycans enzymatically and the O-glycans chemically (reducing and nonreducing conditions). For the structural analysis of the N- and O-glycans a combination of MALDI-TOF-MS, one-dimensional (1)H NMR spectroscopy, Wisteria floribunda agglutinin affinity chromatography, HPAEC-PAD profiling, and HPLC-FD profiling (2-aminobenzamide derivatives), together with exoglycosidase treatments, were used. A mixture of over 60 N-glycans and 10 O-glycans was characterized, giving a detailed insight into the glycosylation of a bovine whey protein product, Deminal 90, which is applied as an ingredient for infant formulas. PMID:23194161

  3. Theoretical modeling of the kinetics of fibrilar aggregation of bovine ?-lactoglobulin at pH 2

    NASA Astrophysics Data System (ADS)

    Arnaudov, Luben N.; de Vries, Renko

    2007-04-01

    The authors propose a kinetic model for the heat-induced fibrilar aggregation of bovine ?-lactoglobulin at pH 2.0. The model involves a nucleation step and a simple addition reaction for the growth of the fibrils, as well as a side reaction leading to the irreversible denaturation and inactivation of a part of the protein molecules. For the early stages of the aggregation reaction, the authors obtain an analytical solution of the model. In agreement with their experimental results, the model predicts a critical protein concentration below where almost no fibrils are formed. The model agrees well with their experimental data from in situ light scattering. By fitting the experimental data with the model, the authors obtain the ionic strength dependent kinetic rate constants for ?-lactoglobulin fibrilar aggregation and the size of the critical nucleus.

  4. Purification of homologous protein carboxyl methyltransferase isozymes from human and bovine erythrocytes

    SciTech Connect

    Gilbert, J.M.; Fowler, A.; Bleibaum, J.; Clarke, S.

    1988-07-12

    The authors have purified the two major isozymes of the L-isoaspartyl/D-aspartyl protein methyltransferase from both human and bovine erythrocytes. These four enzymes all have polypeptide molecular weights of approximately 26,500 and appear to be monomers in solution. Each of these enzymes cross-reacts with antibodies directed against protein carboxyl methyltransferase I from bovine brain. Their structures also appear to be similar when analyzed by dodecyl sulfate gel electrophoresis for the large fragments produced by digestion with Staphylococcus aureus protease V8 or when analyzed by high-performance liquid chromatography (HPLC) for tryptic peptides. The structural relatedness of these enzymes was confirmed by sequence analysis of a total of 433 residues in 32 tryptic fragments of the human erythrocyte isozymes I and II and of the bovine erythrocyte isozyme II. They found sequence identity or probable identity in 111 out of 112 residues when they compared the human isozymes I and II and identities in 127 out of 134 residues when the human and bovine isozymes II were compared. These results suggest that the erythrocyte isozymes from both organisms may have nearly identical structures and confirm the similarities in the function of these methyltransferases that have been previously demonstrated.

  5. Improved mass spectrometric characterization of protein glycosylation reveals unusual glycosylation of maize-derived bovine trypsin

    PubMed Central

    Zhang, Hao; Huang, Richard Y-C; Jalili, Pegah R.; Irungu, Janet W.; Nicol, Gordon R.; Ray, Kevin B.; Rohrs, Henry W.; Gross, Michael L.

    2010-01-01

    Although bottom-up proteomics using tryptic digests is widely used to locate posttranslational modifications (PTM) in proteins, there are cases where the protein has several potential modification sites within a tryptic fragment, and MS2 strategies fail to pinpoint the location. We report here a method using two proteolytic enzymes, trypsin and pepsin, in combination followed by tandem mass spectrometric analysis to provide fragments that allow one to locate the modification sites. We used this strategy to find a glycosylation site on bovine trypsin expressed in maize (TrypZean™). Several glycans are present, and all are attached to a nonconsensus N-glycosylation site on the protein. PMID:21077632

  6. Surface diffusion of interacting proteins. Effect of concentration on the lateral mobility of adsorbed bovine serum albumin.

    PubMed Central

    Tilton, R D; Gast, A P; Robertson, C R

    1990-01-01

    Surface diffusion of bovine serum albumin absorbed from aqueous solution to poly(methylmethacrylate) surfaces is significantly hindered by protein-protein lateral interactions. The long-time self diffusion coefficient measured by fluorescence recovery after pattern photobleaching decreases by approximately one order of magnitude as the surface area fraction occupied by protein increases from 0.10 to 0.69. Qualitative features of the surface concentration dependence of the self diffusion coefficient can be described by several recent models for lateral diffusion of interacting species. The mobile fraction is independent of the surface concentration, and both the self diffusion coefficient and the mobile fraction are constant between 15 min and 7 h of adsorption. PMID:2291948

  7. Cross-Reactivity Between the Soybean Protein P34 and Bovine Caseins

    PubMed Central

    Smaldini, Paola Lorena; Curciarello, Renata; Cauerhff, Ana; Fossati, Carlos Alberto

    2015-01-01

    Purpose Soy-based formulas are widely used as dairy substitutes to treat milk allergy patients. However, reactions to soy have been reported in a small proportion of patients with IgE-mediated milk allergies. The aim of this work was to explore whether P34, a mayor soybean allergen, is involved in this cross-reactivity. Methods In vitro recognition of P34 was evaluated by immunoblotting, competitive ELISA and basophil activation tests (BAT) using sera from allergic patients. In vivo cross-reactivity was examined using an IgE-mediated milk allergy mouse model. Results P34 was recognized by IgE antibodies from the sera of milk allergic patients, casein-specific monoclonal antibodies, and sera from milk-allergic mice. Spleen cells from sensitized mice incubated with milk, soy or P34 secreted IL-5 and IL-13, while IFN-? remained unchanged. In addition, the cutaneous test was positive with cow's milk proteins (CMP) and P34 in the milk allergy mouse model. Moreover, milk-sensitized mice developed immediate symptoms following sublingual exposure to P34. Conclusions Our results demonstrate that P34 shares epitopes with bovine casein, which is responsible for inducing hypersensitivity symptoms in milk allergic mice. This is the first report of the in vivo cross-allergenicity of P34. PMID:25553264

  8. Production and Functional Analysis of Recombinant Bovine Morphogenic Protein 15

    E-print Network

    Burns, Gregory Willis

    2013-11-21

    ovarian tissue. For improved detection and purification of the biologically active recombinant protein, a FLAG tag peptide sequence (Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys) was incorporated into the wild type BMP15 gene by PCR. This modified protein was cloned...

  9. Inhibition of the bovine papillomavirus E2 protein activity by peptide nucleic acid

    Microsoft Academic Search

    Reet Kurg; Ülo Langel; Mart Ustav

    2000-01-01

    The bovine papillomavirus type-1 E2 protein is the master regulator of the papillomavirus transcription and replication, the activity of which is regulated through sequence-specific DNA binding. Peptide nucleic acid (PNA) is a nucleic acid analogue, which associates with high affinity to complementary DNA, RNA or PNA, yielding in formation of stable complexes. The potential use of PNA as a sequence-specific

  10. Sequence homology of rat and human HCNP precursor proteins, bovine phosphatidylethanolamine-binding protein and rat 23-kDa protein associated with the opioid-binding protein

    Microsoft Academic Search

    Naoki Tohdoh; Shinichiro Tojo; Hideo Agui; Kosei Ojika

    1995-01-01

    The hippocampal cholinergic neurostimulating peptide (HCNP) enhances acetylcholine synthesis in rat medial septal tissues. We have cloned the cDNAs of the precursor proteins of rat and human HCNP and deduced their respective amino acid sequences. The HCNP sequences aligned at the N-terminal regions of their precursors. The deduced amino acid sequences showed homology with those of the bovine brain phosphatidylethanolamine-binding

  11. Measurement of protein synthesis and degradation in C 2 C 22 myoblasts using extracts of muscle from hormone treated bovine

    Microsoft Academic Search

    J. L. Montgomery; W. M. Harper; M. F. Miller; K. J. Morrow; J. R. Blanton

    2002-01-01

    A detailed methodology is described for determination of treatment effects on muscle cell protein synthesis and muscle cell protein degradation in a cell culture system. C2C22 mouse myoblasts were treated with growth media containing muscle extracts from bovine treated with different pharmaceutical agents. Radiolabeled amino acids were added to the growth media to determine treatment effects on protein synthesis and

  12. Association analysis of bovine bactericidal/permeability-increasing protein gene polymorphisms with somatic cell score in Holstein cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bactericidal/permeability-increasing (BPI) protein is expressed primarily in bovine neutrophils and epithelial cells and functions as a binding protein of bacterial lipopolysaccharide produced by Gram-negative bacteria. The protein is important in host defense against bacterial infections and may pl...

  13. A bovine model for polycystic ovary syndrome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polycystic ovary syndrome (PCOS) results in the greatest single cause of anovulatory infertility in reproductive age women (affecting 5-10%). Previously, research groups have created animal models utilizing non-human primates and sheep to better understand the mechanisms involved in PCOS. However, c...

  14. In vivo delivery of bovine viral diahorrea virus, E2 protein using hollow mesoporous silica nanoparticles

    NASA Astrophysics Data System (ADS)

    Mahony, D.; Cavallaro, A. S.; Mody, K. T.; Xiong, L.; Mahony, T. J.; Qiao, S. Z.; Mitter, N.

    2014-05-01

    Our work focuses on the application of mesoporous silica nanoparticles as a combined delivery vehicle and adjuvant for vaccine applications. Here we present results using the viral protein, E2, from bovine viral diarrhoea virus (BVDV). BVDV infection occurs in the target species of cattle and sheep herds worldwide and is therefore of economic importance. E2 is a major immunogenic determinant of BVDV and is an ideal candidate for the development of a subunit based nanovaccine using mesoporous silica nanoparticles. Hollow type mesoporous silica nanoparticles with surface amino functionalisation (termed HMSA) were characterised and assessed for adsorption and desorption of E2. A codon-optimised version of the E2 protein (termed Opti-E2) was produced in Escherichia coli. HMSA (120 nm) had an adsorption capacity of 80 ?g Opti-E2 per mg HMSA and once bound E2 did not dissociate from the HMSA. Immunisation studies in mice with a 20 ?g dose of E2 adsorbed to 250 ?g HMSA was compared to immunisation with Opti-E2 (50 ?g) together with the traditional adjuvant Quillaja saponaria Molina tree saponins (QuilA, 10 ?g). The humoral responses with the Opti-E2/HMSA nanovaccine although slightly lower than those obtained for the Opti-E2 + QuilA group demonstrated that HMSA particles are an effective adjuvant that stimulated E2-specific antibody responses. Importantly the cell-mediated immune responses were consistently high in all mice immunised with Opti-E2/HMSA nanovaccine formulation. Therefore we have shown the Opti-E2/HMSA nanoformulation acts as an excellent adjuvant that gives both T-helper 1 and T-helper 2 mediated responses in a small animal model. This study has provided proof-of-concept towards the development of an E2 subunit nanoparticle based vaccine.Our work focuses on the application of mesoporous silica nanoparticles as a combined delivery vehicle and adjuvant for vaccine applications. Here we present results using the viral protein, E2, from bovine viral diarrhoea virus (BVDV). BVDV infection occurs in the target species of cattle and sheep herds worldwide and is therefore of economic importance. E2 is a major immunogenic determinant of BVDV and is an ideal candidate for the development of a subunit based nanovaccine using mesoporous silica nanoparticles. Hollow type mesoporous silica nanoparticles with surface amino functionalisation (termed HMSA) were characterised and assessed for adsorption and desorption of E2. A codon-optimised version of the E2 protein (termed Opti-E2) was produced in Escherichia coli. HMSA (120 nm) had an adsorption capacity of 80 ?g Opti-E2 per mg HMSA and once bound E2 did not dissociate from the HMSA. Immunisation studies in mice with a 20 ?g dose of E2 adsorbed to 250 ?g HMSA was compared to immunisation with Opti-E2 (50 ?g) together with the traditional adjuvant Quillaja saponaria Molina tree saponins (QuilA, 10 ?g). The humoral responses with the Opti-E2/HMSA nanovaccine although slightly lower than those obtained for the Opti-E2 + QuilA group demonstrated that HMSA particles are an effective adjuvant that stimulated E2-specific antibody responses. Importantly the cell-mediated immune responses were consistently high in all mice immunised with Opti-E2/HMSA nanovaccine formulation. Therefore we have shown the Opti-E2/HMSA nanoformulation acts as an excellent adjuvant that gives both T-helper 1 and T-helper 2 mediated responses in a small animal model. This study has provided proof-of-concept towards the development of an E2 subunit nanoparticle based vaccine. Electronic supplementary information (ESI) available: Desorption studies of Opti-E2 bound HMSA. See DOI: 10.1039/c4nr01202j

  15. Isolation of genomic and cDNA clones encoding bovine poly(A) binding protein II.

    PubMed

    Nemeth, A; Krause, S; Blank, D; Jenny, A; Jenö, P; Lustig, A; Wahle, E

    1995-10-25

    cDNA clones for bovine poly(A) binding protein II (PAB II) were isolated. Their sequence predicts a protein of 32.8 kDa, revising earlier estimates of molecular mass. The protein contains one putative RNA-binding domain of the RNP type, an acidic N-terminal and a basic C-terminal domain. Analyses of authentic PAB II were in good agreement with all predictions from the cDNA sequence except that a number of arginine residues appeared to be post-translationally modified. Poly(A) binding protein II expressed in Escherichia coli was active in poly(A) binding and reconstitution of processive polyadenylation, including poly(A) tail length control. The cDNA clones showed a number of potential PAB II binding sites in the 3' untranslated sequence. Bovine poly(A)+RNA contained two mRNAs hybridizing to a PAB II-specific probe. Analysis of a genomic clone revealed six introns in the coding sequence. The revised molecular mass led to a demonstration of PAB II oligomer formation and a reinterpretation of earlier data concerning the protein's binding to poly(A). PMID:7479061

  16. Isolation of genomic and cDNA clones encoding bovine poly(A) binding protein II.

    PubMed Central

    Nemeth, A; Krause, S; Blank, D; Jenny, A; Jenö, P; Lustig, A; Wahle, E

    1995-01-01

    cDNA clones for bovine poly(A) binding protein II (PAB II) were isolated. Their sequence predicts a protein of 32.8 kDa, revising earlier estimates of molecular mass. The protein contains one putative RNA-binding domain of the RNP type, an acidic N-terminal and a basic C-terminal domain. Analyses of authentic PAB II were in good agreement with all predictions from the cDNA sequence except that a number of arginine residues appeared to be post-translationally modified. Poly(A) binding protein II expressed in Escherichia coli was active in poly(A) binding and reconstitution of processive polyadenylation, including poly(A) tail length control. The cDNA clones showed a number of potential PAB II binding sites in the 3' untranslated sequence. Bovine poly(A)+RNA contained two mRNAs hybridizing to a PAB II-specific probe. Analysis of a genomic clone revealed six introns in the coding sequence. The revised molecular mass led to a demonstration of PAB II oligomer formation and a reinterpretation of earlier data concerning the protein's binding to poly(A). Images PMID:7479061

  17. Epithelial and endothelial expression of the green fluorescent protein reporter gene under the control of bovine prion protein (PrP) gene regulatory sequences in transgenic mice

    NASA Astrophysics Data System (ADS)

    Lemaire-Vieille, Catherine; Schulze, Tobias; Podevin-Dimster, Valérie; Follet, Jérome; Bailly, Yannick; Blanquet-Grossard, Françoise; Decavel, Jean-Pierre; Heinen, Ernst; Cesbron, Jean-Yves

    2000-05-01

    The expression of the cellular form of the prion protein (PrPc) gene is required for prion replication and neuroinvasion in transmissible spongiform encephalopathies. The identification of the cell types expressing PrPc is necessary to understanding how the agent replicates and spreads from peripheral sites to the central nervous system. To determine the nature of the cell types expressing PrPc, a green fluorescent protein reporter gene was expressed in transgenic mice under the control of 6.9 kb of the bovine PrP gene regulatory sequences. It was shown that the bovine PrP gene is expressed as two populations of mRNA differing by alternative splicing of one 115-bp 5' untranslated exon in 17 different bovine tissues. The analysis of transgenic mice showed reporter gene expression in some cells that have been identified as expressing PrP, such as cerebellar Purkinje cells, lymphocytes, and keratinocytes. In addition, expression of green fluorescent protein was observed in the plexus of the enteric nervous system and in a restricted subset of cells not yet clearly identified as expressing PrP: the epithelial cells of the thymic medullary and the endothelial cells of both the mucosal capillaries of the intestine and the renal capillaries. These data provide valuable information on the distribution of PrPc at the cellular level and argue for roles of the epithelial and endothelial cells in the spread of infection from the periphery to the brain. Moreover, the transgenic mice described in this paper provide a model that will allow for the study of the transcriptional activity of the PrP gene promoter in response to scrapie infection.

  18. The bovine seminal plasma protein PDC109 extracts phosphorylcholine-containing lipids from the outer membrane leaflet

    Microsoft Academic Search

    Astrid Tannert; Anke Kurz; Karl-Rudolf Erlemann; Karin Müller; Andreas Herrmann; Jürgen Schiller; Edda Töpfer-Petersen; Puttaswamy Manjunath; Peter Müller

    2007-01-01

    The bovine seminal plasma protein PDC-109 modulates the maturation of bull sperm cells by removing lipids, mainly phosphatidylcholine\\u000a and cholesterol, from their cellular membrane. Here, we have characterized the process of extraction of endogenous phospholipids\\u000a and of their respective analogues. By measuring the PDC-109-mediated release of fluorescent phospholipid analogues from lipid\\u000a vesicles and from biological membranes (human erythrocytes, bovine epididymal

  19. Rapid Separation and Quantification of Major Caseins and Whey Proteins of Bovine Milk by Polyacrylamide Gel Electrophoresis

    Microsoft Academic Search

    K. F. Ng-Kwai-Hang; E. M. Kroeker

    1984-01-01

    A rapid polyacrylamide gel electro- phoretic method was developed for separating and quantifying major pro- teins in casein and whey protein fractions of bovine milk. For casein separation, best results were achieved by an 8% poly- acrylamide gel containing 4 M urea and a top layer of large pore sample gel; for whey protein the most_ satisfactory separation was with

  20. Sequence analysis of the turkey enteric coronavirus nucleocapsid and membrane protein genes: a close genomic relationship with bovine coronavirus

    Microsoft Academic Search

    Arnold Verbeek; Peter Tijssen

    1991-01-01

    The 3' end of the turkey coronavirus (TCV) genome and the gene encoding the nucleocapsid protein (N) were cloned and sequenced. The gene encoding the membrane protein (M) was obtained by cloning a polymerase chain reaction (PCR)-amplified fragment obtained using bovine coronavirus (BCV)-specific primers. Furthermore, five TCV DNA fragments, obtained by PCR on RNA from clinical specimens and corresponding to

  1. The major protein of bovine seminal plasma, PDC-109, is a molecular chaperone.

    PubMed

    Sankhala, Rajeshwer Singh; Swamy, Musti J

    2010-05-11

    The major protein of bovine seminal plasma, PDC-109, binds to choline phospholipids on the sperm plasma membrane and induces the efflux of cholesterol and choline phospholipids, which is an important step in sperm capacitation. The high abundance, polydisperse nature and reversibility of thermal unfolding of PDC-109 suggest significant similarities to chaperone-like proteins such as spectrin, alpha-crystallin, and alpha-synuclein. In the present study, biochemical and biophysical approaches were employed to investigate the chaperone-like activity of PDC-109. The effect of various stress factors such as high temperature, chemical denaturant (urea), and acidic pH on target proteins such as lactate dehydrogenase, alcohol dehydrogenase, and insulin were studied in both the presence and absence of PDC-109. The results obtained indicate that PDC-109 exhibits chaperone-like activity, as evidenced by its ability to suppress the nonspecific aggregation of target proteins and direct them into productive folding. Atomic force microscopic studies demonstrate that PDC-109 effectively prevents the fibrillation of insulin, which is of considerable significance since amyloidogenesis has been reported to be a serious problem during sperm maturation in certain species. Binding of phosphorylcholine or high ionic strength in the medium inhibited the chaperone-like activity of PDC-109, suggesting that most likely the aggregation state of the protein is important for the chaperone function. These observations show that PDC-109 functions as a molecular chaperone in vitro, suggesting that it may assist the proper folding of proteins involved in the bovine sperm capacitation pathway. To the best of our knowledge, this is the first study reporting chaperone-like activity of a seminal plasma protein. PMID:20377251

  2. Homology Modeling Study of Bovine ?-Calpain Inhibitor-Binding Domains

    PubMed Central

    Chai, Han-Ha; Lim, Dajeong; Lee, Seung-Hwan; Chai, Hee-Yeoul; Jung, Eunkyoung

    2014-01-01

    The activated mammalian CAPN-structures, the CAPN/CAST complex in particular, have become an invaluable target model using the structure-based virtual screening of drug candidates from the discovery phase to development for over-activated CAPN linked to several diseases, such as post-ischemic injury and cataract formation. The effect of Ca2+-binding to the enzyme is thought to include activation, as well as the dissociation, aggregation, and autolysis of small regular subunits. Unfortunately, the Ca2+-activated enzyme tends to aggregate when provided as a divalent ion at the high-concentration required for the protease crystallization. This is also makes it very difficult to crystallize the whole-length enzyme itself, as well as the enzyme-inhibitor complex. Several parameters that influence CAPN activity have been investigated to determine its roles in Ca2+-modulation, autoproteolysis, phosphorylation, and intracellular distribution and inhibition by its endogenous inhibitor CAST. CAST binds and inhibits CAPN via its CAPN-inhibitor domains (four repeating domains 1–4; CAST1–4) when CAPN is activated by Ca2+-binding. An important key to understanding CAPN1 inhibition by CAST is to determine how CAST interacts at the molecular level with CAPN1 to inhibit its protease activity. In this study, a 3D structure model of a CAPN1 bound bovine CAST4 complex was built by comparative modeling based on the only known template structure of a rat CAPN2/CAST4 complex. The complex model suggests certain residues of bovine CAST4, notably, the TIPPKYQ motif sequence, and the structural elements of these residues, which are important for CAPN1 inhibition. In particular, as CAST4 docks near the flexible active site of CAPN1, conformational changes at the interaction site after binding could be directly related to CAST4 inhibitory activity. These functional interfaces can serve as a guide to the site-mutagenesis in research on bovine CAPN1 structure-function relationships for the design of small molecules inhibitors to prevent uncontrolled and unspecific degradation in the proteolysis of key protease substrates. PMID:24806345

  3. Development of a Western Blot Assay for Detection of Bovine Immunodeficiency-Like Virus Using Capsid and Transmembrane Envelope Proteins Expressed from Recombinant Baculovirus

    PubMed Central

    Abed, Y.; St-Laurent, G.; Zhang, H.; Jacobs, R. M.; Archambault, D.

    1999-01-01

    A 120-amino-acid polypeptide selected from the transmembrane protein region (tTM) and the major capsid protein p26 of bovine immunodeficiency-like virus (BIV) were expressed as fusion proteins from recombinant baculoviruses. The antigenic reactivity of both recombinant fusion proteins was confirmed by Western blot with bovine and rabbit antisera to BIV. BIV-negative bovine sera and animal sera positive for bovine syncytial virus and bovine leukemia virus failed to recognize the recombinant fusion proteins, thereby showing the specificity of the BIV Western blot. One hundred and five bovine serum samples were tested for the presence of anti-BIV antibodies by the recombinant protein-based Western blot and a reference Western blot assay using cell culture-derived virions as test antigens. There was a 100% concordance when the p26 fusion protein was used in the Western blot. However, the Western blot using the tTM fusion protein as its test antigen identified four BIV-positive bovine sera which had tested negative in both the p26 recombinant-protein-based and the reference Western blot assays. This resulted in the lower concordance of 96.2% between the tTM-protein-based and reference Western blot assays. The results of this study showed that the recombinant p26 and tTM proteins can be used as test antigens for the serodetection of BIV-infection in animals. PMID:10066648

  4. Temperature effect on isometric tension is mediated by regulatory proteins tropomyosin and troponin in bovine myocardium

    PubMed Central

    Fujita, Hideaki; Kawai, Masataka

    2002-01-01

    The effect of temperature on isometric tension with and without the regulatory proteins tropomyosin and troponin was studied in bovine myocardium using a thin filament removal and reconstitution protocol. In control bovine myocardium, isometric tension increased linearly with temperature in the range 5–40 °C: isometric tension at 10 and 30 °C was 0.65 and 1.28 times that at 20 °C, respectively, with a Q10 of about 1.4. In actin filament-reconstituted myocardium without regulatory proteins, the temperature effect on isometric tension was less: isometric tension at 10 and 30 °C was 0.96 and 1.17 times that at 20 °C, respectively, with a Q10 of about 1.1. The temperature dependence of the apparent rate constants was studied using sinusoidal analysis. The temperature dependence of 2?b (rate constant of delayed tension phase) did not vary significantly with the regulatory proteins under the standard activating condition (5 mm MgATP, 8 mm Pi, 200 mm ionic strength, pCa 4.66, pH 7.00). Q10 for 2?b in control and actin filament-reconstituted myocardium was 3.8 and 4.0, respectively. There were two phases to the temperature dependence of 2?c (rate constant of quick recovery). In control and thin filament-reconstituted myocardium, Q10 for 2?c was approximately 5.5 in the low temperature range (? 25 °C) and 2.7 in the high temperature range (? 30 °C). In actin filament-reconstituted myocardium, Q10 for 2?c was 8.5 in the low temperature range and 3.6 in the high temperature range. The above results demonstrate that regulatory proteins augment the temperature dependence of isometric tension, indicating that the regulatory proteins may modify the actomyosin interaction. PMID:11850518

  5. Nicotinic Acid Increases Adiponectin Secretion from Differentiated Bovine Preadipocytes through G-Protein Coupled Receptor Signaling

    PubMed Central

    Kopp, Christina; Hosseini, Afshin; Singh, Shiva P.; Regenhard, Petra; Khalilvandi-Behroozyar, Hamed; Sauerwein, Helga; Mielenz, Manfred

    2014-01-01

    The transition period in dairy cows (3 weeks prepartum until 3 weeks postpartum) is associated with substantial mobilization of energy stores, which is often associated with metabolic diseases. Nicotinic acid (NA) is an antilipolytic and lipid-lowering compound used to treat dyslipidaemia in humans, and it also reduces non-esterified fatty acids in cattle. In mice the G-protein coupled receptor 109A (GPR109A) ligand NA positively affects the secretion of adiponectin, an important modulator of glucose and fat metabolism. In cattle, the corresponding data linking NA to adiponectin are missing. Our objective was to examine the effects of NA on adiponectin and AMPK protein abundance and the expression of mRNAs of related genes such as chemerin, an adipokine that enhances adiponectin secretion in vitro. Differentiated bovine adipocytes were incubated with pertussis toxin (PTX) to verify the involvement of GPR signaling, and treated with 10 or 15 µM NA for 12 or 24 h. NA increased adiponectin concentrations (p ? 0.001) and the mRNA abundances of GPR109A (p ? 0.05) and chemerin (p ? 0.01). Pre-incubation with PTX reduced the adiponectin response to NA (p ? 0.001). The NA-stimulated secretion of adiponectin and the mRNA expression of chemerin in the bovine adipocytes were suggestive of GPR signaling-dependent improved insulin sensitivity and/or adipocyte metabolism in dairy cows. PMID:25411802

  6. Nicotinic acid increases adiponectin secretion from differentiated bovine preadipocytes through G-protein coupled receptor signaling.

    PubMed

    Kopp, Christina; Hosseini, Afshin; Singh, Shiva P; Regenhard, Petra; Khalilvandi-Behroozyar, Hamed; Sauerwein, Helga; Mielenz, Manfred

    2014-01-01

    The transition period in dairy cows (3 weeks prepartum until 3 weeks postpartum) is associated with substantial mobilization of energy stores, which is often associated with metabolic diseases. Nicotinic acid (NA) is an antilipolytic and lipid-lowering compound used to treat dyslipidaemia in humans, and it also reduces non-esterified fatty acids in cattle. In mice the G-protein coupled receptor 109A (GPR109A) ligand NA positively affects the secretion of adiponectin, an important modulator of glucose and fat metabolism. In cattle, the corresponding data linking NA to adiponectin are missing. Our objective was to examine the effects of NA on adiponectin and AMPK protein abundance and the expression of mRNAs of related genes such as chemerin, an adipokine that enhances adiponectin secretion in vitro. Differentiated bovine adipocytes were incubated with pertussis toxin (PTX) to verify the involvement of GPR signaling, and treated with 10 or 15 µM NA for 12 or 24 h. NA increased adiponectin concentrations (p ? 0.001) and the mRNA abundances of GPR109A (p ? 0.05) and chemerin (p ? 0.01). Pre-incubation with PTX reduced the adiponectin response to NA (p ? 0.001). The NA-stimulated secretion of adiponectin and the mRNA expression of chemerin in the bovine adipocytes were suggestive of GPR signaling-dependent improved insulin sensitivity and/or adipocyte metabolism in dairy cows. PMID:25411802

  7. Factors regulating the bovine, caprine, rat and human ovarian aromatase promoters in a bovine granulosa cell model.

    PubMed

    Sahmi, Fatiha; Nicola, Edmir S; Zamberlam, Gustavo O; Gonçalves, Paulo D B; Vanselow, Jens; Price, Christopher A

    2014-05-01

    The ovarian promoter of the primate and rodent genes encoding cytochrome P450 aromatase (CYP19A1) are robustly responsive to forskolin in luteinized cell models, whereas the ruminant ovarian promoter is minimally active. We explored this discrepancy by investigating the activity of the bovine ovarian promoter in two bovine granulosa cell models, luteinizing and non-luteinizing cells in vitro. In non-luteinizing cells, both FSH and IGF1 increased abundance of transcripts derived from the ovarian promoter. Comparison of the activity of promoters of several species in response to transcription factors (forskolin, NR5A2, FOXL2) in luteinizing cells demonstrated that a rat ovarian promoter-luciferase reporter was regulated mainly by forskolin (18-fold increase over basal expression) and addition of NR5A2 or FOXL2 had no further effect. Activity of a human promoter was significantly increased by NR5A2 plus forskolin (153-fold) compared with forskolin alone (71-fold over basal); addition of FOXL2 did not significantly increase promoter activity. Forskolin alone provoked minor activation of caprine and bovine promoter reporters (3-fold over basal), and addition of NR5A2 increased activity (7- to 11-fold). When forskolin, NR5A2 and FOXL2 treatments were combined, the activity of the caprine and bovine promoters increased to 20- and 34-fold, respectively. These data suggest that a major reason why CYP19A1 is not expressed in luteinized cells (and the corpus luteum) of ruminants may be the stimulatory effect of FOXL2, which does not appear to be the case in the human and rat. PMID:24556528

  8. Profile of bovine proteins in retained and normally expelled placenta in dairy cows.

    PubMed

    Kankofer, M; Wawrzykowski, J; Hoedemaker, M

    2014-04-01

    Tissue-specific protein profile is determined by its function, structure, intensity of metabolism and usefulness. This profile remains under hormonal control. Any disturbance in the general metabolism may be reflected in changes in both protein quantity and quality. These changes can be of low or high specificity, and some can be used as clinical markers of pathological conditions. The aim of this study was to describe and to compare the protein profile of caruncle and foetal villi of bovine placenta that was either properly released or retained. Placental tissues were collected from healthy cows, divided into releasing and retaining foetal membranes, homogenized and subjected to 1D and 2D electrophoresis. Computer-aided analysis of gel images showed essential qualitative and quantitative alterations in protein profile between tissues that were properly released and retained. Alterations concerned both the number of fractions and spots as well as the intensity of staining. This preliminary study provides a general overview of the differences in the protein profile between released and retained foetal membranes. It may allow for selecting the group of proteins or single molecules, which should be further analysed in detail as possible markers differentiating the retention of foetal membranes in cows from placentas that were released spontaneously. The continuation of the study for the identification of particular spots detected in 2D gels is necessary. PMID:24325199

  9. Development of reduced fat minced meats using inulin and bovine plasma proteins as fat replacers.

    PubMed

    Rodriguez Furlán, Laura T; Padilla, Antonio Pérez; Campderrós, Mercedes E

    2014-02-01

    This work deals with the effect of the addition of inulin and bovine plasma proteins as fat replacers, on the quality of minced meat. The proteins are obtained by ultrafiltration and freeze-drying. The following determinations were carried out: chemical composition, sensorial analysis (color, flavor, taste and consistency), emulsion stability and instrumental texture analysis of samples. The resulting formulations were compared with full-fat minced meat, as control. The results showed an increase of protein contents after fat replacement, while a fat reduction of 20-35% produced light products enriched with proteins and inulin as the functional ingredient. No change was observed in color, flavor, or taste among the samples. However, the sensory analysis showed that the combination of plasma protein (2.5%w/w) and inulin (2%w/w) had the best acceptability with respect to consistency, and had a lower fat drain from the emulsion. Texture profile analysis revealed that this formulation assimilated the control texture properties, being that this result is required for adequate consumer acceptance. PMID:24200568

  10. Association of the GTP-binding protein Rab3A with bovine adrenal chromaffin granules

    SciTech Connect

    Darchen, F.; Hammel, F.; Monteils, M.P.; Scherman, D. (Centre National de la Recherche Scientifique, Paris (France)); Zahraoui, A.; Tavitian, A. (Institut National de la Sante et de la Recherche Medicale, Paris (France))

    1990-08-01

    The Rab3A protein belongs to a large family of small GTP-binding proteins that are present in eukaryotic cells and that share amino acid identities with the Ras proteins (products of the ras protooncogenes). Rab3A, which is specifically located in nervous and endocrine tissues, is suspected to play a key role in secretion. Its localization was investigated in bovine adrenal gland by using a polyclonal antibody. Rab3A was detected in adrenal medulla but not in adrenal cortex. In cultured adrenal medulla cells, Rab3A was specifically expressed in the catecholamine-secreting chromaffin cells. Subcellular fractionation suggested that Rab3A is about 30% cytosolic and that particulate Rab3A is associated with the membrane of chromaffin granules (the catecholamine storage organelles) and with a second compartment likely to be the plasma membrane. The Rab3A localization on chromaffin granule membranes was confirmed by immunoadsorption with an antibody against dopamine {beta}-hydroxylase. Rab3A was not extracted from this membrane by NaCl or KBr but was partially extracted by urea and totally solubilized by Triton X-100, suggesting either an interaction with an intrinsic protein or a membrane association through fatty acid acylation. This study suggests that Rab3A, which may also be located on other secretory vesicles containing noncharacterized small GTP-binding proteins, is involved in their biogenesis or in the regulated secretion process.

  11. Removal of Available Decorin Core-Protein from Powdered Bovine Hide by Treatments used to Process Intact Hides into Leather

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using a modification of a previously developed sandwich Elisa procedure to measure decorin core-protein (DCP), we determined the available decorin content of a sample of raw powdered bovine hide before and after treatment with the reagents used in the early steps of the process for converting a hide...

  12. THE EFFECT OF DIETARY LIPOIC ACID ON METABOLIC HORMONES AND ACUTE PHASE PROTEINS IN STEERS CHALLENGED WITH INFECTIOUS BOVINE RHINOTRACHEITIS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Crossbred steers challenged with infectious bovine rhinotracheitis virus (IBRV) were used to determine the effect of supplemental lipoic acid (LA) on circulating metabolic hormones and acute phase proteins. Steers (n = 32; BW = 260 + or - 6 kg) were randomly assigned to four treatments: control (CON...

  13. A new bovine conjunctiva model shows that Listeria monocytogenes invasion is associated with lysozyme resistance.

    PubMed

    Warren, Jessica; Owen, A Rhys; Glanvill, Amy; Francis, Asher; Maboni, Grazieli; Nova, Rodrigo J; Wapenaar, Wendela; Rees, Catherine; Tötemeyer, Sabine

    2015-08-31

    Listerial keratoconjunctivitis ('silage eye') is a wide spread problem in ruminants causing economic losses to farmers and impacts negatively on animal welfare. It results from direct entry of Listeria monocytogenes into the eye, often following consumption of contaminated silage. An isolation protocol for bovine conjunctival swabbing was developed and used to sample both infected and healthy eyes bovine eyes (n=46). L. monocytogenes was only isolated from one healthy eye sample, and suggests that this organism can be present without causing disease. To initiate a study of this disease, an infection model was developed using isolated conjunctiva explants obtained from cattle eyes post slaughter. Conjunctiva were cultured and infected for 20h with a range of L. monocytogenes isolates (n=11), including the healthy bovine eye isolate and also strains isolated from other bovine sources, such as milk or clinical infections. Two L. monocytogenes isolates (one from a healthy eye and one from a cattle abortion) were markedly less able to invade conjunctiva explants, but one of those was able to efficiently infect Caco2 cells indicating that it was fully virulent. These two isolates were also significantly more sensitive to lysozyme compared to most other isolates tested, suggesting that lysozyme resistance is an important factor when infecting bovine conjunctiva. In conclusion, we present the first bovine conjunctiva explant model for infection studies and demonstrate that clinical L. monocytogenes isolates from cases of bovine keratoconjunctivitis are able to infect these tissues. PMID:25778543

  14. Association between SSCP haplotypes at the bovine growth hormone gene and milk protein percentage.

    PubMed

    Lagziel, A; Lipkin, E; Soller, M

    1996-03-01

    The bovine Growth Hormone gene (bGH) is an attractive candidate gene for milk production in cattle. Single-strand conformation polymorphisms at bGH were identified and used to define haplotype configurations at this gene in the Israeli Holstein dairy cattle population (Bos taurus) and in the parent animals of the International Bovine Reference Family Panel (a collection of B. taurus and B. indicus crosses). B. taurus and B. indicus haplotypes at the bGH gene differed qualitatively, confirming the previously proposed long evolutionary separation of these cattle subraces. Only a small number of bGH haplotypes were present in the Israel Holstein population. One of the haplotypes, apparently of B. indicus origin, was found to have a highly significant positive effect on milk protein percentage. This illustrates the utility of the haplotype approach for uncovering candidate gene involvement in quantitative genetic variation in agricultural populations. The strong effect of an indicine haplotype in a taurine background raises the possibility that indicine alleles at other candidate genes may comprise a genetic resource for improvement of taurine populations. It is proposed that haplotype analysis may be a useful adjunct to measures of genetic distance for evaluating rare breeds with respect to gene conservation. PMID:8849900

  15. Assessing the susceptibility of transgenic mice overexpressing deer prion protein to bovine spongiform encephalopathy.

    PubMed

    Vickery, Christopher M; Lockey, Richard; Holder, Thomas M; Thorne, Leigh; Beck, Katy E; Wilson, Christina; Denyer, Margaret; Sheehan, John; Marsh, Sarah; Webb, Paul R; Dexter, Ian; Norman, Angela; Popescu, Emma; Schneider, Amanda; Holden, Paul; Griffiths, Peter C; Plater, Jane M; Dagleish, Mark P; Martin, Stuart; Telling, Glenn C; Simmons, Marion M; Spiropoulos, John

    2014-02-01

    Several transgenic mouse models have been developed which facilitate the transmission of chronic wasting disease (CWD) of cervids and allow prion strain discrimination. The present study was designed to assess the susceptibility of the prototypic mouse line, Tg(CerPrP)1536(+/-), to bovine spongiform encephalopathy (BSE) prions, which have the ability to overcome species barriers. Tg(CerPrP)1536(+/-) mice challenged with red deer-adapted BSE resulted in 90% to 100% attack rates, and BSE from cattle failed to transmit, indicating agent adaptation in the deer. PMID:24257620

  16. Assessing the Susceptibility of Transgenic Mice Overexpressing Deer Prion Protein to Bovine Spongiform Encephalopathy

    PubMed Central

    Vickery, Christopher M.; Lockey, Richard; Holder, Thomas M.; Thorne, Leigh; Beck, Katy E.; Wilson, Christina; Denyer, Margaret; Sheehan, John; Marsh, Sarah; Webb, Paul R.; Dexter, Ian; Norman, Angela; Popescu, Emma; Schneider, Amanda; Holden, Paul; Griffiths, Peter C.; Plater, Jane M.; Dagleish, Mark P.; Martin, Stuart; Telling, Glenn C.; Simmons, Marion M.

    2014-01-01

    Several transgenic mouse models have been developed which facilitate the transmission of chronic wasting disease (CWD) of cervids and allow prion strain discrimination. The present study was designed to assess the susceptibility of the prototypic mouse line, Tg(CerPrP)1536+/?, to bovine spongiform encephalopathy (BSE) prions, which have the ability to overcome species barriers. Tg(CerPrP)1536+/? mice challenged with red deer-adapted BSE resulted in 90% to 100% attack rates, and BSE from cattle failed to transmit, indicating agent adaptation in the deer. PMID:24257620

  17. Grafting of bovine serum albumin proteins on plasma-modified polymers for potential application in tissue engineering

    NASA Astrophysics Data System (ADS)

    Kasálková, Nikola Slepi?ková; Slepi?ka, Petr; Kolská, Zde?ka; Hoda?ová, Petra; Ku?ková, Št?pánka; Švor?ík, Václav

    2014-04-01

    In this work, an influence of bovine serum albumin proteins grafting on the surface properties of plasma-treated polyethylene and poly- l-lactic acid was studied. The interaction of the vascular smooth muscle cells with the modified polymer surface was determined. The surface properties were characterized by X-ray photoelectron spectroscopy, atomic force microscopy, nano-LC-ESI-Q-TOF mass spectrometry, electrokinetic analysis, and goniometry. One of the motivations for this work is the idea that by the interaction of the cell with substrate surface, the proteins will form an interlayer between the cell and the substrate. It was proven that when interacting with the plasma-treated high-density polyethylene and poly- l-lactic acid, the bovine serum albumin protein is grafted on the polymer surface. Since the proteins are bonded to the substrate surface, they can stimulate cell adhesion and proliferation.

  18. Characterization of Staphylococcus aureus strains isolated from subclinic bovine mastitis by protein patterns, antibiotic resistance and plasmid profile

    Microsoft Academic Search

    Fatma ÖZTÜRK; Leyla AÇIK

    Summary: A total of 50 Staphylococcus aureus strains isolated from bovine subclinical mastitis in Hatay region were characterized by protein patterns, antibiotic resistance and plasmid profiles. SDS-PAGE analysis of whole-cell protein extracts belong to S. aureus strains produced patterns containing 17-49 discrete bands with molecular weights of >14.4-<116 kDa. S. aureus strains clustered into two cluster on the basis of

  19. Prion protein gene (PRNP) variants and evidence for strong purifying selection in functionally important regions of bovine exon 3

    PubMed Central

    Seabury, Christopher M.; Honeycutt, Rodney L.; Rooney, Alejandro P.; Halbert, Natalie D.; Derr, James N.

    2004-01-01

    Amino acid replacements encoded by the prion protein gene (PRNP) have been associated with transmissible and hereditary spongiform encephalopathies in mammalian species. However, an association between bovine spongiform encephalopathy (BSE) and bovine PRNP exon 3 has not been detected. Moreover, little is currently known regarding the mechanisms of evolution influencing the bovine PRNP gene. Therefore, in this study we evaluated the patterns of nucleotide variation associated with PRNP exon 3 for 36 breeds of domestic cattle and representative samples for 10 additional species of Bovinae. The results of our study indicate that strong purifying selection has intensely constrained PRNP over the long-term evolutionary history of the subfamily Bovinae, especially in regions considered to be of functional, structural, and pathogenic importance in humans as well as other mammals. The driving force behind this intense level of purifying selection remains to be explained. PMID:15477588

  20. Simultaneous separation and quantitation of the major bovine whey proteins including proteose peptone and caseinomacropeptide by reversed-phase high-performance liquid chromatography on polystyrene–divinylbenzene

    Microsoft Academic Search

    David F. Elgar; Carmen S. Norris; John S. Ayers; Mark Pritchard; Don E. Otter; Kate P. Palmano

    2000-01-01

    A precise, sensitive and reliable RP-HPLC method was developed to enable not only unequivocal determination of ?-lactalbumin and ?-lactoglobulin in bovine whey samples, but also simultaneous measurement of proteose peptone, caseinomacropeptide, bovine serum albumin and immunoglobulin G. The optimised method on the Resource RPC column allowed separation of the proteins in 30 min and could be applied to the analysis

  1. Bovine Brain: An in vitro Translational Model in Developmental Neuroscience and Neurodegenerative Research

    PubMed Central

    Peruffo, Antonella; Cozzi, Bruno

    2014-01-01

    Animal models provide convenient and clinically relevant tools in the research on neurodegenerative diseases. Studies on developmental disorders extensively rely on the use of laboratory rodents. The present mini-review proposes an alternative translational model based on the use of fetal bovine brain tissue. The bovine (Bos taurus) possesses a large and highly gyrencephalic brain and the long gestation period (41?weeks) is comparable to human pregnancy (38–40?weeks). Primary cultures obtained from fetal bovine brain constitute a validated in vitro model that allows examinations of neurons and/or glial cells under controlled and reproducible conditions. Physiological processes can be also studied on cultured bovine neural cells incubated with specific substrates or by electrically coupled electrolyte-oxide-semiconductor capacitors that permit direct recording from neuronal cells. Bovine neural cells and specific in vitro cell culture could be an alternative in comparative neuroscience and in neurodegenerative research, useful for studying development of normal and altered circuitry in a long gestation mammalian species. Use of bovine tissues would promote a substantial reduction in the use of laboratory animals. PMID:25072040

  2. A prediction method for the isoelectric point of binary protein mixtures of bovine serum albumin and lysozyme adsorbed on colloidal titania and alumina particles.

    PubMed

    Rezwan, Kurosch; Meier, Lorenz P; Gauckler, Ludwig J

    2005-04-12

    Bovine serum albumin and lysozyme mixtures of different mole fractions were adsorbed to colloidal alumina (116 nm) and titania particle (271 nm) suspensions of 2 vol % solid content for 16 h at pH 7.5. The total protein amount normalized to the powder surface area was 1000 ng/cm2. The zeta potential of the protein-treated suspensions was measured as a function of pH and the isoelectric point (IEP) obtained. A simple prediction model in two refinement steps was derived and evaluated for the obtained IEPs. The best model fit which takes into account moles of protein and surface fractions yielded an average prediction error of 7.5% and a maximum error of 16.7%. PMID:15807593

  3. Elementary steps of the cross-bridge cycle in bovine myocardium with and without regulatory proteins.

    PubMed Central

    Fujita, Hideaki; Sasaki, Daisuke; Ishiwata, Shin'ichi; Kawai, Masataka

    2002-01-01

    The role of regulatory proteins in the elementary steps of the cross-bridge cycle in bovine myocardium was investigated. The thin filament was selectively removed by gelsolin and the actin filament was reconstituted without tropomyosin or troponin. Further reconstitution was achieved by adding tropomyosin and troponin. The effects of MgATP and phosphate (Pi) on the rate constants of exponential processes were studied in control, actin filament-reconstituted, and thin filament-reconstituted myocardium at pCa < or = 4.66, pH 7.00, 25 degrees C. In control myocardium, the MgATP association constant was 9.1 +/- 1.3 mM(-1), and the Pi association constant 0.14 +/- 0.04 mM(-1). The equilibrium constant of the cross-bridge detachment step was 2.6 +/- 0.4, and the equilibrium constant of the force generation step was 0.59 +/- 0.04. In actin filament-reconstituted myocardium without regulatory proteins, the MgATP association constant was approximately the same, and the Pi association constant increased to 2.8x. The equilibrium constant of cross-bridge detachment decreased to 0.2x, but the equilibrium constant of the force generation step increased to 4x. These kinetic constants regained control values after reconstitution of the thin filament. These results indicate that tension/cross-bridge in the presence of regulatory proteins is approximately 1.5-1.7x of that in the absence of regulatory proteins. These results further indicate that regulatory proteins promote detachment of cross-bridges. PMID:11806933

  4. Anaplasma marginale major surface protein 1a: A marker of strain diversity with implications for control of bovine anaplasmosis.

    PubMed

    Cabezas-Cruz, Alejandro; de la Fuente, José

    2015-04-01

    Classification of bacteria is challenging due to the lack of a theory-based framework. In addition, the adaptation of bacteria to ecological niches often results in selection of strains with diverse virulence, pathogenicity and transmission characteristics. Bacterial strain diversity presents challenges for taxonomic classification, which in turn impacts the ability to develop accurate diagnostics and effective vaccines. Over the past decade, the worldwide diversity of Anaplasma marginale, an economically important tick-borne pathogen of cattle, has become apparent. The extent of A. marginale strain diversity, formerly underappreciated, has contributed to the challenges of classification which, in turn, likely impacts the design and development of improved vaccines. Notably, the A. marginale surface protein 1a (MSP1a) is a model molecule for these studies because it serves as a marker for strain identity, is both an adhesin necessary for infection of cells and an immuno-reactive protein and is also an indicator of the evolution of strain diversity. Herein, we discuss a molecular taxonomic approach for classification of A. marginale strain diversity. Taxonomic analysis of this important molecule provides the opportunity to understand A. marginale strain diversity as it relates geographic and ecological factors and to the development of effective vaccines for control of bovine anaplasmosis worldwide. PMID:25802034

  5. Analysis of the binding of bovine and human fibrinogen to ferritin: evidence that fibrinogen is a common ferritin-binding protein in mammals.

    PubMed

    Okada, Akiko; Yoshikawa, Yasunaga; Watanabe, Kiyotaka; Orino, Koichi

    2015-08-01

    Both human and horse fibrinogen are heme-binding proteins, and horse fibrinogen also exhibits heme-mediated ferritin binding. This study found that bovine and human fibrinogen are heme-mediated ferritin-binding proteins and demonstrated direct binding of bovine ferritin to protoporphyrin (PPIX) and its derivatives or to Zn ions. Binding of bovine and human fibrinogen to bovine spleen ferritin coated on microtiter plate wells was detected using an anti-human fibrinogen antibody, and this binding was inhibited in a dose-dependent manner by hemin (iron-PPIX) and also inhibited by Zn-PPIX. PPIX showed less of an inhibitory effect on the binding of bovine and human fibrinogen to bovine ferritin. The inhibitory effect of Sn-PPIX was similar to that of PPIX, but with respect to human fibrinogen, PPIX did not inhibit the binding of human fibrinogen to ferritin. Bovine fibrinogen immobilized on CNBr-activated Sepharose 4B beads showed affinity for hemin, Sn-PPIX, Zn-PPIX, and iron-free PPIX in the order Sn-PPIX < iron-free PPIX < hemin < Zn-PPIX. The fibrinogen beads also directly bound to zinc ions. These results suggest that bovine fibrinogen is a heme- and zinc-binding protein and that binding of circulating mammalian fibrinogen to ferritin is heme mediated. PMID:25860295

  6. Locations and immunoreactivities of phosphorylation sites on bovine and porcine tau proteins and a PHF-tau fragment.

    PubMed

    Poulter, L; Barratt, D; Scott, C W; Caputo, C B

    1993-05-01

    Tau protein is a phosphorylated neuronal microtubule-associated protein. Tau protein is also present in the major pathological lesions of Alzheimer's disease in an insoluble hyperphosphorylated state as paired helical filaments (PHFs). We have investigated the phosphorylation state of control taus and a fragment of PHF-tau. Tau samples were digested with protease, separated by reversed-phase high-performance liquid chromatography, and analyzed by mass spectrometry and Edman microsequencing. The serine homologous with S404 of human tau 441 was phosphorylated on bovine and porcine tau and up to two phosphates were present on a peptide of amino acids 182-240 of bovine tau (193-251 of human tau 441). The serine within the KSPV motif was not phosphorylated on bovine or porcine tau. PHF-tau fragments, isolated from pronase-treated PHFs encompassed a 93-amino acid region within the microtubule binding domain. Enzymatic digestion and mass spectrometric analysis showed no phosphate was present and a second carboxyl terminus was identified at E380. Antibodies T3P and SMI34, which recognize PHF-tau and peptides phosphorylated at the sequence KSPV, both reacted with bovine and porcine tau even though the KSPV sequence was not phosphorylated. These data indicate that the 93-amino acid sequence of F5.5 tau from PHFs is not phosphorylated, and the serine equivalent to S404 of human tau is phosphorylated in bovine and porcine tau. Antibodies T3P and SMI34 react with phosphorylated epitopes that are not unique to PHF-tau and that are not necessarily at the KSPV site. PMID:8486651

  7. Expression of recombinant bovine gamma B-, gamma C- and gamma D-crystallins and correlation with native proteins.

    PubMed

    Hay, R E; Andley, U P; Petrash, J M

    1994-05-01

    Despite the use of bovine gamma-crystallins in numerous biophysical and chemical studies, characterization of these proteins at the molecular level is incomplete. Bovine lenses have at least six gamma-crystallin protein fractions currently assigned as gamma s/I, gamma A/IVb, gamma B/II, gamma C/IIIb, gamma D/IIIa and gamma E/IVa. A lack of primary sequence data for corresponding gamma-crystallin genes and proteins, however, has made these assignments tenuous. To clarify these assignments, we have over-expressed recombinant bovine gamma-crystallin proteins in Escherichia coli using complementary DNAs corresponding to gamma B-, gamma C-, and gamma D-crystallin genes. The recombinant crystallins were characterized by chromatographic and spectroscopic comparisons with native bovine crystallin fractions gamma II, gamma IIIa and gamma IIIb. The elution of recombinant gamma B and native gamma II proteins was identical on cation-exchange chromatography as expected; however, recombinant gamma C coeluted with gamma IIIa and recombinant gamma D co-eluted with gamma IIIb. Sequential Edman degradation through the first 29 residues of the native gamma IIIa and gamma IIIb polypeptides confirmed the colinearity of their sequences with those predicted from the gamma C- and gamma D-crystallin genes, respectively. Absorption and UV circular dichroism (CD) spectra of the recombinant proteins were almost identical to those of their native counterparts, indicating that the secondary and tertiary structures of the recombinant proteins were characteristic for gamma-crystallins. Based on these data the bovine gamma-crystallins proteins and genes are correlated as follows: II/gamma B, IIIa/gamma C and IIIb/gamma D. The assignment of gamma IIIb (previously characterized as having a low critical temperature for phase separation) with gamma D rather than gamma C proves an exception to the hypothesis that the gamma ABC-crystallin group is more resistant to phase separation than the gamma DEF group. These corrected assignments should provide a more substantial base for investigations of residues responsible for phase separation and other biophysical characteristics. Additionally, expression of recombinant gamma-crystallins with structures similar to native proteins may prove to be useful in probing specific structure-function relationships of the gamma-crystallins. PMID:7925695

  8. Oxidative damage of bovine serum albumin and other enzyme proteins by iron-chelate complexes.

    PubMed

    Ogino, T; Okada, S

    1995-12-14

    Direct oxidative protein damage by iron-nitrilotriacetate (NTA), as well as physiological iron complexes, iron-citrate and iron-ADP was studied in the presence or absence of H2O2, using bovine serum albumin (BSA), glucose-6-phosphate dehydrogenase (G-6-PD), glutathione reductase (GSSGRase) and catalase as the target proteins. Both Fe(III)NTA+H2O2 and Fe(II)NTA+H2O2 caused marked BSA fragmentation which accompanied the decrease in the intrinsic tryptophan fluorescence and appearance of bityrosine fluorescence. However, Fe(III)citrate+H2O2 showed only slight BSA fragmentation. In the absence of H2O2, Fe(II) NTA but not Fe(III)NTA caused similar but slight BSA fragmentation, which depended on the molecular oxygen. Fe(II)citrate also showed O2-dependent BSA fragmentation to a comparable degree, however, Fe(II)ADP showed no detectable BSA damage. BSA fragmentation by Fe(II)NTA+O2 and by Fe(III)NTA+H2O2 resulted in the appearance of the new alpha-amino groups. Electron spin resonance study using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) as a spin trapping reagent showed DMPO-OH spin adduct, which suggests the presence of hydroxyl radical, in Fe(III)NTA+H2O2, but not in Fe(II)NTA+O2 system. Fe(II)NTA inactivated G-6-PD and GSSGRase in a O2-dependent manner, however, G-6-PD was more susceptible to the damage. This enzyme inactivation also accompanied the protein fragmentation and was not due to simple sulfhydryl oxidation. Catalase was not significantly inactivated nor fragmented by Fe(II)NTA+O2. These findings suggest that the interaction between proteins and iron-chelate complexes is important in iron catalyzed oxidative damage, and that the structure of the chelating agent may determine the target molecules. PMID:8541312

  9. nutrient Metabolism Nutrient utilization and Protein Turnover in the Hindlimb of Cattle Treated with Bovine Somatotropin1'2

    Microsoft Academic Search

    VVES R. BOISCLAIR; DALE E. BAUMAN; ALAN W. BELL

    Our objectives were to assess the effects of chronic administration of recombinant bovine somatotropin (bST) on nutrient utilization and protein turnover in the hindlimb of growing Holstein steers. Ex ternal iliac vessels were catheterized to allow for hindlimb measurements of arteriovenous differences and blood flow. Animals were used in a single-reversal design with 16-d treatment periods of daily intramus cular

  10. Ultrarapid detection of bovine whey proteins in powdered soybean milk by perfusion reversed-phase high-performance liquid chromatography

    Microsoft Academic Search

    M. C. Garc??a; M. L. Marina; M. Torre

    1998-01-01

    A perfusion reversed-phase high-performance liquid chromatographic method has been developed to simultaneously separate soybean and bovine whey proteins (?-lactalbumin and ?-lactoglobulins (A+B)) in a very short analysis time (?5 min). The method consisted of a linear binary gradient water–acetonitrile–0.10% trifluoroacetic acid at a flow-rate of 3 ml\\/min, with the column thermostated at 60°C, and ultraviolet detection at 254 nm. This

  11. Preparation, Characterization, and Localization Against Bovine MP26, an Integral Protein from Plasma Membrane of Antisera Lens Fiber

    Microsoft Academic Search

    DAVID L. PAUL; DANIEL A. GOODENOUGH

    Polyclonal antisera were prepared in rabbits using both native and chymotrypsin- digested bovine lens fiber plasma membranes. MP26, the principal protein of lens fiber plasma membranes, and CT20, a chymotryptic fragment of MP26, were isolated electrophoretically and used to purify anti-MP26 and anti-CT20 activity from the respective antisera by affinity chromatography. These affinity-purified antisera were characterized by immunoreplica. Im- munofluorescence

  12. Lack of repair of rat skull critical size defect treated with bovine morphometric protein bound to microgranular bioabsorbable hydroxyapatite

    Microsoft Academic Search

    Gabriel Ramalho Ferreira; Tania Mary Cestari; José Mauro Granjeiro; Rumio Taga

    2004-01-01

    The ability of a pool of bovine bone morphogenetic proteins bound to synthetic microgranular hydroxyapatite (BMPb-HA) to stimulate bone repair was determined in rat critical size defects. An 8-mm diameter defect was created in the calvaria of 25 rats. In 15 rats, the defects were filled with BMPb-HA homogenized with blood (experimental group), and in 10 rats the defects were

  13. The kinetics of inflammation and phagocytosis during bovine mastitis induced by Streptococcus agalactiae bearing the protein X

    Microsoft Academic Search

    P. Rainard; Y. Lautrou; P. Sarradin; A. Coulibaly; B. Poutrel

    1991-01-01

    The protein X of Streptococcus agalactiae is a surface antigen borne by a high proportion of strains isolated from bovine mastitis. We have tested the capacity of two strains of X-bearing Streptococcus agalactiae to induce mastitis in dairy cows. The reference X-strain (411.07) produced an intramammary infection with local clinical signs in the three inoculated quarters. Another X-bearing strain (443.31)

  14. ISOLATION AND PURIFICATION OF BOVINE IMMUNOGLOBULINS: USE OF SEPHACRYL S-300 FILTRATION AVOIDS PROTEIN

    E-print Network

    Paris-Sud XI, Université de

    ISOLATION AND PURIFICATION OF BOVINE IMMUNOGLOBULINS: USE OF SEPHACRYL S-300 FILTRATION AVOIDS Résumé ISOLEMENT ET PURIFICATION DES IMMUNOGLOBULINES BOVINES: L'UTILISATION DE LA FILTRATION SUR'une filtration sur le gel Sephacryl S-300. Le sérum de colostrum appliqué à cette colonne peut être séparé en

  15. Identification of cell membrane proteins linked to susceptibility to bovine viral diarrhoea virus infection

    Microsoft Academic Search

    C. Schelp; I. Greiser-Wilke; G. Wolf; M. Beer; V. Moennig; B. Liess

    1995-01-01

    Summary Three monoclonal antibodies directed against cell surface molecules of bovine cells inhibited subsequent infections with bovine viral diarrhoea virus (BVDV). They specifically blocked the infectivity of three non-cytopathogenic and three cytopathogenic BVDV strains. These results showed that an important mechanism for virus uptake was inhibited. The ligand of the monoclonal antibody BVD\\/CA 17, which blocked infectivity most efficiently, was

  16. Human Milk Protein Production in Xenografts of Genetically Engineered Bovine Mammary Epithelial Stem Cells

    Microsoft Academic Search

    Eugenio Martignani; Peter Eirew; Paolo Accornero; Connie J. Eaves; Mario Baratta; Joseph Najbauer

    2010-01-01

    BackgroundIn the bovine species milk production is well known to correlate with mammary tissue mass. However, most advances in optimizing milk production relied on improvements of breeding and husbandry practices. A better understanding of the cells that generate bovine mammary tissue could facilitate important advances in milk production and have global economic impact. With this possibility in mind, we show

  17. Small-angle neutron scattering study of differences in phase behavior of silica nanoparticles in the presence of lysozyme and bovine serum albumin proteins

    NASA Astrophysics Data System (ADS)

    Yadav, Indresh; Kumar, Sugam; Aswal, V. K.; Kohlbrecher, J.

    2014-03-01

    The differences in phase behavior of anionic silica nanoparticles (88 Å) in the presence of two globular proteins [cationic lysozyme (molecular weight (MW) 14.7 kD) and anionic bovine serum albumin (BSA) (MW 66.4 kD)] have been studied by small-angle neutron scattering. The measurements were carried out on a fixed concentration (1 wt %) of Ludox silica nanoparticles with varying concentrations of proteins (0-5 wt %) at pH = 7. It is found that, despite having different natures (opposite charges), both proteins can render to the same kind of aggregation of silica nanoparticles. However, the concentration regions over which the aggregation is observed are widely different for the two proteins. Lysozyme with very small amounts (e.g., 0.01 wt %) leads to the aggregation of silica nanoparticles. On the other hand, silica nanoparticles coexist with BSA as independent entities at low protein concentrations and turn to aggregates at high protein concentrations (>1 wt %). In the case of lysozyme, the charge neutralization by the protein on the nanoparticles gives rise to the protein-mediated aggregation of the nanoparticles. The nanoparticle aggregates coexist with unaggregated nanoparticles at low protein concentrations, whereas, they coexist with a free protein at higher protein concentrations. For BSA, the nonadsorbing nature of the protein produces the depletion force that causes the aggregation of the nanoparticles at higher protein concentrations. The evolution of the interaction is modeled by the two Yukawa potential, taking account of both attractive and repulsive terms of the interaction in these systems. The nanoparticle aggregation is found to be governed by the short-range attraction for lysozyme and the long-range attraction for BSA. The aggregates are characterized by the diffusion limited aggregate type of mass fractal morphology.

  18. A comparative analysis of rapid methods for purification and refolding of recombinant bovine prion protein

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacterially-produced recombinant prion protein (rPrP) is a frequently used model system for the study of the properties of wild-type and mutant prion proteins by biochemical and biophysical approaches. A range of approaches have been developed for the purification and refolding of untagged, overexpr...

  19. Identification of a nuclear export signal sequence for bovine papillomavirus E1 protein

    SciTech Connect

    Rosas-Acosta, German; Wilson, Van G., E-mail: wilson@medicine.tamhsc.edu

    2008-03-30

    Recent studies have demonstrated nuclear export by papillomavirus E1 proteins, but the requisite export sequence(s) for bovine papillomavirus (BPV) E1 were not defined. In this report we identify three functional nuclear export sequences (NES) present in BPV E1, with NES2 being the strongest in reporter assays. Nuclear localization of BPV1 E1 was modulated by over- or under-expression of CRM1, the major cellular exportin, and export was strongly reduced by the CRM1 inhibitor, Leptomycin B, indicating that E1 export occurs primarily through a CRM1-dependent process. Consistent with the in vivo functional results, E1 bound CRM1 in an in vitro pull-down assay. In addition, sumoylated E1 bound CRM1 more effectively than unmodified E1, suggesting that E1 export may be regulated by SUMO modification. Lastly, an E1 NES2 mutant accumulated in the nucleus to a greater extent than wild-type E1, yet was defective for viral origin replication in vivo. However, NES2 exhibited no intrinsic replication defect in an in vitro replication assay, implying that nucleocytoplasmic shuttling may be required to maintain E1 in a replication competent state.

  20. Chemical structure of the arabinogalactan protein from gum ghatti and its interaction with bovine serum albumin.

    PubMed

    Ghosh, Kanika; Ray, Sayani; Ghosh, Debjani; Ray, Bimalendu

    2015-03-01

    Exudate gums, because of their beneficial properties, have been significant items of international trade in various industries for centuries. This manuscript sets out to gain insight into the fine structural details of an arabinogalactan protein (AGP) of gum ghatti (Anogeissus latifolia gum). The presence of a highly branched 554 kDa AGP having 1,6-linked Galp, 1,2-linked Manp, 1,3-linked Araf and 1,4-linked GlcpA main chain, substituted at O-4,6 of 1,2-linked Manp, and O-3/O-3,4 of 1,6-linked Galp residues by Araf, Arap and Galp units was revealed by chemical, chromatographic, ESMS, and NMR analyses. In particular, ESMS analysis of per acetylated oligomeric fragments derived from AGP by Smith degradation followed by acetylation was described as a commanding tool for providing critical structural information on a spectrum of glycerol tagged oligosaccharides. In addition, formation of an electrostatically driven complex between the isolated AGP and bovine serum albumin resulting in changes in the microenvironment around the tryptophan residues of BSA was established. A moderate radical scavenging activity comparable with those of standard antioxidants was observed from the AGP fraction (?94% at 1 mg/mL) that could be valuable in foods or pharmaceutical products as alternatives to synthetic antioxidants. PMID:25498648

  1. Unfolding protein response signaling is involved in development, maintenance, and regression of the corpus luteum during the bovine estrous cycle.

    PubMed

    Park, Hyo-Jin; Park, Sun-Ji; Koo, Deog-Bon; Kong, Il-Keun; Kim, Min Kyu; Kim, Jin-Man; Choi, Myung-Sook; Park, Young-Ho; Kim, Sun-Uk; Chang, Kyu-Tae; Park, Choon-Keun; Chae, Jung-Il; Lee, Dong-Seok

    2013-11-15

    The corpus luteum (CL) is a transient endocrine organ. Development, maintenance, and regression of CL are effectively controlled by dynamic changes in gene expression. However, it is unknown what types of gene are affected during the CL life span of the estrous cycle in bovine. Here, we determined whether unfolded protein response (UPR) signaling via eIF2?/ATF4/GADD34, p90ATF6/p50ATF6, and IRE1/XBP1, which is a cellular stress response associated with the endoplasmic reticulum (ER), is involved in the bovine CL life span. Our results indicated that expression of Grp78/Bip, the master UPR regulator, was increased during the maintenance stage and rapidly decreased at the regression stage. Additionally, UPR signaling pathways genes were found to be involved in luteal phase progression during the estrous cycle. Our findings suggested that Grp78/Bip, ATF6, and XBP1 act as ER chaperones for initiating CL development and maintaining the CL. In addition, we investigated whether ER stress-mediated apoptosis is occurred through three UPR signaling pathways in CL regression stage. Interestingly, pIRE1 and CHOP were found to be involved in both the adaptive response and ER stress-mediated apoptosis. During the CL regression stage, increased expression of pJNK and CHOP, two components of ER stress-mediated apoptotic cascades, occurred before increased level of cleaved caspase 3 were observed. The present investigation was performed to identify a functional link between UPR signaling and CL life span during the bovine estrous cycle. Taken together, results from this study demonstrated that UPR protein/gene expression levels were different at various stages of the bovine CL life span. Variations in the expression of these protein/genes may play important roles in luteal stage progression during the estrous cycle. PMID:24161737

  2. Eliminating bovine tuberculosis in cattle and badgers: insights from a dynamic model

    E-print Network

    Brooks-Pollock, Ellen; Wood, James L. N.

    2015-05-13

    Bovine tuberculosis (BTB) is a multi-species infection that commonly affects cattle and badgers in Great Britain. Despite years of study, the impact of badgers on BTB incidence in cattle is poorly understood. Using a two-host transmission model...

  3. Protein Hydrolysates from Non-bovine and Plant Sources Replaces Tryptone in Microbiological Media

    Microsoft Academic Search

    Yamini Ranganathan; Shifa Patel; Vijai K. Pasupuleti; R. Meganathan

    2010-01-01

    \\u000a Tryptone (pancreatic digest of casein) is a common ingredient in laboratory and fermentation media for growing wild-type and\\u000a genetically modified microorganisms. Many of the commercially manufactured products such as human growth hormone, antibiotics,\\u000a insulin, etc. are produced by recombinant strains grown on materials derived from bovine sources. With the emergence of Bovine\\u000a Spongiform Encephalopathy (BSE) and the consequent increase in

  4. Effects of electrical stimulation on the myofibril protein degradation and fragmentation in bovine longissimus dorsi muscle and its relationship to tenderness 

    E-print Network

    Procknor, Maria Lucia Salles Cunha

    1984-01-01

    EFFECTS OF ELECTRICAL STIMULATION ON THE MYOFIBRIL PROTEIN DEGRADATION AND FRAGMENTATION IN BOVINE LONGISSIMUS DORSI MUSCLE AND ITS RELATIONSHIP TO TENDERNESS A Thesis by Maria Lucia Salles Cunha Procknor Submitted to the Graduate College... IN BOVINE LONGISSIMUS DORSI MUSCLE AND ITS RELATIONSHIP TO TENDERNESS A Thesis by MARIA LUCIA SALLES CUNHA PROCKNOR Approved as to style and content by: Thayne R. Dutson (Co-Chairman) Jeffrey W. Savell (Co-Chairman) Konrad A. Eugster (Member...

  5. Leucine and histidine independently regulate milk protein synthesis in bovine mammary epithelial cells via mTOR signaling pathway.

    PubMed

    Gao, Hai-Na; Hu, Han; Zheng, Nan; Wang, Jia-Qi

    2015-06-01

    The aim of this study is to investigate the effects of leucine (Leu) and histidine (His) on the expression of both the mammalian target of rapamycin (mTOR) signaling pathway-related proteins and caseins in immortalized bovine mammary epithelial cells (CMEC-H), using a single supplement through Western blotting. The Earle's balanced salt solution (EBSS) was set as the control group and other treatment groups, based on the EBSS, were added with different concentrations of Leu or His, respectively. The results showed that, compared with the control group, the expression of caseins and the phosphorylation of mTOR (Ser(2481)), Raptor (Ser(792)), eIF4E (Ser(209)), and eEF2 (Thr(56)) increased with the Leu concentrations ranging from 0.45 to 10.80 mmol/L (P<0.01). The P-4EBP1 (Thr(37)) at 10.80 mmol/L Leu, and P-RPS6 (Ser(235/236)) at 5.40 to 10.80 mmol/L Leu all decreased. Similarly, the His supplementation from 0.15 to 9.60 mmol/L increased the expression of ?s2-casein, ?-casein, ?-casein, P-mTOR (Ser(2481)), P-Raptor (Ser(792)), P-S6K1 (Thr(389)), P-4EBP1 (Thr(37)), P-eIF4E (Ser(209)), and P-eEF2 (Thr(56)) (P<0.01) in CMEC-H, whereas the ?s1-casein expression was only reduced at 9.60 mmol/L His, G protein ? subunit-like protein (G?L) at 0.15 and 9.60 mmol/L His, and P-RPS6 at 4.80 to 9.60 mmol/L His. Our linear regression model assay suggested that the ?s1-casein expression was positively correlated with P-mTOR (P<0.01), P-S6K1 (P<0.01), and P-eEF2 (P<0.01) for the addition of Leu, while the expressions of ?-casein (P<0.01) and ?-casein (P<0.01) were positively correlated with P-eEF2 for the addition of His. In conclusion, the milk protein synthesis was up-regulated through activation of the mTOR pathway with the addition of Leu and His in CMEC-H. PMID:26055918

  6. Leucine and histidine independently regulate milk protein synthesis in bovine mammary epithelial cells via mTOR signaling pathway*

    PubMed Central

    Gao, Hai-na; Hu, Han; Zheng, Nan; Wang, Jia-qi

    2015-01-01

    The aim of this study is to investigate the effects of leucine (Leu) and histidine (His) on the expression of both the mammalian target of rapamycin (mTOR) signaling pathway-related proteins and caseins in immortalized bovine mammary epithelial cells (CMEC-H), using a single supplement through Western blotting. The Earle’s balanced salt solution (EBSS) was set as the control group and other treatment groups, based on the EBSS, were added with different concentrations of Leu or His, respectively. The results showed that, compared with the control group, the expression of caseins and the phosphorylation of mTOR (Ser2481), Raptor (Ser792), eIF4E (Ser209), and eEF2 (Thr56) increased with the Leu concentrations ranging from 0.45 to 10.80 mmol/L (P<0.01). The P-4EBP1 (Thr37) at 10.80 mmol/L Leu, and P-RPS6 (Ser235/236) at 5.40 to 10.80 mmol/L Leu all decreased. Similarly, the His supplementation from 0.15 to 9.60 mmol/L increased the expression of ?s2-casein, ?-casein, ?-casein, P-mTOR (Ser2481), P-Raptor (Ser792), P-S6K1 (Thr389), P-4EBP1 (Thr37), P-eIF4E (Ser209), and P-eEF2 (Thr56) (P<0.01) in CMEC-H, whereas the ?s1-casein expression was only reduced at 9.60 mmol/L His, G protein ? subunit-like protein (G?L) at 0.15 and 9.60 mmol/L His, and P-RPS6 at 4.80 to 9.60 mmol/L His. Our linear regression model assay suggested that the ?s1-casein expression was positively correlated with P-mTOR (P<0.01), P-S6K1 (P<0.01), and P-eEF2 (P<0.01) for the addition of Leu, while the expressions of ?-casein (P<0.01) and ?-casein (P<0.01) were positively correlated with P-eEF2 for the addition of His. In conclusion, the milk protein synthesis was up-regulated through activation of the mTOR pathway with the addition of Leu and His in CMEC-H. PMID:26055918

  7. Bovine leukemia virus nucleocapsid protein is an efficient nucleic acid chaperone.

    PubMed

    Qualley, Dominic F; Sokolove, Victoria L; Ross, James L

    2015-03-13

    Nucleocapsid proteins (NCs) direct the rearrangement of nucleic acids to form the most thermodynamically stable structure, and facilitate many steps throughout the life cycle of retroviruses. NCs bind strongly to nucleic acids (NAs) and promote NA aggregation by virtue of their cationic nature; they also destabilize the NA duplex via highly structured zinc-binding motifs. Thus, they are considered to be NA chaperones. While most retroviral NCs are structurally similar, differences are observed both within and between retroviral genera. In this work, we compare the NA binding and chaperone activity of bovine leukemia virus (BLV) NC to that of two other retroviral NCs: human immunodeficiency virus type 1 (HIV-1) NC, which is structurally similar to BLV NC but from a different retrovirus genus, and human T-cell leukemia virus type 1 (HTLV-1) NC, which possesses several key structural differences from BLV NC but is from the same genus. Our data show that BLV and HIV-1 NCs bind to NAs with stronger affinity in relation to HTLV-1 NC, and that they also accelerate the annealing of complementary stem-loop structures to a greater extent. Analysis of kinetic parameters derived from the annealing data suggests that while all three NCs stimulate annealing by a two-step mechanism as previously reported, the relative contributions of each step to the overall annealing equilibrium are conserved between BLV and HIV-1 NCs but are different for HTLV-1 NC. It is concluded that while BLV and HTLV-1 belong to the same genus of retroviruses, processes that rely on NC may not be directly comparable. PMID:25686502

  8. The glass transition behavior of the globular protein bovine serum albumin.

    PubMed

    Brownsey, Geoffrey J; Noel, Timothy R; Parker, Roger; Ring, Stephen G

    2003-12-01

    The glass-like transition behavior of concentrated aqueous solutions of bovine serum albumin was examined using rheological techniques. At mass fractions >0.4, there was a marked concentration dependence of viscosity with a glass-like kinetic arrest observed at mass fractions in the region of 0.55. At mass fractions >0.6 the material behaved as a solid with a Young's modulus rising from approximately 20 MPa at a mass fraction of 0.62-1.1 GPa at 0.86. The solid was viscoelastic and exhibited stress relaxation with relaxation times increasing from 33 to 610 s over the same concentration range. The concentration dependence of the osmotic pressure was measured, at intermediate concentrations, using an osmotic stress technique and could be described using a hard sphere model, indicating that the intermolecular interactions were predominantly repulsive. In summary, a major structural relaxation results from the collective motion of the globules at the supra-globule length scale and, at 20 degrees C, this is arrested at water contents of 40% w/w. This appears to be analogous to the glass transition in colloidal hard spheres. PMID:14645083

  9. A model of the spread of the bovine viral-diarrhoea virus within a dairy herd

    Microsoft Academic Search

    Anne-France Viet; Christine Fourichon; Henri Seegers; Christine Jacob; Chantal Guihenneuc-Jouyaux

    2004-01-01

    Wet BVDSim (a stochastic simulation model) was developed to study the dynamics of the spread of the bovine viral-diarrhoea virus (BVDV) within a dairy herd. This model took into account herd-management factors (common in several countries), which influence BVDV spread. BVDSim was designed as a discrete-entity and discrete-event simulation model. It relied on two processes defined at the individual-animal level,

  10. Metal Ion and Guanine Nucleotide Modulations of Agonist Interaction in G-Protein-Coupled Serotonin 1A Receptors from Bovine Hippocampus

    Microsoft Academic Search

    K. G. Harikumar; Amitabha Chattopadhyay

    1998-01-01

    1. The serotonin type 1A (5-HT1A) receptors are members of a superfamily of seven transmembrane domain receptors that couple to GTP-binding regulatory proteins (G-proteins). We have studied the modulation of agonist binding to 5-HT1A receptors from bovine hippocampus by metal ions and guanine nucleotide.

  11. The Positively Charged Termini of L2 Minor Capsid Protein Required for Bovine Papillomavirus Infection Function Separately in Nuclear Import and DNA Binding

    Microsoft Academic Search

    Alyson Fay; William H. Yutzy; Richard B. S. Roden; Junona Moroianu

    2004-01-01

    During the papillomavirus (PV) life cycle, the L2 minor capsid protein enters the nucleus twice: in the initial phase after entry of virions into cells and in the productive phase to mediate encapsidation of the newly replicated viral genome. Therefore, we investigated the interactions of the L2 protein of bovine PV type 1 (BPV1) with the nuclear import machinery and

  12. Polymer-Induced Heteronucleation for Protein Single Crystal Growth: Structural Elucidation of Bovine Liver Catalase and Concanavalin A Forms

    SciTech Connect

    Foroughi, Leila M.; Kang, You-Na; Matzger, Adam J. (Michigan)

    2012-05-09

    Obtaining single crystals for X-ray diffraction remains a major bottleneck in structural biology; when existing crystal growth methods fail to yield suitable crystals, often the target rather than the crystallization approach is reconsidered. Here we demonstrate that polymer-induced heteronucleation, a powerful technique that has been used for small molecule crystallization form discovery, can be applied to protein crystallization by optimizing the heteronucleant composition and crystallization formats for crystallizing a wide range of protein targets. Applying these advances to two benchmark proteins resulted in dramatically increased crystal size, enabling structure determination, for a half century old form of bovine liver catalase (BLC) that had previously only been characterized by electron microscopy, and the discovery of two new forms of concanavalin A (conA) from the Jack bean and accompanying structural elucidation of one of these forms.

  13. A Bovine Model of Respiratory Chlamydia psittaci Infection: Challenge Dose Titration

    PubMed Central

    Reinhold, Petra; Ostermann, Carola; Liebler-Tenorio, Elisabeth; Berndt, Angela; Vogel, Anette; Lambertz, Jacqueline; Rothe, Michael; Rüttger, Anke; Schubert, Evelyn; Sachse, Konrad

    2012-01-01

    This study aimed to establish and evaluate a bovine respiratory model of experimentally induced acute C. psittaci infection. Calves are natural hosts and pathogenesis may resemble the situation in humans. Intrabronchial inoculation of C. psittaci strain DC15 was performed in calves aged 2–3 months via bronchoscope at four different challenge doses from 106 to 109 inclusion-forming units (ifu) per animal. Control groups received either UV-inactivated C. psittaci or cell culture medium. While 106 ifu/calf resulted in a mild respiratory infection only, the doses of 107 and 108 induced fever, tachypnea, dry cough, and tachycardia that became apparent 2–3 days post inoculation (dpi) and lasted for about one week. In calves exposed to 109 ifu C. psittaci, the respiratory disease was accompanied by severe systemic illness (apathy, tremor, markedly reduced appetite). At the time point of most pronounced clinical signs (3 dpi) the extent of lung lesions was below 10% of pulmonary tissue in calves inoculated with 106 and 107 ifu, about 15% in calves inoculated with 108 and more than 30% in calves inoculated with 109 ifu C. psittaci. Beside clinical signs and pathologic lesions, the bacterial load of lung tissue and markers of pulmonary inflammation (i.e., cell counts, concentration of proteins and eicosanoids in broncho-alveolar lavage fluid) were positively associated with ifu of viable C. psittaci. While any effect of endotoxin has been ruled out, all effects could be attributed to infection by the replicating bacteria. In conclusion, the calf represents a suitable model of respiratory chlamydial infection. Dose titration revealed that both clinically latent and clinically manifest infection can be reproduced experimentally by either 106 or 108 ifu/calf of C. psittaci DC15 while doses above 108 ifu C. psittaci cannot be recommended for further studies for ethical reasons. This defined model of different clinical expressions of chlamydial infection allows studying host-pathogen interactions. PMID:22299031

  14. Cryopreservation for bovine embryos in serum-free freezing medium containing silk protein sericin.

    PubMed

    Isobe, Tomohiro; Ikebata, Yoshihisa; Onitsuka, Takeshi; Do, Lanh Thi Kim; Sato, Yoko; Taniguchi, Masayasu; Otoi, Takeshige

    2013-10-01

    Because the use of serum in the embryo cryopreservation increases the probability of animal health problems such as bovine spongiform encephalopathy (BSE) and viral infections, this study was conducted to examine the effects of sericin supplementation for serum-free freezing medium on the survival and development of bovine embryos after freezing-thawing and direct transfer to recipients. When in vitro-produced bovine embryos were frozen conventionally in the freezing medium supplemented with various concentrations (0.1%, 0.5%, and 1.0%) of sericin, the percentages of damaged zona pellucida, survival, and development of frozen-thawed embryos were similar to those of embryos frozen in freezing medium supplemented with 0.4% bovine serum albumin (BSA) and 20% fetal bovine serum (FBS) (0.4BSA/20F; control). When in vivo-derived embryos were frozen with 0.4BSA/20F (control), 0.5% sericin +20% FBS (0.5S/20F) or 0.5% sericin (0.5S) and were subsequently transferred directly to recipients, the percentages of recipients with pregnancy and normal calving in the 0.5S/20F group were higher than those in the control group (47.3% vs. 40.1% and 94.6% vs. 87.3%, respectively). Moreover, the percentages of recipients with pregnancy and normal calving (42.2% and 92.4%, respectively) in the 0.5S group were similar with those of other groups. In conclusion, these results indicate that serum-free freezing medium supplemented with sericin is available for the cryopreservation of bovine embryos and that it is beneficial for the elimination of a potential source of biological contamination by serum or BSA. PMID:23850826

  15. A guinea pig model of bovine pneumonic pasteurellosis.

    PubMed Central

    Morck, D W; Costerton, J W; Bolingbroke, D O; Ceri, H; Boyd, N D; Olson, M E

    1990-01-01

    The induction of pneumonic pasteurellosis in guinea pigs (Cavia porcellus) was examined. Specific pathogen free male guinea pigs were anesthetized and a tracheostomy performed to introduce 10(5), 10(4) or 10(3) Pasteurella haemolytica-A1 into the left principal bronchus. The surgical site was closed with tissue adhesive and staples and the animals were monitored for signs of respiratory tract infection. Within 24 hours after inoculation they became depressed, anorectic, pyretic and dyspneic. Fibrinous pleuropneumonia with prominent areas of necrosis and hemorrhage was present. Pericardial effusion was a frequent finding. There was infiltration of the pleura and alveoli with degenerate heterophils and macrophages, a hyperplastic mesothelium and fibrin exudation on the pleura and within alveoli. Hemorrhage, congestion, consolidation, edema and fibrin exudation were prominent in the hilar region of the lungs. Bacterial colonies were evident in all airways. More bacteria were recovered from infected lungs than were inoculated (p less than 0.05) indicating P. haemolytica was actively multiplying in the lungs. Hematological and clinical chemistry data were consistent with fibrinous pneumonia, however, blood cultures were positive for P. haemolytica in 61% (11/18) of animals sampled. Examination of pneumonic pasteurellosis in guinea pigs may be useful in studying pathogenetic and pathological features applicable to bovine pneumonic pasteurellosis (shipping fever pneumonia). Images Fig. 1. Fig. 2. Fig. 4. Fig. 5. Fig. 7. Fig. 8. Fig. 9. PMID:2306663

  16. Bovine NK Cells Can Produce Gamma Interferon in Response to the Secreted Mycobacterial Proteins ESAT-6 and MPP14 but Not in Response to MPB70

    Microsoft Academic Search

    Ingrid Olsen; Preben Boysen; Siri Kulberg; Jayne C. Hope; Gregers Jungersen; Anne K. Storset

    2005-01-01

    Bovine NK cells have recently been characterized and the present study describes the interaction between NK cells, antigen-presenting cells, and secreted mycobacterial proteins. Gamma interferon (IFN-) production by NK cells was seen in approximately 30% of noninfected calves in response to the Mycobacterium tuberculosis complex-specific protein ESAT-6, MPP14 from Mycobacterium avium subsp. paratuberculosis, and purified protein derivative (PPD) from M.

  17. Variation in the bovine FABP4 gene affects milk yield and milk protein content in dairy cows.

    PubMed

    Zhou, H; Cheng, L; Azimu, W; Hodge, S; Edwards, G R; Hickford, J G H

    2015-01-01

    Fatty acid binding proteins (FABPs) bind long-chain fatty acids and are involved in their intracellular transport. Of the known bovine FABP genes, FABP4 has been mapped to a region on chromosome 14 that contains quantitative trait loci for milk traits. This study investigated the association of FABP4 haplotypes with milk production traits in 719 Holstein-Friesian?×?Jersey cows. Polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis of a variable region of the gene revealed three haplotypes (A, B and C). Five single nucleotide polymorphisms (SNPs) were identified: two in exon 3 and three in intron 3. A was associated (P?=?0.032) with increased milk protein percentage (present: 4.00?±?0.02%; absent: 3.95?±?0.02%) and B was associated (P?=?0.009) with increased milk yield (present: 23.81?±?0.23?kg/d; absent: 23.06?±?0.21?kg/d), but tended to be associated with a decrease in protein percentage and an increase in protein yield. Cows with genotypes AA, AB and AC produced less milk, but with a higher protein percentage than BC cows. This suggest that FABP4 affects milk yield and milk protein content, both economically important traits, and that further study of this gene is warranted. PMID:26067182

  18. Variation in the bovine FABP4 gene affects milk yield and milk protein content in dairy cows

    PubMed Central

    Zhou, H.; Cheng, L.; Azimu, W.; Hodge, S.; Edwards, G. R.; Hickford, J. G. H.

    2015-01-01

    Fatty acid binding proteins (FABPs) bind long-chain fatty acids and are involved in their intracellular transport. Of the known bovine FABP genes, FABP4 has been mapped to a region on chromosome 14 that contains quantitative trait loci for milk traits. This study investigated the association of FABP4 haplotypes with milk production traits in 719 Holstein-Friesian?×?Jersey cows. Polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis of a variable region of the gene revealed three haplotypes (A, B and C). Five single nucleotide polymorphisms (SNPs) were identified: two in exon 3 and three in intron 3. A was associated (P?=?0.032) with increased milk protein percentage (present: 4.00?±?0.02%; absent: 3.95?±?0.02%) and B was associated (P?=?0.009) with increased milk yield (present: 23.81?±?0.23?kg/d; absent: 23.06?±?0.21?kg/d), but tended to be associated with a decrease in protein percentage and an increase in protein yield. Cows with genotypes AA, AB and AC produced less milk, but with a higher protein percentage than BC cows. This suggest that FABP4 affects milk yield and milk protein content, both economically important traits, and that further study of this gene is warranted. PMID:26067182

  19. Electrolyte effect on the phase behavior of silica nanoparticles with lysozyme and bovine-serum-albumin proteins

    NASA Astrophysics Data System (ADS)

    Yadav, Indresh; Aswal, V. K.; Kohlbrecher, J.

    2015-05-01

    Small-angle neutron scattering (SANS) and dynamic light scattering (DLS) studies have been carried out to investigate the effect of an electrolyte on the phase behavior of anionic silica nanoparticles with two globular proteins—cationic lysozyme [molecular weight (MW) 14.7 kDa] and anionic bovine serum albumin (MW 66.4 kDa). The results are compared with our earlier published work on similar systems without any electrolyte [I. Yadav, S. Kumar, V. K. Aswal, and J. Kohlbrecher, Phys. Rev. E 89, 032304 (2014), 10.1103/PhysRevE.89.032304]. Both the nanoparticle-protein systems transform to two phase at lower concentration of protein in the presence of an electrolyte. The autocorrelation function in DLS suggests that the diffusion coefficient (D) of a nanoparticle-protein system decreases in approaching two phase with the increase in protein concentration. This variation in D can be attributed to increase in attractive interaction and/or overall increase in the size. Further, these two contributions (interaction and structure) are determined from the SANS data. The changes in the phase behavior of nanoparticle-protein systems in the presence of an electrolyte are explained in terms of modifications in both the repulsive and attractive components of interaction between nanoparticles. In a two-phase system individual silica nanoparticles coexist along with their fractal aggregates.

  20. Quantitative and qualitative analysis of cellular prion protein (PrP(C)) expression in bovine somatic tissues.

    PubMed

    Peralta, Oscar A; Eyestone, Willard H

    2009-01-01

    The host encoded cellular prion protein (PrP(C)) is an N-linked glycoprotein tethered to the cell membrane by a glycophosphatidylinositol (GPI) anchor. Under certain conditions, PrP(C) can undergo conversion into a conformationally-altered isoform (PrP(Sc)) widely believed to be the pathogenic agent of transmissible spongiform encephalopathies (TSEs). Understanding the tissue-specific expression of PrP(C) is crucial considering that cells expressing high levels of PrP(C) bear a risk for conversion and accumulation of PrP(Sc). In the present study, fifteen bovine somatic tissues were analyzed for PrP(C) expression by quantitative western blot and immunohistochemistry. Quantitative western blot analysis revealed highest expression of PrP(C) in cerebellum, obex and spinal cord. Intermediate levels were detected in thymus, intestine, nerve, heart and spleen, and lower levels in lung, muscle, kidney, lymph node, skin, pancreas and liver. Immunohistochemical analysis detected intense cellular-specific PrP(C) staining in neurons, thymocytes and lymphocytes. PrP(C) was also detected in the enteric wall, pancreatic islets of langerhans, myocardium, pulmonary alveolar sacs, renal glomeruli and dermal epithelial cells. This study demonstrated the quantitatively varied, wide-spread, tissue- and cell-specific expression pattern of PrP(C) in bovine somatic tissues. The importance of this study is to lay the foundation for understanding the tissue-specific expression of PrP(C) and to consider the potential participation of more bovine tissues in the transmission of BSE infection. PMID:19806026

  1. Viscomagnetic Study of Bovine Serum Albumin and Egg Albumin Proteins in Aqueous Solutions

    Microsoft Academic Search

    Man Singh

    2007-01-01

    Intrinsic viscosities [?] for 0.2, 0.4, 0.6, and 0.8 mg% aqueous of each Bovine serum albumin (BSA) and egg albumin (EA) solutions under zero, 0.5 and 0.7 magnetic flux at 298.15 K and 1 atmosphere are reported. The [?] data for BSA are higher than EA and values of both increase with magnetic flux strengthening molecular interactions.

  2. Effects of embryo size at transfer (whole versus demi) and early pregnancy progesterone supplementation on embryo growth and pregnancy-specific protein bovine concentrations in recipient dairy heifers

    Microsoft Academic Search

    L. Lopes-da-Costa; J. Chagas e Silva; M. C. Deloche; N. Jeanguyot; P. Humblot; A. E. M. Horta

    2011-01-01

    The objectives of this study were to evaluate embryonic size and survival, plasma progesterone (P4) and pregnancy-specific protein bovine (PSPB) concentrations in early pregnancies (n = 99) following the transfer of one whole (n = 66) or one demi (n = 33) embryo to recipient virgin dairy heifers. The experiment was designed to evaluate the fixed effects of embryo size

  3. Expression of the bovine viral diarrhoea virus Osloss p80 protein: its use as ELISA antigen for cattle serum antibody detection

    Microsoft Academic Search

    Nathalie Vanderheijden; L. De Moerlooze; D. Vandenbergh; G. Chappuis; A. Renard; C. Lecomte

    1993-01-01

    The putative gene encoding the cytopathic bovine viral diarrhoea virus (BVDV) Osloss strain p80 protein was amplified by PCR and inserted into a T7 promoter- based vector for expression in Escherichia coli. Bacterial expression led to cytoplasmic insoluble inclusion bodies which were denatured by urea treatment and renatured by dialysis. Rabbit antisera were raised against this p80 recombinant antigen and

  4. Comparative Protein Structure Modeling Using Modeller

    PubMed Central

    Eswar, Narayanan; Marti-Renom, Marc A.; Madhusudhan, M.S.; Eramian, David; Shen, Min-yi; Pieper, Ursula

    2014-01-01

    Functional characterization of a protein sequence is one of the most frequent problems in biology. This task is usually facilitated by accurate three-dimensional (3-D) structure of the studied protein. In the absence of an experimentally determined structure, comparative or homology modeling can sometimes provide a useful 3-D model for a protein that is related to at least one known protein structure. Comparative modeling predicts the 3-D structure of a given protein sequence (target) based primarily on its alignment to one or more proteins of known structure (templates). The prediction process consists of fold assignment, target-template alignment, model building, and model evaluation. This unit describes how to calculate comparative models using the program MODELLER and discusses all four steps of comparative modeling, frequently observed errors, and some applications. Modeling lactate dehydrogenase from Trichomonas vaginalis (TvLDH) is described as an example. The download and installation of the MODELLER software is also described. PMID:18428767

  5. Studies on substantially increased proteins in follicular fluid of bovine ovarian follicular cysts using 2-D PAGE and MALDI-TOF MS

    PubMed Central

    Maniwa, Jiro; Izumi, Shunsuke; Isobe, Naoki; Terada, Takato

    2005-01-01

    Background The objective of this study was to identify substantially increased proteins in bovine cystic follicular fluid (FF) in order to clarify the pathology and etiology of bovine ovarian follicular cysts (BOFC). Methods Proteins in normal and cystic FF samples were subjected to two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and were compared using silver stained gel images with PDQuest image analysis software. Peptides from these increased spots were analyzed by matrix assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), and were identified based on the NCBI database by a peptide mass fingerprinting method. Results Comparative proteomic analysis showed 8 increased protein spots present in cystic FF. MS analysis and database searching revealed that the increased proteins in cystic FF were bovine mitochondrial f1-atpase (BMFA), erythroid associated factor (EAF), methionine synthase (MeS), VEGF-receptor, glyceraldehydes 3-phosphate dehydrogenase (GAPDH), heat shock protein 70 (HSP70), ?-lactoglobulin (BLG) and succinate dehydrogenase Ip subunit (SD). Conclusion Our results suggest that these proteins are overexpressed in BOFC, and that they may play important roles in the pathogenesis of BOFC. Furthermore, these proteins in the FF could be useful biomarkers for BOFC. PMID:15941490

  6. Improvement of gluten-free bread properties by the incorporation of bovine plasma proteins and different saccharides into the matrix.

    PubMed

    Rodriguez Furlán, Laura T; Pérez Padilla, Antonio; Campderrós, Mercedes E

    2015-03-01

    The aim of this work was to improve the quality of gluten-free bread, incorporating plasma bovine proteins concentrated by ultrafiltration and freeze-dried with saccharides (inulin and sucrose). The influence of these compounds on textural properties and final bread quality was assessed. The textural studies revealed that with the addition of proteins and inulin, homogeneous and smaller air cells were achieved improving the textural properties while the bread hardness was comparable with breads with gluten. The volume of gluten-free breads increased with increasing proteins and inulin concentrations, reaching a maximum at a protein concentration of 3.5% (w/w). The addition of the enhancers improved moisture retention of the loaves after cooking and an increase of lightness of crumb with respect to the control was observed. The sensory analysis found no statistically significant difference in sensory attributes evaluated with respect to the control, so these ingredients do not negatively affect the organoleptic properties of bread. PMID:25306343

  7. Evaluation of Multiplexed Foot-and-Mouth Disease Nonstructural Protein Antibody Assay Against Standardized Bovine Serum Panel

    SciTech Connect

    Perkins, J; Parida, S; Clavijo, A

    2007-05-14

    Liquid array technology has previously been used to show proof-of-principle of a multiplexed non structural protein serological assay to differentiate foot-and-mouth infected and vaccinated animals. The current multiplexed assay consists of synthetically produced peptide signatures 3A, 3B and 3D and recombinant protein signature 3ABC in combination with four controls. To determine diagnostic specificity of each signature in the multiplex, the assay was evaluated against a naive population (n = 104) and a vaccinated population (n = 94). Subsequently, the multiplexed assay was assessed using a panel of bovine sera generated by the World Reference Laboratory for foot-and-mouth disease in Pirbright, UK. This sera panel has been used to assess the performance of other singleplex ELISA-based non-structural protein antibody assays. The 3ABC signature in the multiplexed assay showed comparative performance to a commercially available non-structural protein 3ABC ELISA (Cedi test{reg_sign}) and additional information pertaining to the relative diagnostic sensitivity of each signature in the multiplex is acquired in one experiment. The encouraging results of the evaluation of the multiplexed assay against a panel of diagnostically relevant samples promotes further assay development and optimization to generate an assay for routine use in foot-and-mouth disease surveillance.

  8. Protein crystal growth in microgravity review of large scale temperature induction method: Bovine insulin, human insulin and human ?-interferon

    NASA Astrophysics Data System (ADS)

    Long, Marianna M.; Bishop, John Bradford; Delucas, Lawrence J.; Nagabhushan, Tattanhalli L.; Reichert, Paul; Smith, G. David

    1997-01-01

    The Protein Crystal Growth Facility (PCF) is space-flight hardware that accommodates large scale protein crystal growth experiments using temperature change as the inductive step. Recent modifications include specialized instrumentation for monitoring crystal nucleation with laser light scattering. This paper reviews results from its first seven flights on the Space Shuttle, the last with laser light scattering instrumentation in place. The PCF's objective is twofold: (1) the production of high quality protein crystals for x-ray analysis and subsequent structure-based drug design and (2) preparation of a large quantity of relatively contaminant free crystals for use as time-release protein pharmaceuticals. The first three Shuttle flights with bovine insulin constituted the PCF's proof of concept, demonstrating that the space-grown crystals were larger and diffracted to higher resolution than their earth-grown counterparts. The later four PCF missions were used to grow recombinant human insulin crystals for x-ray analysis and continue productions trials aimed at the development of a processing facility for crystalline recombinant a-interferon.

  9. Bone enhancing effect of titanium-binding proteins isolated from bovine bone and implanted into rat calvaria with titanium scaffold.

    PubMed

    Kuboki, Yoshinori; Furusawa, Toshitake; Sato, Masaaki; Sun, Yongkun; Unuma, Hidero; Abe, Shigeaki; Fujisawa, Ryuichi; Akasaka, Tsukasa; Watari, Fumio; Takita, Hiroko; Sammons, Rachel

    2014-01-01

    Based on our previous finding that a chromatography with titanium beads selectively binds phosphoproteins, including caseins, phosvitin and dentin phosphoproteins, we investigated whether bone phosphoproteins also bind to titanium. Bovine bone matrix proteins were extracted with 2 M urea/PBS after demineralization. The 2 M urea extract was directly applied to the titanium chromatography column as reported. The chromatogram showed an initial large peak at breakthrough position (non-binding fraction) and a smaller second peak eluted later (titanium-binding fraction). Both peaks were analyzed by SDS polyacrylamide gel electrophoresis. Stains-all staining which preferentially identifies phospho-proteins revealed that the first peak contained no positively stained band, while the second peak showed 4 or 5 distinctive bands indicative of bone phosphoproteins. To investigate the biological functions of the titanium-binding bone proteins (TiBP), we implanted them into calvaria of rats, combined with titanium web (TW), a highly porous titanium scaffold of thin titanium-fibers. Bone TiBP induced significantly enhanced bone formation, and new bone appeared connected directly to titanium fibers, accompanied by active blood vessel formations. Control TW alone did not induce bone formation within the titanium framework. These results demonstrate that the bone titanium-binding proteins include phosphoproteins which enhance bone formation when implanted into bone with titanium. PMID:24840192

  10. A novel in vitro bovine cartilage punch model for assessing the regeneration of focal cartilage defects with biocompatible bacterial nanocellulose

    PubMed Central

    2013-01-01

    Introduction Current therapies for articular cartilage defects fail to achieve qualitatively sufficient tissue regeneration, possibly because of a mismatch between the speed of cartilage rebuilding and the resorption of degradable implant polymers. The present study focused on the self-healing capacity of resident cartilage cells in conjunction with cell-free and biocompatible (but non-resorbable) bacterial nanocellulose (BNC). This was tested in a novel in vitro bovine cartilage punch model. Methods Standardized bovine cartilage discs with a central defect filled with BNC were cultured for up to eight weeks with/without stimulation with transforming growth factor-?1 (TGF-?1. Cartilage formation and integrity were analyzed by histology, immunohistochemistry and electron microscopy. Content, release and neosynthesis of the matrix molecules proteoglycan/aggrecan, collagen II and collagen I were also quantified. Finally, gene expression of these molecules was profiled in resident chondrocytes and chondrocytes migrated onto the cartilage surface or the implant material. Results Non-stimulated and especially TGF-?1-stimulated cartilage discs displayed a preserved structural and functional integrity of the chondrocytes and surrounding matrix, remained vital in long-term culture (eight weeks) without signs of degeneration and showed substantial synthesis of cartilage-specific molecules at the protein and mRNA level. Whereas mobilization of chondrocytes from the matrix onto the surface of cartilage and implant was pivotal for successful seeding of cell-free BNC, chondrocytes did not immigrate into the central BNC area, possibly due to the relatively small diameter of its pores (2 to 5 ?m). Chondrocytes on the BNC surface showed signs of successful redifferentiation over time, including increase of aggrecan/collagen type II mRNA, decrease of collagen type I mRNA and initial deposition of proteoglycan and collagen type II in long-term high-density pellet cultures. Although TGF-?1 stimulation showed protective effects on matrix integrity, effects on other parameters were limited. Conclusions The present bovine cartilage punch model represents a robust, reproducible and highly suitable tool for the long-term culture of cartilage, maintaining matrix integrity and homoeostasis. As an alternative to animal studies, this model may closely reflect early stages of cartilage regeneration, allowing the evaluation of promising biomaterials with/without chondrogenic factors. PMID:23673274

  11. Expression of SULT1E1 protein in bovine placentomes: Evidence for localization in uninucleated trophoblast cells

    Microsoft Academic Search

    P. Khatri; G. Frenette; R. Sullivan; B. Hoffmann; G. Schuler

    2011-01-01

    The bovine placenta produces considerable amounts of pregnancy-associated estrogens, predominantly estrone sulfate, which does not interact with classical nuclear estrogen receptors. Thus, bovine placental estrogens may rather act as local regulatory factors than as hormones in a classical sense. To obtain information on the local availability of free estrogens in bovine placentomes, the expression pattern of the estrogen specific sulfotransferase

  12. Perfusion-Cultured Bovine Anterior Segments as an Ex Vivo Model for Studying Glucocorticoid-Induced Ocular Hypertension and Glaucoma

    PubMed Central

    Tovar-Vidales, Tara; Yorio, Thomas; Wordinger, Robert J.; Clark, Abbot F.

    2011-01-01

    Purpose. To determine whether perfusion-cultured bovine anterior segments would be a suitable model for glaucoma research. Methods. Fresh bovine eyes were dissected and sealed on a custom-made acrylic dish with an O-ring. Perfusion medium was infused by a syringe pump at a constant infusion rate of 5 ?L/min. After intraocular pressure (IOP) was stable, bovine eyes were perfused with medium containing either a vehicle control (0.1% ethanol [ETH]) or dexamethasone (DEX) for up to 7 days. IOP was recorded by a pressure transducer and a computerized system. Perfusion medium was collected for Western immunoblot analysis of myocilin (MYOC). Results. The morphology of the bovine trabecular meshwork after perfusion culture was similar to that of freshly dissected, nonperfused bovine eyes. Treatment with DEX elevated IOP in some bovine eyes, whereas others showed little change. The authors analyzed the data from 18 ETH-treated control eyes and defined 2.82 mm Hg as the threshold of ocular hypertension (OHT), which equals mean pressure change + 2× SD. Approximately 40% (12/29) of the bovine eyes were DEX responders, which is very close to the DEX-responsive rates observed in human and monkey eyes. Western blot data showed that DEX treatment induced the expression of the DEX-inducible gene MYOC only in the perfusion-cultured anterior segments with DEX-induced OHT. Conclusions. OHT can be induced by DEX in perfusion-cultured bovine anterior segments. This is a fast, convenient, affordable, and reliable model for studying DEX-induced OHT and the mechanisms of trabecular outflow. PMID:21911581

  13. Spectroscopic probe analysis for exploring probe-protein interaction: a mapping of native, unfolding and refolding of protein bovine serum albumin by extrinsic fluorescence probe.

    PubMed

    Samanta, Anuva; Paul, Bijan Kumar; Guchhait, Nikhil

    2011-07-01

    Steady state and dynamic fluorescence measurements have been used to investigate interaction between Bovine Serum Albumin (BSA) and fluorescence probe para-N,N-dimethylamino orthohydroxy benzaldehyde (PDOHBA), a structurally important molecule exhibiting excited state coupled proton transfer (PT) and charge transfer (CT) reaction. Fluorescence anisotropy, acrylamide quenching, and time resolved fluorescence measurements corroborate the binding nature of the probe with protein. The binding constant between BSA and PDOHBA has been determined by using Benesi-Hildebrand and Stern-Volmer equations. The negative value of ?G indicates the spontaneity of this probe-protein complexation process. Observations from synchronous, three dimensional fluorescence spectra and circular dichroism spectra point toward the fact that the hydrophobicity as well as ?-helix content of BSA are altered in presence of probe PDOHBA. The PT band of PDOHBA is found to be an excellent reporter for the mapping of destructive and protective behavior of SDS with variation of chaotrope concentration. PMID:21514035

  14. Demonstration of membrane estrogen binding proteins in rat brain by ligand blotting using a 17?-estradiol-[ 125I]bovine serum albumin conjugate

    Microsoft Academic Search

    Jianbiao Zheng; Victor D. Ramirez

    1997-01-01

    This paper describes a ligand blotting procedure to visualize membrane estrogen receptors\\/binding proteins immobilized on nitrocellulose membranes. Using 17?-estradiol covalently linked with [125I]bovine serum albumin (BSA) at the C-6 position (17?-E-6-[125I]BSA) as a ligand, three major binding proteins with molecular masses of approximately 23, 28, and 32 kDa were identified from crude synaptosomal fractions (P2) of female rat brains. The

  15. Bovine herpes virus gD protein produced in plants using a recombinant tobacco mosaic virus (TMV) vector possesses authentic antigenicity

    Microsoft Academic Search

    D. M Pérez Filgueira; P. I Zamorano; M. G Dom??nguez; O Taboga; M. P Del Médico Zajac; M Puntel; S. A Romera; T. J Morris; M. V Borca; A. M Sadir

    2003-01-01

    A tobacco mosaic virus (TMV)-based vector was utilized for expression of a cytosolic form of the bovine herpesvirus type 1 (BHV-1) protein glycoprotein D (gDc). Nicotiana benthamiana plants were harvested 7 days after inoculation with RNA transcripts derived from the TMV-gDc recombinant virus. Recombinant gDc protein of expected electrophoretic mobility accumulated in inoculated leaves to a concentration of about 20?g\\/g

  16. Artificial Transmembrane Oncoproteins Smaller than the Bovine Papillomavirus E5 Protein Redefine Sequence Requirements for Activation of the Platelet-Derived Growth Factor   Receptor

    Microsoft Academic Search

    Kristina Talbert-Slagle; Sara Marlatt; Francisco N. Barrera; Ekta Khurana; Joanne Oates; Mark Gerstein; Donald M. Engelman; Ann M. Dixon; Daniel DiMaio

    2009-01-01

    The bovine papillomavirus E5 protein (BPV E5) is a 44-amino-acid homodimeric transmembrane protein that binds directly to the transmembrane domain of the platelet-derived growth factor (PDGF) receptor and induces ligand-independent receptor activation. Three specific features of BPV E5 are considered important for its ability to activate the PDGF receptor and transform mouse fibroblasts: a pair of C-terminal cysteines, a transmembrane

  17. Isolation and characterization of two novel peptide amides originating from myelin basic protein in bovine brain

    Microsoft Academic Search

    Ken Takamatsu; Kazuhiko Tatemoto I

    1992-01-01

    During a systematic search for peptides that possess the C-terminal amide structure, two novel peptide amides, one with a tyrosine amide and the other with an alanine amide were isolated from bovine brain by acid extraction and sequential steps of reversed phase HPLC. Microsequence, amino acid and mass spectral analyses revealed the structures: Ac-Ala-Ala-Gln-Lys-Arg-Pro-Ser-Gln-Arg-Ser-Lys-Tyr-amide and Ac-Ala-Ala-Gln-Lys-Arg-Pro-Ser-Gln-Arg-Ser-Lys-Tyr-Leu-Ala-Ser-Ala-amide. These 12 and 16

  18. Comparison of capillary electrophoresis with traditional methods to analyse bovine whey proteins

    Microsoft Academic Search

    Nicki M. Kinghorn; Carmen S. Norris; Geoff R. Paterson; Don E. Otter

    1995-01-01

    The separation of the four major whey proteins by sodium dodecyl sulphate (SDS)-capillary gel electrophoresis (CGE) is described. Whilst commercially purified whey proteins could be analysed using the recommended protocol, the more complex nature of an acid whey and a reconstituted whey protein concentrate (WPC) powder necessitated considerable refinement of the CGE sample buffer. Individual whey proteins in the acid

  19. Sclera-Choroid-RPE Transport of Eight ?-Blockers in Human, Bovine, Porcine, Rabbit, and Rat Models

    PubMed Central

    Kadam, Rajendra S.; Cheruvu, Narayan P. S.; Edelhauser, Henry F.

    2011-01-01

    Purpose. To determine the influence of drug lipophilicity, ocular pigmentation, and species differences on transscleral solute transport. Methods. The transport of eight ?-blockers across excised sclera/sclera-choroid-RPE (SCRPE) of albino rabbit, pigmented rabbit, human, porcine, and bovine eyes was determined over 6 hours. The ex vivo transscleral ?-blocker transport to the vitreous at the end of 6 hours was determined in euthanatized, pigmented Brown Norway rats. The thicknesses of the sclera and SCRPE and the melanin content in choroid-RPE (CRPE) were measured to determine whether species differences in drug transport can be explained on this basis. Results. Solute lipophilicity inversely correlated with the SCRPE cumulative percentage of transport in all species (R2 ? 0.80). The CRPE impeded the SCRPE transport of all ?-blockers (51%–64% resistance in the rabbits; 84%–99.8% in the bovine and porcine eyes) more than the sclera, with the impedance increasing with lipophilicity. SCRPE transport followed the trend albino rabbit > pigmented rabbit > human > porcine > bovine, and a cross-species comparison showed good Spearman's rho correlation (R2 ? 0.85). Bovine (R2 = 0.84), porcine (R2 = 0.84), and human (R2 = 0.71) SCRPE transport was more predictive than that in the rabbit models (R2 = 0.60–0.61) of transscleral solute transport to the vitreous in rats. The CRPE concentrations were higher in pigmented rabbits than in albino rabbits. The melanin content of the CRPE exhibited the trend albino rabbit ? pigmented rabbit < porcine ? bovine < rat. Normalization to scleral thickness abolished the species differences in scleral transport. Normalization to SCRPE thickness and melanin content significantly reduced species differences in SCRPE transport. Conclusions. Owing to the presence of pigment and drug binding, choroid-RPE is the principal barrier to transscleral ?-blocker transport, with the barrier being more significant for lipophilic ?-blockers. Although different in magnitude between species, sclera/SCRPE transport can be correlated between species. Tissue thickness accounts for the species differences in scleral transport. Differences in tissue thickness and melanin content largely account for the species differences in SCRPE transport. PMID:21282583

  20. Crystal structure of the phosphatidylethanolamine-binding protein from bovine brain: a novel structural class of phospholipid-binding proteins

    Microsoft Academic Search

    Laurence Serre; Béatrice Vallée; Nicole Bureaud; Françoise Schoentgen; Charles Zelwer

    1998-01-01

    Background: Phosphatidylethanolamine-binding protein (PEBP) is a basic protein found in numerous tissues from a wide range of species. The screening of gene and protein data banks defines a family of PEBP-related proteins that are present in a variety of organisms, including Drosophila and inferior eukaryotes. PEBP binds to phosphatidylethanolamine and nucleotides in vitro, but its biological function in vivo is

  1. The acute phase response of haptoglobin and serum amyloid A (SAA) in cattle undergoing experimental infection with bovine respiratory syncytial virus

    Microsoft Academic Search

    Peter M. H Heegaard; Dale L Godson; Mathilda J. M Toussaint; Kirsten Tjørnehøj; Lars E Larsen; Birgitte Viuff; Leif Rønsholt

    2000-01-01

    The ability of a pure virus infection to induce an acute phase protein response is of interest as viral infections are normally considered to be less efficient in inducing an acute phase protein response than bacterial infections. This was studied in a bovine model for infection with bovine respiratory syncytial virus (BRSV), analysing the induction of the two most dominant

  2. Microwave Ablation Compared with Radiofrequency Ablation for Breast Tissue in an Ex Vivo Bovine Udder Model

    Microsoft Academic Search

    Toshihiro TanakaSaskia; Saskia Westphal; Peter Isfort; Till Braunschweig; Tobias Penzkofer; Philipp Bruners; Kimihiko Kichikawa; Thomas Schmitz-Rode; Andreas H. Mahnken

    Purpose  To compare the effectiveness of microwave (MW) ablation with radiofrequency (RF) ablation for treating breast tissue in a\\u000a nonperfused ex vivo model of healthy bovine udder tissue.\\u000a \\u000a \\u000a \\u000a \\u000a Materials and Methods  MW ablations were performed at power outputs of 25W, 35W, and 45W using a 915-MHz frequency generator and a 2-cm active tip\\u000a antenna. RF ablations were performed with a bipolar RF

  3. Confirmation of Free Solvent Model Assumptions in Predicting the Osmotic Pressure of Concentrated Globular Proteins

    Microsoft Academic Search

    M. A. Yousef; R. Datta; V. G. J. Rodgers

    2001-01-01

    Previously the osmotic pressure of the concentrated globular proteins immuno-?-globulin, bovine serum albumin, hen egg lysozyme, and ovalbumin in moderate-ionic-strength solutions was successfully modeled using a free solvent model (Yousef et al., J. Colloid Interface Sci.197, 108, 1998; 207, 273, 1998; AIChE J., 2001). This model considered the average solute–solvent and microion–solute interactions, represented the hydrated macromolecule and the cooperative

  4. Commercial Bovine Proteoglycan Is Highly Arthritogenic and Can Be Used as an Alternative Antigen Source for PGIA Model

    PubMed Central

    Ishikawa, Larissa Lumi Watanabe; Colavite, Priscila Maria; da Rosa, Larissa Camargo; França, Thais Graziela Donegá; Zorzella-Pezavento, Sofia Fernanda Gonçalves; Chiuso-Minicucci, Fernanda; Sartori, Alexandrina

    2014-01-01

    Rheumatoid arthritis (RA) is the most common systemic autoimmune disease. It affects mainly the joints, causing synovitis, cartilage destruction, and bone erosion. Many experimental models are used to study the mechanisms involved in immunopathogenesis and new therapies for this disease. Proteoglycan-induced arthritis (PGIA) is a widely used model based on the cross-reactivity of injected foreign (usually human) PG and mice self-PG. Considering the complexity of the extraction and purification of human PG, in this study we evaluated the arthritogenicity of bovine PG that is commercially available. Bovine PG was highly arthritogenic, triggering 100% incidence of arthritis in female BALB/c retired breeder mice. Animals immunized with bovine PG presented clinical symptoms and histopathological features similar to human RA and other experimental models. Moreover, bovine PG immunization determined higher levels of proinflammatory and anti-inflammatory cytokines in arthritic mice compared to healthy ones. As expected, only the arthritic group produced IgG1 and IgG2a antibodies against PG. Thus, commercial bovine PG can be used as an alternative antigenic source to PGIA for the study of many RA aspects, including the immunopathogenesis of the disease and also the development of new therapies. PMID:24971313

  5. In vitro cross-linking of bovine lens proteins photosensitized by promazines

    SciTech Connect

    Merville, M.P.; Decuyper, J.; Piette, J.; Calberg-Bacq, C.M.; Van de Vorst, A.

    1984-05-01

    Promazine derivatives induce cross-linking of bovine lens crystallins in vitro by irradiation with near-ultraviolet (UV) light in the presence of O/sub 2/, as revealed by electrophoresis after denaturation. With the five derivatives tested (promazine (PZ), chlorpromazine (CPZ), triflupromazine (TFPZ), methoxypromazine (MTPZ), and acepromazine (ACPZ)), single-hit kinetics are observed. Evidence implicating the cation radicals of the PZ derivatives as the causative agent of this in vitro effect is presented. Hydroxyl radicals do not appear to be involved in the photo-cross-linking reaction. Sodium ascorbate protects against damage induced either by PZ derivatives plus light or by PZ cation radicals in the dark. These findings are discussed with respect to development of cataracts induced by these drugs in vivo.

  6. Anisotropy in the compressive mechanical properties of bovine cortical bone and the mineral and protein constituents.

    PubMed

    Novitskaya, Ekaterina; Chen, Po-Yu; Lee, Steve; Castro-Ceseña, Ana; Hirata, Gustavo; Lubarda, Vlado A; McKittrick, Joanna

    2011-08-01

    The mechanical properties of fully demineralized, fully deproteinized and untreated cortical bovine femur bone were investigated by compression testing in three anatomical directions (longitudinal, radial and transverse). The weighted sum of the stress-strain curves of the treated bones was far lower than that of the untreated bone, indicating a strong molecular and/or mechanical interaction between the collagen matrix and the mineral phase. Demineralization and deproteinization of the bone demonstrated that contiguous, stand-alone structures result, showing that bone can be considered an interpenetrating composite material. Structural features of the samples from all groups were studied by optical and scanning electron microscopy. Anisotropic mechanical properties were observed: the radial direction was found to be the strongest for untreated bone, while the longitudinal one was found to be the strongest for deproteinized and demineralized bones. A possible explanation for this phenomenon is the difference in bone microstructure in the radial and longitudinal directions. PMID:21571104

  7. Unraveling the binding mechanism of polyoxyethylene sorbitan esters with bovine serum albumin: a novel theoretical model based on molecular dynamic simulations.

    PubMed

    Delgado-Magnero, Karelia H; Valiente, Pedro A; Ruiz-Peña, Miriam; Pérez-Gramatges, Aurora; Pons, Tirso

    2014-04-01

    To gain a better understanding of the interactions governing the binding mechanism of proteins with non-ionic surfactants, the association processes of Tween 20 and Tween 80 with the bovine serum albumin (BSA) protein were investigated using molecular dynamics (MD) simulations. Protein:surfactant molar ratios were chosen according to the critical micelle concentration (CMC) of each surfactant in the presence of BSA. It was found that both the hydrophilic and the hydrophobic groups of the BSA equally contribute to the surface area of interaction with the non-ionic surfactants. A novel theoretical model for the interactions between BSA and these surfactants at the atomic level is proposed, where both surfactants bind to non-specific domains of the BSA three-dimensional structure mainly through their polyoxyethylene groups, by hydrogen bonds and van der Waals interactions. This is well supported by the strong electrostatic and van der Waals interaction energies obtained in the calculations involving surfactant polyoxyethylene groups and different protein regions. The results obtained from the MD simulations suggest that the formation of surfactant clusters over the BSA structure, due to further cooperative self-assembly of Tween molecules, could increase the protein conformational stability. These results extend the current knowledge on molecular interactions between globular proteins and non-ionic surfactants, and contribute to the fine-tuning design of protein formulations using polysorbates as excipients for minimizing the undesirable effects of protein adsorption and aggregation. PMID:24309134

  8. A cloned bovine cDNA encodes an alternate form of the catalytic subunit of cAMP-dependent protein kinase.

    PubMed

    Showers, M O; Maurer, R A

    1986-12-15

    While attempting to isolate a cDNA clone for the catalytic subunit of the bovine cAMP-dependent protein kinase, we have isolated cDNAs which code for a protein slightly different than the known amino acid sequence. The alternate cDNA was identified by screening a bovine pituitary cDNA library using synthetic oligonucleotides predicted from the known amino acid sequence of the catalytic subunit. The cDNA which we identified, encodes a protein which is 93% identical to the known amino acid sequence of the bovine catalytic subunit. It seems likely that this cDNA represents a previously undiscovered catalytic subunit of the cAMP-dependent protein kinase. The mRNA for the alternate catalytic subunit is different in size from the mRNA coding for the previously known catalytic subunit and also has a different tissue distribution. These findings suggest that there are at least two different genes for the catalytic subunit. The differences in amino acid sequence and tissue distribution suggest the possibility of important functional differences in the two enzymes. PMID:3023347

  9. Protein nutrition for ruminants in European countries, in the light of animal feeding regulations linked to bovine spongiform encephalopathy.

    PubMed

    Sellier, P

    2003-04-01

    The outbreak of bovine spongiform encephalopathy (BSE) and the discovery of the central role played by meat-and-bone meal (MBM) as the vehicle of infection resulted, from the late 1980s onwards, in the implementation of new regulations on the incorporation of animal proteins, and then of most fats of animal origin, into diets fed to ruminants and other farmed animals. The BSE-related feed ban, which has gradually been reinforced over time, has led to the investigation of cost-effective routes for adequately replacing MBM and tallow by new sources of dietary proteins, minerals and lipids in the formulation of manufactured concentrates. As far as the technical fulfilment of the nutritive requirements of growing and lactating ruminants is concerned, efficient alternative solutions, based principally on recourse to food materials from vegetals already exist or hopefully will soon be available in most of the situations prevailing in Europe. However, related aspects, such as animal feed-processing, availability and traceability of certain food materials, quality of animal products, environmental constraints or disposal of animal waste from the meat industry give cause for concern. The expected consequences of the BSE-related feeding regulations on the organisational and economic framework of animal and crop production sectors throughout Europe and at world level must also be evaluated. PMID:12793784

  10. Generation of the Bovine Viral Diarrhea Virus E0 Protein in Transgenic Astragalus and Its Immunogenicity in Sika Deer

    PubMed Central

    Gao, Yugang; Zang, Pu; Liu, Qun; Wei, Gongqing

    2014-01-01

    The bovine viral diarrhea virus (BVDV), a single-stranded RNA virus, can cause fatal diarrhea syndrome, respiratory problems, and reproductive disorders in herds. Over the past few years, it has become clear that the BVDV infection rates are increasing and it is likely that an effective vaccine for BVDV will be needed. In this study, transgenic Astragalus was used as an alternative productive platform for the expression of glycoprotein E0. The immunogenicity of glycoprotein E0 expressed in transgenic Astragalus was detected in deer. The presence of pBI121-E0 was confirmed by polymerase chain reaction (PCR), transcription was verified by reverse transcription- (RT-) PCR, and recombinant protein expression was confirmed by ELISA and Western blot analyses. Deer that were immunized subcutaneously with the transgenic plant vaccine developed specific humoral and cell-mediated immune responses against BVDV. This study provides a new method for a protein with weak immunogenicity to be used as part of a transgenic plant vaccine. PMID:24963321

  11. Mutations in domain a? of protein disulfide isomerase affect the folding pathway of bovine pancreatic ribonuclease A

    PubMed Central

    Ruoppolo, Margherita; Orrù, Stefania; Talamo, Fabio; Ljung, Johanna; Pirneskoski, Annamari; Kivirikko, Kari I.; Marino, Gennaro; Koivunen, Peppi

    2003-01-01

    Protein disulfide isomerase (PDI, EC 5.3.4.1), an enzyme and chaperone, catalyses disulfide bond formation and rearrangements in protein folding. It is also a subunit in two proteins, the enzyme collagen prolyl 4-hydroxylase and the microsomal triglyceride transfer protein. It consists of two catalytically active domains, a and a?, and two inactive ones, b and b?, all four domains having the thioredoxin fold. Domain b? contains the primary peptide binding site, but a? is also critical for several of the major PDI functions. Mass spectrometry was used here to follow the folding pathway of bovine pancreatic ribonuclease A (RNase A) in the presence of three PDI mutants, F449R, ?455–457, and abb?, and the individual domains a and a?. The first two mutants contained alterations in the last ? helix of domain a?, while the third lacked the entire domain a?. All mutants produced genuine, correctly folded RNase A, but the appearance rate of 50% of the product, as compared to wild-type PDI, was reduced 2.5-fold in the case of PDI ?455–457, 7.5-fold to eightfold in the cases of PDI F449R and PDI abb?, and over 15-fold in the cases of the individual domains a and a?. In addition, PDI F449R and PDI abb? affected the distribution of folding intermediates. Domains a and a? catalyzed the early steps in the folding but no disulfide rearrangements, and therefore the rate observed in the presence of these individual domains was similar to that of the spontaneous process. PMID:12717017

  12. 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 ?-lactalbumin (ALA), ?-lactoglobulin (BLG), ovalbumin (OVA), bovin serum albumin (BSA) and HEWL were chosen for this investigation.

  13. Proteins other than the Locus of Enterocyte Effacement-encoded proteins may contribute to Escherichia coli O157:H7 adherence to bovine rectoanal junction stratified squamous epithelial cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this study, the Type III Secretion System (TTSS) proteins considered critical for Escherichia coli O157 (O157) adherence to the follicle-associated epithelial (FAE) cells at the bovine recto-anal junction (RAJ), did not appear to contribute to O157 adherence to the RAJ squamous epithelial (RSE) ...

  14. Amino-terminal sequence of bovine leukemia virus major internal protein: homology with mammalian type C virus p30 structural proteins.

    PubMed Central

    Oroszlan, S; Copeland, T D; Henderson, L E; Stephenson, J R; Gilden, R V

    1979-01-01

    The amino acid composition, the COOH-terminal amino acid, and the NH2-terminal amino acid sequence of the first 55 residues of the major internal structural protein, p24, of bovine leukemia virus (BLV) were determined. The compositional data and the results of end-group analysis revealed that, although BLV p24 is chemically distinct, it more closely resembles the p30 structural proteins than the other gag gene products of mammalian retroviruses. It was found that BLV p24 shares the common NH2-terminal proline and COOH-terminal leucine but lacks the common prolylleucylarginine tripeptide and the larger conserved region found near the NH2 terminus of all mammalian type C viral p30s. Alignment of the amino acid sequence of BLV p24 with the previously determined sequence of feline leukemia virus p27 revealed a statistically significant sequence homology. A more distant relationship was found between BLV p24 and other mammalian p30s. The finding of a definite sequence homology between BLV p24 and mammalian type C virus p30s clearly establishes the origin of these contemporary viral proteins from common progenitor genes. Images PMID:223166

  15. Fatty acid-binding protein activities in bovine muscle, liver and adipose tissue

    SciTech Connect

    Smith, S.B.; Ekeren, P.A.; Sanders, J.O.

    1985-11-01

    Subcutaneous adipose tissue, sternomandibularis muscle and liver were obtained from steers immediately postmortem. Muscle strips and adipose tissue snips were incubated with 0.75 mM (1- UC)palmitate and 5 mM glucose. Muscle strips esterified palmitate at the rate of 2.5 nmol/min per gram tissue, which was 30% of the rate observed for adipose tissue. Fatty acid-binding protein activity was measured in 104,000 x g supernatant fractions of liver, muscle and adipose tissue homogenates. Muscle and adipose tissue fractions bound 840 and 140 pmol (1- UC)palmitoyl-CoA per gram tissue, respectively. Fatty acid-binding protein activity was greater in adipose tissue than in muscle when data were expressed per milligram protein. Fatty acid binding-protein activity was correlated with the rate of palmitate esterification within each tissue. Liver contained the highest fatty acid-binding protein activity.

  16. Bovine Respiratory Syncytial Virus Nonstructural Proteins NS1 and NS2 Cooperatively Antagonize Alpha\\/Beta Interferon-Induced Antiviral Response

    Microsoft Academic Search

    JORG SCHLENDER; BIRGIT BOSSERT; URSULA BUCHHOLZ; KARL-KLAUS CONZELMANN

    2000-01-01

    The functions of bovine respiratory syncytial virus (BRSV) nonstructural proteins NS1 and NS2 were studied by generation and analysis of recombinant BRSV carrying single and double gene deletions. Whereas in MDBK cells the lack of either or both NS genes resulted in a 5,000- to 10,000-fold reduction of virus titers, in Vero cells a moderate (10-fold) reduction was observed. Interestingly,

  17. Polymorphic Forms of Expressed Bovine Interferon  Genes: Relative Transcript Abundance during Early Placental Development, Promoter Sequences of Genes and Biological Activity of Protein Products

    Microsoft Academic Search

    ALAN D. EALY; SANDRA F. LARSON; LIMIN LIU; ANDREI P. ALEXENKO; GAIL L. WINKELMAN; H. MICHAEL KUBISCH; JAMES A. BIXBY; R. MICHAEL ROBERTS

    2001-01-01

    Multiple interferon (IFN)-t genes exist in cattle, but it has re- mained unclear how many are expressed, the extent of their variation, and whether different genes exhibit similar patterns of expression and code for proteins with similar biological activities. A total of 118 complementary DNA (cDNA) were bi-directionally sequenced from reverse-transcribed bovine (bo) conceptus RNA over the period from blastocyst

  18. Study on acute phase proteins (haptoglobin, serum amyloid A, fibrinogen, and ceruloplasmin) changes and their diagnostic values in bovine tropical theileriosis

    Microsoft Academic Search

    S. Nazifi; S. M. Razavi; Z. Esmailnejad; H. Gheisari

    2009-01-01

    This study was conducted to assess the pattern of changes and the relative value of acute phase proteins (haptoglobin (Hp),\\u000a serum amyloid A (SAA), ceruloplasmin and fibrinogen) in bovine tropical theileriosis. The diseased group comprised 50 dairy\\u000a Holstein cattle, 2?3 years old, naturally infected with Theileria annulata. Infected animals were divided into four subgroups with different parasitemia rates (5%). As a

  19. Molecular characterization of RNA and protein synthesis during a one-step growth curve of bovine viral diarrhoea virus in ovine (SFT-R) cells

    Microsoft Academic Search

    N. Mishra; B. S. Mathapati; K. Rajukumar; R. K. Nema; S. P. Behera; S. C. Dubey

    2010-01-01

    The aim of this study was to determine the kinetics of noncytopathic bovine viral diarrhoea virus (BVDV) multiplication and synthesis of BVDV specific RNA and proteins in ovine cells (SFT-R) during a one-step growth curve. The virus titre and RNA level were determined by focus-forming assay and real time RT-PCR. The RNA synthesis was detected by Northern blot while synthesis

  20. Copper deficiency in the young bovine results in dramatic decreases in brain copper concentration but does not alter brain prion protein biology

    Microsoft Academic Search

    L. R. Legleiter; J. W. Spears; H. C. Liu

    2008-01-01

    An Mn for Cu substitution on cellular prion proteins (PrPc) in the brain that results in bio- chemical changes to PrPc has been implicated in the pathogenesis of transmissible spongiform encephal- opathies. Recent research in the mature bovine does not support this theory. The present study tested this hypothesis by using progeny from gestating cows re- ceiving Cu-deficient diets or

  1. Protein models docking benchmark 2.

    PubMed

    Anishchenko, Ivan; Kundrotas, Petras J; Tuzikov, Alexander V; Vakser, Ilya A

    2015-05-01

    Structural characterization of protein-protein interactions is essential for our ability to understand life processes. However, only a fraction of known proteins have experimentally determined structures. Such structures provide templates for modeling of a large part of the proteome, where individual proteins can be docked by template-free or template-based techniques. Still, the sensitivity of the docking methods to the inherent inaccuracies of protein models, as opposed to the experimentally determined high-resolution structures, remains largely untested, primarily due to the absence of appropriate benchmark set(s). Structures in such a set should have predefined inaccuracy levels and, at the same time, resemble actual protein models in terms of structural motifs/packing. The set should also be large enough to ensure statistical reliability of the benchmarking results. We present a major update of the previously developed benchmark set of protein models. For each interactor, six models were generated with the model-to-native C(?) RMSD in the 1 to 6 Å range. The models in the set were generated by a new approach, which corresponds to the actual modeling of new protein structures in the "real case scenario," as opposed to the previous set, where a significant number of structures were model-like only. In addition, the larger number of complexes (165 vs. 63 in the previous set) increases the statistical reliability of the benchmarking. We estimated the highest accuracy of the predicted complexes (according to CAPRI criteria), which can be attained using the benchmark structures. The set is available at http://dockground.bioinformatics.ku.edu. PMID:25712716

  2. Isolation of calcium-binding proteins on selective adsorbents. Application to purification of bovine calmodulin.

    PubMed

    Chaga, G S; Ersson, B; Porath, J O

    1996-05-01

    We report the fractionation of calcium-binding proteins using immobilized metal ion affinity chromatography (IMAC) with hard metal ions. Various hard metal ions (Mn2+, La3+, Nd3+, Eu(3 were immobilized on cross-linked agarose substituted with Tris(carboxymethyl)ethylenediamine (TED) and used as an adsorbent. After systematic studies, europium was selected for further work on the fractionation of calcium-binding proteins. It was found that the presence of Ca2+ in the sample and the solvent strongly promoted the adsorption and selectivity. Selective elution was accomplished in stepwise mode by the addition of calcium chelators such as malonate, citrate and phosphate. Calmodulin of high purity was isolated from a crude extract. Similar behavior of other calcium-binding proteins indicates that the reported chromatographic procedure can be generally applied to such proteins. PMID:8653201

  3. DNA sequence polymorphisms within the bovine guanine nucleotide-binding protein Gs subunit alpha (Gs?)-encoding (GNAS) genomic imprinting domain are associated with performance traits

    PubMed Central

    2011-01-01

    Background Genes which are epigenetically regulated via genomic imprinting can be potential targets for artificial selection during animal breeding. Indeed, imprinted loci have been shown to underlie some important quantitative traits in domestic mammals, most notably muscle mass and fat deposition. In this candidate gene study, we have identified novel associations between six validated single nucleotide polymorphisms (SNPs) spanning a 97.6 kb region within the bovine guanine nucleotide-binding protein Gs subunit alpha gene (GNAS) domain on bovine chromosome 13 and genetic merit for a range of performance traits in 848 progeny-tested Holstein-Friesian sires. The mammalian GNAS domain consists of a number of reciprocally-imprinted, alternatively-spliced genes which can play a major role in growth, development and disease in mice and humans. Based on the current annotation of the bovine GNAS domain, four of the SNPs analysed (rs43101491, rs43101493, rs43101485 and rs43101486) were located upstream of the GNAS gene, while one SNP (rs41694646) was located in the second intron of the GNAS gene. The final SNP (rs41694656) was located in the first exon of transcripts encoding the putative bovine neuroendocrine-specific protein NESP55, resulting in an aspartic acid-to-asparagine amino acid substitution at amino acid position 192. Results SNP genotype-phenotype association analyses indicate that the single intronic GNAS SNP (rs41694646) is associated (P ? 0.05) with a range of performance traits including milk yield, milk protein yield, the content of fat and protein in milk, culled cow carcass weight and progeny carcass conformation, measures of animal body size, direct calving difficulty (i.e. difficulty in calving due to the size of the calf) and gestation length. Association (P ? 0.01) with direct calving difficulty (i.e. due to calf size) and maternal calving difficulty (i.e. due to the maternal pelvic width size) was also observed at the rs43101491 SNP. Following adjustment for multiple-testing, significant association (q ? 0.05) remained between the rs41694646 SNP and four traits (animal stature, body depth, direct calving difficulty and milk yield) only. Notably, the single SNP in the bovine NESP55 gene (rs41694656) was associated (P ? 0.01) with somatic cell count--an often-cited indicator of resistance to mastitis and overall health status of the mammary system--and previous studies have demonstrated that the chromosomal region to where the GNAS domain maps underlies an important quantitative trait locus for this trait. This association, however, was not significant after adjustment for multiple testing. The three remaining SNPs assayed were not associated with any of the performance traits analysed in this study. Analysis of all pairwise linkage disequilibrium (r2) values suggests that most allele substitution effects for the assayed SNPs observed are independent. Finally, the polymorphic coding SNP in the putative bovine NESP55 gene was used to test the imprinting status of this gene across a range of foetal bovine tissues. Conclusions Previous studies in other mammalian species have shown that DNA sequence variation within the imprinted GNAS gene cluster contributes to several physiological and metabolic disorders, including obesity in humans and mice. Similarly, the results presented here indicate an important role for the imprinted GNAS cluster in underlying complex performance traits in cattle such as animal growth, calving, fertility and health. These findings suggest that GNAS domain-associated polymorphisms may serve as important genetic markers for future livestock breeding programs and support previous studies that candidate imprinted loci may act as molecular targets for the genetic improvement of agricultural populations. In addition, we present new evidence that the bovine NESP55 gene is epigenetically regulated as a maternally expressed imprinted gene in placental and intestinal tissues from 8-10 week old bovine foetuses. PMID:21214909

  4. Calculation of protein conformation as an assembly of stable overlapping segments: application to bovine pancreatic trypsin inhibitor.

    PubMed Central

    Simon, I; Glasser, L; Scheraga, H A

    1991-01-01

    Conformations of bovine pancreatic trypsin inhibitor were calculated by assuming that the final structure as well as properly chosen overlapping segments thereof are simultaneously in low-energy (not necessarily the lowest-energy) conformational states. Therefore, the whole chain can be built up from building blocks whose conformations are determined primarily by short-range interactions. Our earlier buildup procedure was modified by taking account of a statistical analysis of known amino acid sequences that indicates that there is nonrandom pairing of amino acid residues in short segments along the chain, and by carrying out energy minimization on only these segments and on the whole chain [without minimizing the energies of intermediate-size segments (20-30 residues long)]. Results of this statistical analysis were used to determine the variable sizes of the overlapping oligopeptide building blocks used in the calculations; these varied from tripeptides to octapeptides, depending on the amino acid sequence. Successive stages of approximations were used to combine the low-energy conformations of these building blocks in order to keep the number of variables in the computations to a manageable size. The calculations led to a limited number of conformations of the protein (only two different groups, with very similar structure within each group), most residues of which were in the same conformational state as in the native structure. PMID:2023916

  5. Preparation and characterization of bovine serum albumin surface-imprinted thermosensitive magnetic polymer microsphere and its application for protein recognition.

    PubMed

    Li, Xiangjie; Zhang, Baoliang; Li, Wei; Lei, Xingfeng; Fan, Xinlong; Tian, Lei; Zhang, Hepeng; Zhang, Qiuyu

    2014-01-15

    A novel bovine serum albumin surface-imprinted thermosensitive magnetic composite microsphere was successfully prepared by the surface grafting copolymerization method in the presence of temperature-sensitive monomer N-isopropylacrylamide (NIPAM), functional monomer methacrylic acid (MAA) and cross-linking agent N,N'-methylenebisacrylamide (MBA). The structure and component of the thermosensitive magnetic molecularly imprinted microsphere were investigated by transmission electron microscopy (TEM), Fourier transform infrared (FT-IR), vibrating sample magnetometer (VSM) and thermogravimetric analysis (TGA). The results of thermosensitivity, adsorption capacity, selectivity and reusability showed the formation of a thermosensitivity grafting polymer layer P(NIPAM-MAA-MBA) on the surface of Fe3O4@SiO2 and the good adsorption capacity and specific recognition for template protein. When the adsorption temperature was higher than the lower critical solution temperature (LCST) of poly(N-isopropylacrylamide) (PNIPAM), shape memory effect of imprinted cavities would be more effective. In other words, it was more conducive to capture template molecules under this condition and the imprinting factor would be higher. On the other hand, when the desorption temperature was lower than LCST of PNIPAM, the decrease of shape memory effect between imprinted cavities and template molecules would facilitate the release of template molecules from the imprinted cavities. Based on this property, the adsorption and desorption of template molecules could be regulated by system temperature indirectly which benefited from the existence of thermosensitivity imprinting layer. PMID:23973936

  6. An innovative bovine odorant binding protein-based filtering cartridge for the removal of triazine herbicides from water.

    PubMed

    Bianchi, Federica; Basini, Giuseppina; Grolli, Stefano; Conti, Virna; Bianchi, Francesco; Grasselli, Francesca; Careri, Maria; Ramoni, Roberto

    2013-01-01

    Odorant binding protein (OBP) is a multi-functional scavenger for small hydrophobic molecules dissolved in the mucus lining the nasal epithelia of mammals, characterized by broad ligand binding specificity towards a large number of structurally unrelated natural and synthetic molecules of different chemical classes. Here, we demonstrate for the first time the application of OBP as the active element of an innovative filtering matrix for the removal of environmental pollutants such as triazine herbicides from water samples. The filtering device, obtained by coupling histidine-tagged bovine OBP to a nickel nitrilotriacetic acid (Ni-NTA) agarose resin, was characterized in terms of retention capacity for the herbicides atrazine, simazine, and propazine. Analysis of these herbicides at trace levels with solid-phase microextraction followed by gas chromatography-mass spectrometry using the selected ion monitoring mode proved the capabilities of the proposed device for the decontamination of surface and groundwater samples in the 0.2-2,300 ?g/L concentration range, obtaining a reduction in the triazine content greater than 97 %, thus suggesting its possible use for the potabilization of water. PMID:23104315

  7. Some nutritional effects of folate-binding protein in bovine milk on the bioavailability of folate to rats

    SciTech Connect

    Tani, M.; Iwai, K.

    1984-04-01

    The excretions of folate compounds into both the urine and bile were investigated in rats after the administration of pteroylglutamic acid (PteGlu) with or without the folate-binding protein (FBP) prepared from bovine milk. When the sample solution, containing either free or bound (/sup 3/H)PteGlu (i.e., bound to the FBP from milk), was delivered to rats intragastrically via oral intubation, the amounts of (/sup 3/H)PteGlu excreted into the feces did not change. On the other hand, the urinary excretion of /sup 3/H-labeled folate compounds, especially (/sup 3/H)5-methyltetrahydrofolic acid (5-CH/sub 3/-H/sub 4/PteGlu), after the administration of bound (/sup 3/H)PteGlu was significantly lower (P less than 0.01) than that after the administration of free (/sup 3/H)PteGlu. The urinary excretion of (/sup 3/H)5-CH/sub 3/-H/sub 4/PteGlu was directly proportional to the initial amount of free (/sup 3/H)PteGlu administered. The similar effect of FBP was also observed when the biliary excretion of /sup 3/H-labeled folate compounds was investigated in situ. Furthermore, the incorporation of (/sup 3/H)PteGlu into folate-requiring intestinal microorganisms was considerably reduced when it was bound to FBP. These results suggest that milk FBP has some nutritional effects on the bioavailability of folate in vivo.

  8. Transmission of scrapie and sheep-passaged bovine spongiform encephalopathy prions to transgenic mice expressing elk prion protein

    PubMed Central

    Tamgüney, Gültekin; Miller, Michael W.; Giles, Kurt; Lemus, Azucena; Glidden, David V.; DeArmond, Stephen J.; Prusiner, Stanley B.

    2009-01-01

    Chronic wasting disease (CWD) is a transmissible, fatal prion disease of cervids and is largely confined to North America. The origin of CWD continues to pose a conundrum: does the disease arise spontaneously or result from some other naturally occurring reservoir? To address whether prions from sheep might be able to cause disease in cervids, we inoculated mice expressing the elk prion protein (PrP) transgene [Tg(ElkPrP) mice] with two scrapie prion isolates. The SSBP/1 scrapie isolate transmitted disease to Tg(ElkPrP) mice with a median incubation time of 270?days, but a second isolate failed to produce neurological dysfunction in these mice. Although prions from cattle with bovine spongiform encephalopathy (BSE) did not transmit to the Tg(ElkPrP) mice, they did transmit after being passaged through sheep. In Tg(ElkPrP) mice, the sheep-passaged BSE prions exhibited an incubation time of approximately 300?days. SSBP/1 prions produced abundant deposits of the disease-causing PrP isoform, denoted PrPSc, in the cerebellum and pons of Tg(ElkPrP) mice, whereas PrPSc accumulation in Tg mice inoculated with sheep-passaged BSE prions was confined to the deep cerebellar nuclei, habenula and the brainstem. The susceptibility of ‘cervidized’ mice to ‘ovinized’ prions raises the question about why CWD has not been reported in other parts of the world where cervids and scrapie-infected sheep coexist. PMID:19264659

  9. Binding of a substrate analog to a domain swapping protein: X-ray structure of the complex of bovine seminal ribonuclease with uridylyl(2',5')adenosine.

    PubMed Central

    Vitagliano, L.; Adinolfi, S.; Riccio, A.; Sica, F.; Zagari, A.; Mazzarella, L.

    1998-01-01

    Bovine seminal ribonuclease (BS-RNase) is a unique member of the pancreatic-like ribonuclease superfamily. The native enzyme is a mixture of two dimeric forms with distinct structural features. The most abundant form is characterized by the swapping of N-terminal fragments. In this paper, the crystal structure of the complex between the swapping dimer and uridylyl(2',5')adenosine is reported at 2.06 A resolution. The refined model has a crystallographic R-factor of 0.184 and good stereochemistry. The quality of the electron density maps enables the structure of both the inhibitor and active site residues to be unambiguously determined. The overall architecture of the active site is similar to that of RNase A. The dinucleotide adopts an extended conformation with the pyrimidine and purine base interacting with Thr45 and Asn71, respectively. Several residues (Gln11, His12, Lys41, His119, and Phe120) bind the oxygens of the phosphate group. The structural similarity of the active sites of BS-RNase and RNase A includes some specific water molecules believed to be relevant to catalytic activity. Upon binding of the dinucleotide, small but significant modifications of the tertiary and quaternary structure of the protein are observed. The ensuing correlation of these modifications with the catalytic activity of the enzyme is discussed. PMID:10082366

  10. The comparative specificity of 3 oestradiol-binding proteins. Rat alpha-foetoprotein, rat liver 17beta-hydroxy steroid dehydrogenase and anti-(oestradiol-6-carboxymethyloxime-bovine serum albumin) antiserum.

    PubMed Central

    Laurant, C; de Lauzon, S D; Cittanova, N; Nunez, E; Jayle, M F

    1975-01-01

    1. The specificity of 3 oestradiol-binding proteins was studied. Two of these proteins are naturally occurring (rat alpha-foetoprotein and rat liver microsomal 17beta-hydroxy steroid dehydrogenase) and the third is an artificially induced model, anti-(oestradiol-6-carboxymethyloxime-bovine serum albumin) gamma-globulins. 2. A specific binding procedure for each protein model permitted a determination of its affinity for oestradiol and for 30 other steroids. 3. The results obtained have brought to light the different areas of the steroid molecule that are important for its recognition by each of the three proteins. The two naturally occurring proteins (alpha-foetoprotein and 17beta-hydroxy steroid dehydrogenase) recognize the edge of the steroid defined by C-4, C-6, C-8 and C-15. On the other hand, the gamma-globulins recognize the opposite edge, i.e. that defined by C-2, C-10, C-11 and C-17. 4. Diethylstilboestrol, whose structure is analogous to that of a steroid, is only recognized by the two naturally occurring proteins. PMID:56174

  11. EFFECTS OF PREPARTUM PROTEIN NUTRITION AND BIRTH WEIGHT ON BASAL METABOLISM IN BOVINE NEONATES x

    Microsoft Academic Search

    G. E. Carstens; D. E. Johnson; M. D. Holland; K. G. Odde

    Recipient beef heifers, pregnant with single demi-embryos, were paired according to identical twin or full-sib embryo. Within pair, recipient heifers were assigned to one of two isocaloric diets containing a control or restricted level of protein (91 vs 55% of National Research Council recom- mendations) on d 190 of gestation. Following parturition, calves were weighed, fed 1 liter of colo-

  12. The Positively Charged Termini of L2 Minor Capsid Protein Required for Bovine Papillomavirus Infection Function Separately in Nuclear Import and DNA Binding

    PubMed Central

    Fay, Alyson; Yutzy, William H.; Roden, Richard B. S.; Moroianu, Junona

    2004-01-01

    During the papillomavirus (PV) life cycle, the L2 minor capsid protein enters the nucleus twice: in the initial phase after entry of virions into cells and in the productive phase to mediate encapsidation of the newly replicated viral genome. Therefore, we investigated the interactions of the L2 protein of bovine PV type 1 (BPV1) with the nuclear import machinery and the viral DNA. We found that BPV1 L2 bound to the karyopherin ?2 (Kap ?2) adapter and formed a complex with Kap ?2?1 heterodimers. Previous data have shown that the positively charged termini of BPV1 L2 are required for BPV1 infection after the binding of the virions to the cell surface. We determined that these BPV1 L2 termini function as nuclear localization signals (NLSs). Both the N-terminal NLS (nNLS) and the C-terminal NLS (cNLS) interacted with Kap ?2, formed a complex with Kap ?2?1 heterodimers, and mediated nuclear import via a Kap ?2?1 pathway. Interestingly, the cNLS was also the major DNA binding site of BPV1 L2. Consistent with the promiscuous DNA encapsidation by BPV1 pseudovirions, this DNA binding occurred without nucleotide sequence specificity. Moreover, an L2 mutant encoding a scrambled version of the cNLS, which supports production of virions, rescued the DNA binding but not the Kap ?2 interaction. These data support a model in which BPV1 L2 functions as an adapter between the viral DNA via the cNLS and the Kaps via the nNLS and facilitates nuclear import of the DNA during infection. PMID:15564455

  13. Expression and characterization of novel ovine orthologs of bovine placental prolactin-related proteins

    Microsoft Academic Search

    Koichi Ushizawa; Toru Takahashi; Misa Hosoe; Katsuhiro Ohkoshi; Kazuyoshi Hashizume

    2007-01-01

    BACKGROUND: The prolactin-related proteins (PRPs) are non-classical placental-specific members of the prolactin\\/growth hormone family. Among ruminants, they are expressed in the cotyledonary villi of cattle and goat. We investigated placental PRP in sheep in order to gain a comprehensive understanding of the function and evolution of these molecules. We also examined the sequence properties, expression and lactogenic activation of the

  14. Immobilisation of bovine enterokinase and application of the immobilised enzyme in fusion protein cleavage

    Microsoft Academic Search

    Tina Kubitzki; Thomas Noll; Stephan Lütz

    2008-01-01

    Two immobilisation methods for enterokinase were developed, which yielded high remaining activities for the cleavage of the\\u000a fusion protein MUC1-IgG Fc. Different carrier materials were compared regarding remaining enzyme activity and storage stability.\\u000a Immobilisation procedures involving support material activation using glutardialdehyde were found to result in low remaining\\u000a activities. Applying less aggressive activation procedures, remaining activities of ?60% were received

  15. Microwave Ablation Compared with Radiofrequency Ablation for Breast Tissue in an Ex Vivo Bovine Udder Model

    SciTech Connect

    Tanaka, Toshihiro, E-mail: toshihir@bf6.so-net.ne.jp [RWTH Aachen University, Applied Medical Engineering, Helmholtz-Institute Aachen (Germany); Westphal, Saskia, E-mail: swestphal@ukaachen.de [RWTH Aachen University, Department of Pathology Aachen University Hospital (Germany); Isfort, Peter, E-mail: isfort@hia.rwth-aachen.de [RWTH Aachen University, Applied Medical Engineering, Helmholtz-Institute Aachen (Germany); Braunschweig, Till, E-mail: tbraunschweig@ukaachen.de [RWTH Aachen University, Department of Pathology Aachen University Hospital (Germany); Penzkofer, Tobias, E-mail: penzkofer@hia.rwth-aachen.de; Bruners, Philipp, E-mail: bruners@rad.rwth-aachen.de [RWTH Aachen University, Applied Medical Engineering, Helmholtz-Institute Aachen (Germany); Kichikawa, Kimihiko, E-mail: kkichika@naramed-u.ac.jp [Nara Medical University, Department of Radiology (Japan); Schmitz-Rode, Thomas, E-mail: smiro@hia.rwth-aachen.de; Mahnken, Andreas H., E-mail: mahnken@rad.rwth-aachen.de [RWTH Aachen University, Applied Medical Engineering, Helmholtz-Institute Aachen (Germany)

    2012-08-15

    Purpose: To compare the effectiveness of microwave (MW) ablation with radiofrequency (RF) ablation for treating breast tissue in a nonperfused ex vivo model of healthy bovine udder tissue. Materials and Methods: MW ablations were performed at power outputs of 25W, 35W, and 45W using a 915-MHz frequency generator and a 2-cm active tip antenna. RF ablations were performed with a bipolar RF system with 2- and 3-cm active tip electrodes. Tissue temperatures were continuously monitored during ablation. Results: The mean short-axis diameters of the coagulation zones were 1.34 {+-} 0.14, 1.45 {+-} 0.13, and 1.74 {+-} 0.11 cm for MW ablation at outputs of 25W, 35W, and 45W. For RF ablation, the corresponding values were 1.16 {+-} 0.09 and 1.26 {+-} 0.14 cm with electrodes having 2- and 3-cm active tips, respectively. The mean coagulation volumes were 2.27 {+-} 0.65, 2.85 {+-} 0.72, and 4.45 {+-} 0.47 cm{sup 3} for MW ablation at outputs of 25W, 35W, and 45W and 1.18 {+-} 0.30 and 2.29 {+-} 0.55 cm{sup 3} got RF ablation with 2- and 3-cm electrodes, respectively. MW ablations at 35W and 45W achieved significantly longer short-axis diameters than RF ablations (P < 0.05). The highest tissue temperature was achieved with MW ablation at 45W (P < 0.05). On histological examination, the extent of the ablation zone in MW ablations was less affected by tissue heterogeneity than that in RF ablations. Conclusion: MW ablation appears to be advantageous with respect to the volume of ablation and the shape of the margin of necrosis compared with RF ablation in an ex vivo bovine udder.

  16. Effects of electrical stimulation high temperature pre-rigor conditioning on myofibrillar proteins of bovine muscle 

    E-print Network

    Adams, Keith LeRoy

    1977-01-01

    C and various pH's. 19 The absorbance due to CAF treatment as related to shear force values of cooked samples. 60 CHAPTFR I INTRODUCTION A large amount of the research effort in the field of meats is directed toward producing a more palatable... stimulation immediately postmortem and with beef conditioned at 21 C. Both CAF (Calcium Activated Factor) and cathepsins (neutral and acidic) have been shown to degrade some of the myofibrillar proteins in vitro and could cause these changes in intact...

  17. Mouse model of membranous nephropathy induced by cationic bovine serum albumin: antigen dose-response relations and strain differences

    Microsoft Academic Search

    Jin-Shuen Chen; Ann Chen; Li-Chien Chang; Wun-Shaing Wayne Chang; Herng-Sheng Lee; Shih-Hua Lin; Yuh-Feng Lin

    Background. Few well-characterized animal models have been developed to study the pathogenesis of membranous nephropathy (MN). We have developed a mouse model of MN induced by cationic bovine serum albumin (cBSA), and examined the role of genetic background on disease induction by assessing different mouse strains. Methods. cBSA in an optimum dose was given intravenously to 8-week-old female ICR, BALB\\/c

  18. Straightforward isolation of phosphatidyl-ethanolamine-binding protein-1 (PEBP-1) and ubiquitin from bovine testis by hydrophobic-interaction chromatography (HIC).

    PubMed

    Haj Hassan, Maya; Klett, Danièle; Cahoreau, Claire; Combarnous, Yves

    2011-10-01

    Isolation of phosphatidyl-ethanolamine-binding protein-1 (PEBP-1) from bovine brain was described almost three decades ago but it required a large number of steps to reach high purity. After the fractionation of bovine testis proteins by ammonium sulfate precipitation we found that PEBP-1, detected by Western blotting, was among the very few proteins still soluble at 80% ammonium sulfate saturation (3.2M). This soluble fraction (S80) was directly loaded onto a phenyl sepharose column equilibrated at the same ammonium sulfate concentration (3.2M). A stepwise elution of the retained material at 1.0, 0.5, 0.2, 0.1M ammonium sulfate in ammonium hydrogen carbonate was performed and then with ammonium hydrogen carbonate alone and finally with 50% ethylene glycol. All fractions were analyzed by SDS-PAGE and Western blotting and the fractions containing PEBP-1 was further fractionated by size exclusion chromatography on a HR75 Superdex column permitting the isolation of ubiquitin in addition to PEBP-1 as demonstrated by Western blotting and mass spectrometry. This study shows the feasibility of hydrophobic interaction chromatography (HIC) on phenyl sepharose at a very high ammonium sulfate concentration (3.2M; 80% saturation) to efficiently purify the proteins that are still soluble in these extreme conditions. PMID:21917533

  19. Diffusion models of protein folding

    PubMed Central

    2012-01-01

    In theory and in the analysis of experiments, protein folding is often described as diffusion along a single coordinate. We explore here the application of a one-dimensional diffusion model to interpret simulations of protein folding, where the parameters of a model that “best” describes the simulation trajectories are determined using a Bayesian analysis. We discuss the requirements for such a model to be a good approximation to the global dynamics, and several methods for testing its accuracy. For example, one test considers the effect of an added bias potential on the fitted free energies and diffusion coefficients. Such a bias may also be used to extend our approach to determining parameters for the model to systems which would not normally explore the full coordinate range on accessible time scales. Alternatively, the propagators predicted from the model at different “lag” times may be compared with observations from simulation. We then present some applications of the model to protein folding, including Kramers-like turnover in folding rates of coarse-grained models, the effect of non-native interactions on folding, and the effect of the chosen coordinate on the observed position-dependence of the diffusion coefficients. Lastly, we consider how our results are useful for the interpretation of experiments, and how this type of Bayesian analysis may eventually be applied directly to analyse experimental data. PMID:21842082

  20. 4528 J. Phys. Chem. 1993,97, 4528-4534 Electrophoretic and Quasi-ElasticLight Scattering of Soluble Protein-Polyelectrolyte Complexes

    E-print Network

    Dubin, Paul D.

    Form: November 30, I992 Complexation between globular proteins (bovine serum albumin, bovine pancreas ribonuclease, and chicken egg lysozyme) and a number of synthetic polyelectrolytes was studied by quasi of bovine serum albumin (BSA) and quaternized poly(4-vinylpyridines), Kabanov22proposed a model in which

  1. Tissue expression and predicted protein structures of the bovine ANGPTL3 and association of novel SNPs with growth and meat quality traits.

    PubMed

    Chen, N B; Ma, Y; Yang, T; Lin, F; Fu, W W; Xu, Y J; Li, F; Li, J Y; Gao, S X

    2015-08-01

    Angiopoietin-like protein 3 (ANGPTL3) is a secreted protein that regulates lipid, glucose and energy metabolism. This study was conducted to better understand the effect of ANGPTL3 on important economic traits in cattle. First, transcript profiles for ANGPTL3 were measured in nine different Jiaxian cattle tissues. Second, polymorphisms were identified in the complete coding region and promoter region of the bovine ANGPTL3 gene in 707 cattle samples. Finally, an association study was carried out utilizing these single nucleotide polymorphisms (SNPs) to determine the effect of these SNPs on the growth and meat quality traits. Quantitative real-time PCR analysis showed that ANGPTL3 was mainly expressed in the liver. The promoter of the bovine ANGPTL3 contained several putative transcription factor binding sites (SF1, HNF-1, LXR?, NF??, HNF-3 and C/EBP). In total, four SNPs of the bovine ANGPTL3 gene were identified by direct sequencing. SNP1 (rs469906272: g.-38T>C) was identified in the promoter, SNP2 (rs451104723:g.104A>T) and SNP3 (rs482516226: g.509A>G) were identified in exon 1, and SNP4 (rs477165942: g.8661T>C) was identified in exon 6. Changes in predicted protein structures due to non-synonymous SNPs were analyzed. Haplotype frequencies and linkage disequilibrium were also investigated. Analysis of four SNPs in cattle from different native Chinese breeds (Nanyang (NY) and Jiaxian (JX)) and commercial breeds (Angus (AG), Hereford (HF), Limousin (LM), Luxi (LX), Simmental (ST) and Jinnan (JN)) revealed a significant association with growth traits (including: BW and hipbone width) and meat quality traits (including: Warner-Bratzler shear force and ribeye area). Therefore, implementation of these four mutations in selection indices in the beef industry may be beneficial in selecting individuals with superior growth and meat quality traits. PMID:25951897

  2. High-fat diet based on dried bovine brain: an effective animal model of dyslipidemia and insulin resistance.

    PubMed

    Araújo, Tiago Gomes; Leite, Ana Catarina Rezende; Martins da Fonseca, Caíque Silveira; Carvalho, Bruno Melo; Schuler, Alexandre Ricardo Pereira; Lima, Vera Lúcia de Menezes

    2011-09-01

    Currently, there are no reports in the literature demonstrating any animal model that ingests one of the fattiest animal food source, the bovine brain. We hypothesized that a high-fat diet (HFD), based on dried bovine brain, could be used to develop an animal model possessing a spectrum of insulin resistance-related features. The HFD was formulated with 40% dried bovine brain plus 16.4% butter fat, prepared in-house. Furthermore, the diet contained 52% calories as fat and 73% of total fatty acids were saturated. Swiss mice weighing about 40 g were assigned to two dietary groups (n=6/group), one group received a standard chow diet and the other was given HFD for 3 months. The body weight and biochemical parameters of the animals were measured initially and at monthly intervals until the end of the experiment. Animals fed on a HFD showed a significant increase in the body and adipose tissue weight, serum total cholesterol and triglyceride levels, when compared with mice fed on the control diet. Additionally, the HFD group showed higher circulating levels of liver transaminases, such as alanine aminotransferase and aspartate aminotransferase, compared with the control group. Finally, to illustrate the usefulness of this model, we report that the HFD induced mild hyperglycemia, fasting hyperinsulinemia, and increased the homeostasis model of assessment (HOMA-IR), in comparison with the control group. In conclusion, our results show that HFD, based on dried bovine brain, causes insulin resistance-related metabolic disturbances. Thus, this may be a suitable model to study disturbances in energy metabolism and their consequences. PMID:21437730

  3. Viscoelastic properties of bovine orbital connective tissue and fat: constitutive models

    PubMed Central

    Yoo, Lawrence; Gupta, Vijay; Lee, Choongyeop; Kavehpore, Pirouz

    2012-01-01

    Reported mechanical properties of orbital connective tissue and fat have been too sparse to model strain–stress relationships underlying biomechanical interactions in strabismus. We performed rheological tests to develop a multi-mode upper convected Maxwell (UCM) model of these tissues under shear loading. From 20 fresh bovine orbits, 30 samples of connective tissue were taken from rectus pulley regions and 30 samples of fatty tissues from the posterior orbit. Additional samples were defatted to determine connective tissue weight proportion, which was verified histologically. Mechanical testing in shear employed a triborheometer to perform: strain sweeps at 0.5–2.0 Hz; shear stress relaxation with 1% strain; viscometry at 0.01–0.5 s?1 strain rate; and shear oscillation at 1% strain. Average connective tissue weight proportion was 98% for predominantly connective tissue and 76% for fatty tissue. Connective tissue specimens reached a long-term relaxation modulus of 668 Pa after 1,500 s, while corresponding values for fatty tissue specimens were 290 Pa and 1,100 s. Shear stress magnitude for connective tissue exceeded that of fatty tissue by five-fold. Based on these data, we developed a multimode UCM model with variable viscosities and time constants, and a damped hyperelastic response that accurately described measured properties of both connective and fatty tissues. Model parameters differed significantly between the two tissues. Viscoelastic properties of predominantly connective orbital tissues under shear loading differ markedly from properties of orbital fat, but both are accurately reflected using UCM models. These viscoelastic models will facilitate realistic global modeling of EOM behavior in binocular alignment and strabismus. PMID:21207094

  4. Speciation of trace elements in proteins in human and bovine serum by size exclusion chromatography and inductively coupled plasma-mass spectrometry with a magnetic sector mass spectrometer.

    PubMed

    Wang, J; Houk, R S; Dreessen, D; Wiederin, D R

    1999-10-01

    Proteins are separated by size exclusion chromatography while atomic ions from the inorganic elements are detected on-line by inductively coupled plasma-mass spectrometry. A double focusing mass analyzer provides very high sensitivity, low background, and sufficient spectral resolution to separate the atomic ions of interest from most polyatomic ions at the same nominal m/z value. The chromatograms show the distribution of the elements of interest between protein-bound and free fractions and provide the approximate molecular weights of those protein fractions that contain the elements monitored. The distribution of various elements, including V, Mo, Fe, Co, Mn, and lanthanides, in human or bovine serum samples are shown. Alkali metals and Tl are present primarily as free metal ions and are not bound to proteins. Inorganic elements spiked into the serum samples can be followed into various proteins. EDTA does not remove Fe, Pb, Sn, or Th from the proteins but does extract Mn from some proteins. Procedures for determining the effects of breaking disulfide linkages on the metal binding characteristics of proteins are also described. PMID:10550683

  5. Metabolism of soluble rapeseed meal (Brassica rapa L.) protein during incubations with buffered bovine rumen contents in vitro.

    PubMed

    Stefa?ski, T; Ahvenjärvi, S; Huhtanen, P; Shingfield, K J

    2013-01-01

    Accurate quantitative information on the fate of dietary protein in the rumen is central to modern metabolizable protein systems developed to improve the efficiency of nitrogen utilization in ruminants. An in vitro method was developed to estimate the rate of soluble rapeseed meal (Brassica rapa L.) protein (SRMP) degradation. Unlabeled and (15)N-labeled solvent-extracted rapeseed meal were incubated alone or as an equal mixture (125 mg of N/L) with buffered rumen contents and a mixture of carbohydrates formulated to provide a constant source of fermentable energy during the course of all incubations. Incubations were made over 0.33, 0.67, 1.0, 1.5, 2.0, 2.5, 3.0, 4.0, 6.0, 8.0, and 10.0 h. Enrichment of (14)N and (15)N isotopes in total N of ammonia (AN), soluble nonammonia (SNAN), and insoluble (ISN) fractions liberated during incubations with test proteins was determined. A model with 4 pools that accounted for both intracellular and extracellular N transformations was used to estimate the rate of SRMP degradation. Parameter values used in the model were adjusted based on the size of A(14)N, A(15)N, SNA(14)N, SNA(15)N, IS(14)N, and IS(15)N pools, measured at different time points during incubations with buffered rumen fluid. The mean rate of N degradation for SRMP was estimated at 0.126 (SD 0.0499) h(-1). No substantive difference in the rate of protein degradation or microbial protein synthesis was observed during incubations of labeled and unlabeled substrates with rumen fluid. In conclusion, combined use of data from incubations of unlabeled and (15)N-labeled rapeseed protein with buffered rumen inoculum provided sufficient information to allow for estimation of parameter values in a complex dynamic model of soluble protein degradation. Results indicate the potential of the technique to evaluate the degradability of SNAN of other dietary protein sources and implicate ruminal escape of soluble rapeseed protein as an important source of amino acids in ruminants. PMID:23127902

  6. Experimentally-based multiscale model of the elastic moduli of bovine trabecular bone and its constituents.

    PubMed

    Hamed, Elham; Novitskaya, Ekaterina; Li, Jun; Jasiuk, Iwona; McKittrick, Joanna

    2015-09-01

    The elastic moduli of trabecular bone were modeled using an analytical multiscale approach. Trabecular bone was represented as a porous nanocomposite material with a hierarchical structure spanning from the collagen-mineral level to the trabecular architecture level. In parallel, compression testing was done on bovine femoral trabecular bone samples in two anatomical directions, parallel to the femoral neck axis and perpendicular to it, and the measured elastic moduli were compared with the corresponding theoretical results. To gain insights on the interaction of collagen and minerals at the nanoscale, bone samples were deproteinized or demineralized. After such processing, the treated samples remained as self-standing structures and were tested in compression. Micro-computed tomography was used to characterize the hierarchical structure of these three bone types and to quantify the amount of bone porosity. The obtained experimental data served as inputs to the multiscale model and guided us to represent bone as an interpenetrating composite material. Good agreement was found between the theory and experiments for the elastic moduli of the untreated, deproteinized, and demineralized trabecular bone. PMID:26046284

  7. A phenomenological model of protein folding

    E-print Network

    Danielsson, Ulf H; Niemi, Antti J

    2009-01-01

    We construct a phenomenological effective field theory model that describes the universality class of biologically active single-strand proteins. The model allows both for an explicit construction of native state protein conformations, and a dynamical description of protein folding and unfolding processes. The model reveals a connection between homochirality and protein collapse, and enables the theoretical investigation of various other aspects of protein folding even in the case of very long polypeptide chains where other methods are not available.

  8. Bone temperature during cementation with a heatsink: a bovine model pilot study

    PubMed Central

    2014-01-01

    Background Bone cement is an effective means of supporting implants, but reaches high temperatures while undergoing polymerisation. Bone has been shown to be sensitive to thermal injury with osteonecrosis reported after one minute at 47°C. Necrosis during cementing may lead to loosening of the prosthesis. Some surgeons fill the joint cavity with cool irrigation fluid to provide a heatsink during cementing, but this has not been supported by research. This paper assesses a simple technique to investigate the efficacy of this method. Findings We used a model acetabulum in a bovine humerus to allow measurement of bone temperatures in cementing. Models were prepared with a 50 mm diameter acetabulum and three temperature probe holes; two as close as possible to the acetabular margin at half the depth of the acetabulum and at the full depth of the acetabulum, and one 10 mm from the acetabular rim. Four warmed models were cemented with Palacos RG using a standard mixing system and a 10 mm polyethylene disc to represent an acetabular component. Two of the acetabular models were filled with room temperature water to provide a heatsink. An electronic probe measured temperature at 5 second intervals from the moment of cementing. In the models with no heatsink, peak temperature was 40.3°C. The mean temperature rise was 10.9°C. In the models with a heatsink, there was an average fall in the bone temperature during cementing of 4.4°C. Conclusions These results suggest that using a heatsink while cementing prostheses may reduce the peak bone temperature. This study demonstrates a simple, repeatable technique which may be useful for larger trials. PMID:25099248

  9. Crystal Structure of Bovine Coronavirus Spike Protein Lectin Received for publication,September 10, 2012, and in revised form, October 19, 2012 Published, JBC Papers in Press,October 22, 2012, DOI 10.1074/jbc.M112.418210

    E-print Network

    Li, Fang

    protein-binding NTD of mouse coronavirus. Significance: This study explores origin and evolution or protein receptors. Results: We determined crystal structure of bovine coronavirus NTD and located its sugar-binding site using mutagenesis. Conclusion: Bovine coronavirus NTD shares structural folds

  10. Reconstitution of bovine A1 adenosine receptors and G proteins in phospholipid vesicles: betagamma-subunit composition influences guanine nucleotide exchange and agonist binding.

    PubMed

    Figler, R A; Lindorfer, M A; Graber, S G; Garrison, J C; Linden, J

    1997-12-23

    We have studied the interactions of purified A1 adenosine receptors and G proteins reconstituted into phospholipid vesicles to investigate how the betagamma composition of G protein heterotrimers influences coupling. Recombinant hexahistidine-tagged bovine A1 adenosine receptors were expressed in Sf9 cells and purified to homogeneity by sequential chromatography over heparin-sepharose, xanthine amino congener-agarose, and nickel-nitrilotriacetic acid columns. These receptors were reconstituted with pure recombinant G proteins of defined subunit composition. Receptor-G protein complexes containing alphai2 and beta1gamma2 or beta1gamma3 and stimulated with the agonist, (R)-phenylisopropyladenosine, exchange guanine nucleotide 2-3 times more rapidly than do complexes containing beta1gamma1. This difference is not overcome by increasing the concentration of betagamma subunits. Receptor-G protein complexes containing beta1gamma1 also bind less of the agonist, [125I]-iodoaminobenzyladenosine (125I-ABA), than do complexes containing beta1gamma3. Kinetic experiments show that 125I-ABA dissociates 2-fold more rapidly from receptor-G protein complexes containing beta1gamma1 than from complexes containing the other betagamma subunits. The affinity of the interaction between immobilized Galphai2 subunits and beta1gamma1 or beta1gamma2 measured with an optical biosensor in the absence of receptor is similar. Taken together, these data implicate the gamma-subunit in influencing the interaction between the A1 adenosine receptor and G proteins. PMID:9405064

  11. Effective Hard Particle Model for the Osmotic Pressure of Highly Concentrated Binary Protein Solutions

    PubMed Central

    Minton, Allen P.

    2008-01-01

    The experimentally measured concentration dependence of the osmotic pressure of an equimolar mixture of hen egg ovalbumin and bovine serum albumin at pH 7.0 and 25°C in the presence of 0.15 M NaCl is shown to be quantitatively accounted for by a model in which each protein species is represented by an effective hard sphere. The size of this sphere is determined by analysis of the concentration dependence of the osmotic pressure of the isolated protein. PMID:18212007

  12. Influence of the pore fluid on the phase velocity in bovine trabecular bone In Vitro: Prediction of the biot model

    NASA Astrophysics Data System (ADS)

    Lee, Kang Il

    2013-01-01

    The present study aims to investigate the influence of the pore fluid on the phase velocity in bovine trabecular bone in vitro. The frequency-dependent phase velocity was measured in 20 marrow-filled and water-filled bovine femoral trabecular bone samples. The mean phase velocities at frequencies between 0.6 and 1.2 MHz exhibited significant negative dispersions for both the marrow-filled and the water-filled samples. The magnitudes of the dispersions showed no significant differences between the marrow-filled and the water-filled samples. In contrast, replacement of marrow by water led to a mean increase in the phase velocity of 27 m/s at frequencies from 0.6 to 1.2 MHz. The theoretical phase velocities of the fast wave predicted by using the Biot model for elastic wave propagation in fluid-saturated porous media showed good agreements with the measurements.

  13. Single and binary adsorption of proteins on ion-exchange adsorbent: The effectiveness of isothermal models.

    PubMed

    Liang, Juan; Fieg, Georg; Shi, Qing-Hong; Sun, Yan

    2012-09-01

    Simultaneous and sequential adsorption equilibria of single and binary adsorption of bovine serum albumin and bovine hemoglobin on Q Sepharose FF were investigated in different buffer constituents and initial conditions. The results in simultaneous adsorption showed that both proteins underwent competitive adsorption onto the adsorbent following greatly by protein-surface interaction. Preferentially adsorbed albumin complied with the universal rule of ion-exchange adsorption whereas buffer had no marked influence on hemoglobin adsorption. Moreover, an increase in initial ratios of proteins was benefit to a growth of adsorption density. In sequential adsorption, hemoglobin had the same adsorption densities as single-component adsorption. It was attributed to the displacement of preadsorbed albumin and multiple layer adsorption of hemoglobin. Three isothermal models (i.e. extended Langmuir, steric mass-action, and statistical thermodynamic (ST) models) were introduced to describe the ion-exchange adsorption of albumin and hemoglobin mixtures. The results suggested that extended Langmuir model gave the lowest deviation in describing preferential adsorption of albumin at a given salt concentration while steric mass-action model could very well describe the salt effect in albumin adsorption. For weaker adsorbed hemoglobin, ST model was the preferred choice. In concert with breakthrough data, the research further revealed the complexity in ion-exchange adsorption of proteins. PMID:22888059

  14. Eliminating bovine tuberculosis in cattle and badgers: insight from a dynamic model

    PubMed Central

    Brooks-Pollock, Ellen; Wood, James L. N.

    2015-01-01

    Bovine tuberculosis (BTB) is a multi-species infection that commonly affects cattle and badgers in Great Britain. Despite years of study, the impact of badgers on BTB incidence in cattle is poorly understood. Using a two-host transmission model of BTB in cattle and badgers, we find that published data and parameter estimates are most consistent with a system at the threshold of control. The most consistent explanation for data obtained from cattle and badger populations includes within-host reproduction numbers close to 1 and between-host reproduction numbers of approximately 0.05. In terms of controlling infection in cattle, reducing cattle-to-cattle transmission is essential. In some regions, even large reductions in badger prevalence can have a modest impact on cattle infection and a multi-stranded approach is necessary that also targets badger-to-cattle transmission directly. The new perspective highlighted by this two-host approach provides insight into the control of BTB in Great Britain. PMID:25972466

  15. Effect of bovine lactoferrin in a therapeutic hamster model of hepatic amoebiasis.

    PubMed

    Ordaz-Pichardo, Cynthia; León-Sicairos, Nidia; Hernández-Ramírez, Verónica Ivonne; Talamás-Rohana, Patricia; de la Garza, Mireya

    2012-06-01

    Entamoeba histolytica is the causative agent of amoebiasis, a disease that produces dysentery as a result of the perforation of the large intestine. This parasite often invades other organs, primarily the liver, leading to an amoebic liver abscess (ALA), which can cause death. Metronidazole is the drug of choice for the treatment of ALA; however, it produces toxic side effects in patients. Lactoferrin (Lf) is a glycoprotein of the innate immune response that sequesters iron in the mucosae. Lf possesses immune-regulatory properties, such as antiinflammatory and antioxidant activities. Moreover, the microbicidal activity of apoLf, which lacks bound iron, has been shown. In this study, we evaluated the therapeutic effect of bovine Lf (bLf) against ALA in a model of hepatic amoebiasis in hamsters. Interestingly, hamsters treated intragastrically with Lf (2.5 mg/100 g mass) over a period of 8 days showed no clinical signs of disease and ALA was effectively decreased, with only 0.63% detectable lesion, compared with 63% in untreated animals. Furthermore, liver function and blood cells approached normal levels among those receiving bLf treatment. These results suggest that bLf may aid in the therapy of amoebiasis, likely without producing undesirable effects in patients. PMID:22332957

  16. Use of topical bovine thrombin in an anti-coagulated rat model of hepatic injury.

    PubMed

    Schmiedt, Chad W; Köhler, Rickard; Brainard, Benjamin M

    2012-12-01

    The need for surgical hemostasis in patients treated with anticoagulant medications is a concern. This study assessed a bovine-derived topical hemostat (FastAct, FA) using a partial hepatectomy hemorrhage model in anticoagulated rats. Ninety rats were randomly assigned to receive preoperative heparin, warfarin, or nothing (n=30/treatment). Within each treatment group, FA, saline, direct pressure (DP), electrocautery, or nothing (n=6/group) was applied to the hepatectomy site. Eight additional rats were used for assessment of the preoperative anticoagulant regimen. Rats that were not anticoagulated and received FA had faster clot times and less hemorrhage than those receiving DP (P<0.05). In warfarin-pretreated rats, FA resulted in faster coagulation times than saline or DP and less hemorrhage than saline (P<0.05). No differences were detected in heparinized rats. Across all groups, rats receiving FA lost less blood and formed clots more frequently than saline (P<0.05). FA may be useful to treat hemorrhage from hepatic lacerations in anticoagulated patients. PMID:22633173

  17. Mechanisms of ocular toxicity using the in vitro bovine lens and sodium dodecyl sulfate as a chemical model.

    PubMed

    Bantseev, Vladimir; McCanna, David; Banh, Alice; Wong, Winnie W; Moran, Kelley L; Dixon, D George; Trevithick, John R; Sivak, Jacob G

    2003-05-01

    Previous work using the in vitro bovine lens as a model has shown a correlation between toxicity and lens optical function and showed much higher sensitivity in detecting irritancy of several surfactants at much lower concentrations than the Draize score. In the current study, cultured bovine lenses were used to study the effects of the surfactant sodium dodecyl sulfate (SDS) on lens optical properties and mitochondrial integrity. Bovine lenses were exposed to SDS (0.1 to 0.00625%) for 30 min and cultured for 24 h. Compared to controls (n = 17), loss of sharp focus was evident immediately following exposure to 0.1% SDS (n = 14, p < 0.0001). At 24 h loss of sharp focus became evident in all groups. Loss of lens transparency, significant increase in lens wet weight, and axial length were seen 24 h postexposure in lenses treated with 0.1 to 0.025% SDS. Confocal analysis 24 h postexposure showed SDS concentration-dependent decrease in number and length of the mitochondria in lens epithelial and superficial cortical fiber cells. The results of this study show a correlation between lens optical properties and metabolic function and together provide a sensitive in vitro model of ocular chemical toxicity. Results of confocal analysis suggest that the mitochondrial integrity of the superficial cortical fiber cells is most sensitive to damage caused by SDS. The results further suggest that recovery of lens metabolic function is necessary for the recovery of lens optical properties. PMID:12700424

  18. Stochastic simulation modeling to determine time to detect Bovine Viral Diarrhea antibodies in bulk tank milk.

    PubMed

    Foddai, Alessandro; Enøe, Claes; Krogh, Kaspar; Stockmarr, Anders; Halasa, Tariq

    2014-11-01

    A stochastic simulation model was developed to estimate the time from introduction of Bovine Viral Diarrhea Virus (BVDV) in a herd to detection of antibodies in bulk tank milk (BTM) samples using three ELISAs. We assumed that antibodies could be detected, after a fixed threshold prevalence of seroconverted milking cows was reached in the herd. Different thresholds were set for each ELISA, according to previous studies. For each test, antibody detection was simulated in small (70 cows), medium (150 cows) and large (320 cows) herds. The assays included were: (1) the Danish blocking ELISA, (2) the SVANOVIR(®)BVDV-Ab ELISA, and (3) the ELISA BVD/MD p80 Institute Pourquier. The validation of the model was mainly carried out by comparing the predicted incidence of persistently infected (PI) calves and the predicted detection time, with records from a BVD infected herd. Results showed that the SVANOVIR, which was the most efficient ELISA, could detect antibodies in the BTM of a large herd 280 days (95% prediction interval: 218; 568) after a transiently infected (TI) milking cow has been introduced into the herd. The estimated time to detection after introduction of one PI calf was 111 days (44; 605). With SVANOVIR ELISA the incidence of PIs and dead born calves could be limited and the impact of the disease on the animal welfare and income of farmers (before detection) could be minimized. The results from the simulation modeling can be used to improve the current Danish BVD surveillance program in detecting early infected herds. PMID:25081944

  19. Exploring the affinity binding of alkylmaltoside surfactants to bovine serum albumin and their effect on the protein stability: A spectroscopic approach.

    PubMed

    Hierrezuelo, J M; Carnero Ruiz, C

    2015-08-01

    Steady-state and time-resolved fluorescence together with circular dichroism (CD) spectroscopic studies was performed to examine the interactions between bovine serum albumin (BSA) and two alkylmaltoside surfactants, i.e. n-decyl-?-d-maltoside (?-C10G2) and n-dodecyl-?-d-maltoside (?-C12G2), having identical structures but different tail lengths. Changes in the intrinsic fluorescence of BSA from static as well as dynamic measurements revealed a weak protein-surfactant interaction and gave the corresponding binding curves, suggesting that the binding mechanism of surfactants to protein is essentially cooperative in nature. The behavior of both surfactants is similar, so that the differences detected were attributed to the more hydrophobic nature of ?-C12G2, which favors the adsorption of micelle-like aggregates onto the protein surface. These observations were substantially demonstrated by data derived from synchronous, three-dimensional and anisotropy fluorescence experiments. Changes in the secondary structure of the protein induced by the interaction with surfactants were analyzed by CD to determine the contents of ?-helix and ?-strand. It was noted that whereas the addition of ?-C10G2 appears to stabilize the secondary structure of the protein, ?-C12G2 causes a marginal denaturation of BSA for a protein:surfactant molar ratio as high as 1 to 100. PMID:26042703

  20. Geranylgeranylated Rab proteins terminating in Cys-Ala-Cys, but not Cys-Cys, are carboxyl-methylated by bovine brain membranes in vitro.

    PubMed Central

    Smeland, T E; Seabra, M C; Goldstein, J L; Brown, M S

    1994-01-01

    Geranylgeranylated Rab proteins usually terminate in either Cys-Cys or Cys-Xaa-Cys, where Xaa is Ala, Ser, or Gly. In both classes of proteins, the two cysteines are geranylgeranylated, but only the Cys-Xaa-Cys class has been shown to be carboxyl-methylated on the terminal cysteine in vivo. In the current study, we used recombinant Rab geranylgeranyltransferase and a Rab escort protein (REP-1) to attach geranylgeranyl residues to the two cysteines at the carboxyl terminus of Rab3A (Cys-Ala-Cys) and Rab1A (Cys-Cys). The geranylgeranylated proteins were then incubated with bovine cerebellar membranes that contain an enzyme that transfers [3H]methyl from S-[methyl-3H]adenosyl-L-methionine to geranylgeranylated cysteine. The enzyme transferred [3H]methyl to geranylgeranylated Rab3A but not to geranylgeranylated Rab1A. Replacement of the Cys-Ala-Cys terminus of Rab3A with Cys-Cys abolished methylation, and the opposite result was obtained when the Cys-Cys of Rab1A was replaced with Cys-Ala-Cys. When the Cys-Cys terminus of Rab1A was changed to Ser-Cys, methylation was restored. These studies suggest that the carboxyl-terminal cysteine of Rab proteins terminating in Cys-Xaa-Cys but not Cys-Cys is methylated and that the resistance of Cys-Cys proteins to methylation is attributable to the vicinal geranylgeranylated cysteines. Images PMID:7938016

  1. Identification of three antigen epitopes on the nucleocapsid protein of the genotype C of bovine parainfluenza virus type 3.

    PubMed

    Ren, Jian-Le; Zhu, Yuan-Mao; Zhou, Yue-Hui; Lv, Chuang; Yan, Hao; Ma, Lei; Shi, Hong-Fei; Xue, Fei

    2015-07-01

    Bovine parainfluenza virus type 3 (BPIV3) is an important respiratory tract pathogen for both young and adult cattle. So far, three genotypes A, B and C of BPIV3 have been described on the basis of genetic and phylogenetic analysis. But fine mapping of epitopes of BPIV3 is scant and the antigenic variations among the three genotypes of BPIV3 have not been reported. Nucleocapsid protein (NP) is the most abundant protein in the virion and highly conserved in BPIV3, which is crucial for the induction of protective immunity in host. To identify antigenic determinants of BPIV3 NP, a panel of monoclonal antibodies (mAbs) was tested against a series of overlapping recombinant NP fragments expressed in Escherichia coli. Firstly, six monoclonal antibodies (mAbs) against NP of the genotype C of BPIV3 (BPIV3c) were generated by using the purified BPIV3c strain SD0835 as immunogen and the recombinant NP of SD0835 as screening antigen. Then three antigen epitopes were identified with the six mAbs. One epitope (91)GNNADVKYVIYM(102) was recognized by mAb 5E5. The mAbs 7G5, 7G8, 7G9, and 7H5 were reactive with another epitope (407)FYKPTGG(413). The third epitope (428)ESRGDQDQ(435) was reactive with mAb 6F8. Further analysis showed that the epitope (91-102 amino acids [aa]) was the most conserved and reactive with mAb 5E5 for all three genotypes of BPIV3 and HPIV3. The epitope (407-413 aa) was relatively conserved and reactive with mAbs 7G5, 7G8, 7G9, and 7H5 for BPIV3a, BPIV3c and HPIV3, but not reactive with BPIV3b. The epitope (428-435 aa) was less conserved and was reactive only with mAb 6F8 for BPIV3a and BPIV3c. These results suggested that there were evident antigenic variations among the three genotypes of BPIV3 and HPIV3. The mAb 6F8 could be used to detect BPIV3a and BPIV3c. The mAbs 7G5, 7G8, 7G9, and 7H5 might be used for differentiate BPIV3a, BPIV3c and HPIV3 from BPIV3b. The mAb 5E5 might be used for detecting all three types of BPIV3 and HPIV3. The results in this study would have potential applications in the development of suitable diagnostic techniques for BPIV3, which was prevalent in China. PMID:25960335

  2. Adsorption-induced conformational changes of proteins onto ceramic particles: Differential scanning calorimetry and FTIR analysis

    Microsoft Academic Search

    Natascha Brandes; Petra B. Welzel; Carsten Werner; Lothar W. Kroh

    2006-01-01

    Three model proteins, bovine serum albumin, hen's egg lysozyme and bovine serum fibrinogen, were adsorbed from aqueous solution onto finely dispersed ceramic particles, namely different kinds of alumina and hydroxyapatite particles. The influence of adsorption on protein secondary structure was investigated. The FTIR spectroscopic findings were compared with the results of DSC measurements. In almost all cases it was found

  3. Small Noncoding RNAs Encoded within the Bovine Herpesvirus 1 Latency-Related Gene Can Reduce Steady-State Levels of Infected Cell Protein 0 (bICP0)?

    PubMed Central

    Jaber, Tareq; Workman, Aspen; Jones, Clinton

    2010-01-01

    Following acute infection in mucosal epithelium, bovine herpes virus 1 (BHV-1) establishes lifelong latency in sensory neurons within trigeminal ganglia. The latency-related RNA (LR-RNA) is abundantly expressed in sensory neurons of latently infected calves. Expression of LR proteins is necessary for the latency reactivation cycle because a mutant virus that does not express LR proteins is unable to reactivate from latency after dexamethasone treatment. LR-RNA sequences also inhibit bICP0 expression, productive infection, and cell growth. However, it is unclear how LR-RNA mediates these functions. In this study, we identified a 463-bp region within the LR gene (the XbaI-PstI [XP] fragment) that inhibited bICP0 protein and RNA expression in transiently transfected mouse neuroblastoma cells. Small noncoding RNAs (sncRNAs) encoded within the XP fragment (20 to 90 nucleotides in length) were detected in transiently transfected mouse neuroblastoma cells. Two families of sncRNAs were cloned from this region, and each family was predicted to contain a mature microRNA (miRNA). Both miRNAs were predicted to base pair with bICP0 mRNA sequences, suggesting that they reduce bICP0 levels. To test this prediction, sequences encompassing the respective sncRNAs and mature miRNAs were synthesized and cloned into a small interfering RNA expression vector. Both sncRNA families and their respective miRNAs inhibited bICP0 protein expression in mouse neuroblastoma cells and productive infection in bovine cells. In trigeminal ganglia of latently infected calves, an sncRNA that migrated between nucleotides 20 and 25 hybridized to the XP fragment. During dexamethasone-induced reactivation from latency, XP-specific sncRNA levels were reduced, suggesting that these sncRNAs support the establishment and maintenance of lifelong latency in cattle. PMID:20410286

  4. Estrogen-specific sulfotransferase (SULT1E1) in bovine placentomes: inverse levels of mRNA and protein in uninucleated trophoblast cells and trophoblast giant cells.

    PubMed

    Polei, Marina; Viergutz, Torsten; Tomek, Wolfgang; Schuler, Gerhard; Fürbass, Rainer

    2014-08-01

    The bovine trophoblast produces significant amounts of estrogens. In maternal and fetal blood, estrogens occur predominantly in sulfonated forms, which are unable to bind to estrogen receptors (ESRs). However, estrogens may act as local factors in ESR-positive trophoblast cells or in the adjacent caruncular epithelium, which in addition to ESR highly expresses steroid sulfatase. Estrogen sulfonation is catalyzed by the cytosolic enzyme SULT1E1. Previous studies clearly indicated the trophoblast as the primary site of estrogen sulfonation. However, investigations into the cellular localization of SULT1E1 yielded conflicting results. In situ hybridization studies detected SULT1E1 mRNA only in trophoblast giant cells (TGCs), whereas in immunohistochemical experiments the SULT1E1 protein was virtually restricted to uninucleated trophoblast cells (UTCs). The aim of this work was to resolve this conflict by analyzing SULT1E1 expression in isolated UTCs and TGCs. Highly enriched pools of UTCs and TGCs were obtained from four bovine placentas (Days 118-130 of gestation) using an optimized fluorescence-activated cell sorting procedure. UTC and TGC pools were analyzed by quantitative RT-PCR and Western blot experiments to measure the amounts of SULT1E1 transcript and protein, respectively. In contrast to previously published results, both SULT1E1 transcript and SULT1E1 protein were clearly present in the UTC and TGC pools. However, some evidence indicated a higher transcript concentration in TGCs and a higher amount of protein in UTCs. Thus, our results resolve the conflicting results on the localization of SULT1E1 from earlier studies and suggest that posttranscriptional mechanisms play an important role in the control of SULT1E1 expression during TGC differentiation. PMID:25009209

  5. Modeling of bovine spongiform encephalopathy in a two-species feedback loop.

    PubMed

    Barnes, Richard; Lehman, Clarence

    2013-06-01

    Bovine spongiform encephalopathy, otherwise known as mad cow disease, can spread when an individual cow consumes feed containing the infected tissues of another individual, forming a one-species feedback loop. Such feedback is the primary means of transmission for BSE during epidemic conditions. Following outbreaks in the European Union and elsewhere, many governments enacted legislation designed to limit the spread of such diseases via elimination or reduction of one-species feedback loops in agricultural systems. However, two-species feedback loops-those in which infectious material from one-species is consumed by a secondary species whose tissue is then consumed by the first species-were not universally prohibited and have not been studied before. Here we present a basic ecological disease model which examines the rôle feedback loops may play in the spread of BSE and related diseases. Our model shows that there are critical thresholds between the infection's expansion and decrease related to the lifespan of the hosts, the growth rate of the prions, and the amount of prions circulating between hosts. The ecological disease dynamics can be intrinsically oscillatory, having outbreaks as well as refractory periods which can make it appear that the disease is under control while it is still increasing. We show that non-susceptible species that have been intentionally inserted into a feedback loop to stop the spread of disease do not, strictly by themselves, guarantee its control, though they may give that appearance by increasing the refractory period of an epidemic's oscillations. We suggest ways in which age-related dynamics and cross-species coupling should be considered in continuing evaluations aimed at maintaining a safe food supply. PMID:23746801

  6. Antiviral Efficacy of a Respiratory Syncytial Virus (RSV) Fusion Inhibitor in a Bovine Model of RSV Infection.

    PubMed

    Jordan, Robert; Shao, Matt; Mackman, Richard L; Perron, Michel; Cihlar, Tomas; Lewis, Sandy A; Eisenberg, Eugene J; Carey, Anne; Strickley, Robert G; Chien, Jason W; Anderson, Mark L; McEligot, Heather A; Behrens, Nicole E; Gershwin, Laurel J

    2015-08-01

    Respiratory syncytial virus (RSV) is the leading cause of bronchiolitis and pneumonia in infants. Effective treatment for RSV infection is a significant unmet medical need. While new RSV therapeutics are now in development, there are very few animal models that mimic the pathogenesis of human RSV, making it difficult to evaluate new disease interventions. Experimental infection of Holstein calves with bovine RSV (bRSV) causes a severe respiratory infection that is similar to human RSV infection, providing a relevant model for testing novel therapeutic agents. In this model, viral load is readily detected in nasal secretions by quantitative real-time PCR (qRT-PCR), and cumulative symptom scoring together with histopathology evaluations of infected tissue allow for the assessment of disease severity. The bovine RSV model was used to evaluate the antiviral activity of an RSV fusion inhibitor, GS1, which blocks virus entry by inhibiting the fusion of the viral envelope with the host cell membrane. The efficacy of GS1, a close structural analog of GS-5806 that is being developed to treat RSV infection in humans was evaluated in two randomized, blind, placebo-controlled studies in bRSV-infected calves. Intravenous administration of GS1 at 4 mg/kg of body weight/day for 7 days starting 24 h or 72 h postinoculation provided clear therapeutic benefit by reducing the viral load, disease symptom score, respiration rate, and lung pathology associated with bRSV infection. These data support the use of the bovine RSV model for evaluation of experimental therapeutics for treatment of RSV. PMID:26055364

  7. Adipogenesis of bovine perimuscular preadipocytes

    SciTech Connect

    Taniguchi, Masaaki; Le Luo Guan; Zhang Bing [Department of Agricultural, Food and Nutritional Science, University of Alberta, 410 Ag-For Centre, Edmonton, Alta., T6G 2P5 (Canada); Dodson, Michael V. [Department of Animal Sciences, Washington State University, P.O. Box 646310, Pullman, WA 99164 (United States); Okine, Erasmus [Department of Agricultural, Food and Nutritional Science, University of Alberta, 410 Ag-For Centre, Edmonton, Alta., T6G 2P5 (Canada); Moore, Stephen S. [Department of Agricultural, Food and Nutritional Science, University of Alberta, 410 Ag-For Centre, Edmonton, Alta., T6G 2P5 (Canada)], E-mail: Stephen.moore@ualberta.ca

    2008-02-01

    In this study, non-transformed progeny adipofibroblasts, derived from mature adipocyte dedifferentiation, was used as a novel in vitro model to study adipogenic gene expression in cattle. Adipofibroblasts from dedifferentiated mature perimuscular fat (PMF) tissue were cultured with differentiation stimulants until the cells exhibited morphological differentiation. Treated cells were harvested from day 2 to 16 for RNA extraction, whereas control cells were cultured without addition of stimulants. Results from time course gene expression assays by quantitative real-time PCR revealed that peroxisome proliferator-activated receptor gamma (PPAR-{gamma}), sterol regulatory element binding protein 1 (SREBP-1) and their six down-stream genes were co-expressed at day 2 post-differentiation induction. When compared to other adipogenesis culture systems, the adipogenic gene expression of bovine PMF adipofibroblasts culture was different, especially to the rodent model. Collectively, these results demonstrated PPAR-{gamma} and SREBP-1 cooperatively play a key role to regulate the re-differentiation of bovine adipofibroblasts, during early conversion stages in vitro.

  8. Evidence for protein kinase C in bovine adrenocortical membrane preparations using (/sup 35/S) gamma-thio-ATP as a phosphate donor

    SciTech Connect

    Coyne, M.D.; Luszczynska, H.M.; Kunzi, M.

    1987-01-01

    Thio-substituted ATP is a sensitive probe for detecting protein kinase C activity as demonstrated in bovine adrenocortical cell membrane preparations. A single endogenous protein substrate with a molecular weight of approximately 47 Kd was rapidly phosphorylated with (/sup 35/S) gamma-thio-ATP as phosphate donor. Phosphorylation was significantly increased in 30 seconds and reached a plateau by 3 minutes. The activity of the endogenous membrane kinase was unaffected by ACTH, cAMP, calmodulin or trifluoperazine but was responsive to combinations of calcium (Ca), diolein and phosphatidyl serine (PS). In addition, the kinase was activated by the tumor promoting phorbol ester, 12-0-tetradecanoylphorbol-13-acetate, indicating that the membrane contains a protein kinase C and a single 47 Kd phosphorylatable protein substrate. The same substrate is phosphorylated by Ca/diolein/PS activated kinase in membrane preparations from a broad range of rat tissues. Attempts to identify the substrate indicate that it is neither the type I regulatory subunit of cAMP dependent protein kinase nor mitochondrial cytochrome P450.

  9. Characterization of aggregation and protein expression of bovine corneal endothelial cells as microcarrier cultures in a rotating-wall vessel

    Microsoft Academic Search

    James W. Muhitch; Kim C. O'Connor; Diane A. Blake; Daniel J. Lacks; Nitsa Rosenzweig; Glenn F. Spaulding

    2000-01-01

    Rotating-wall vessels are beneficial to tissue engineering in that the reconstituted tissue formed in these low-shear bioreactors undergoes extensive three-dimensional growth and differentiation. In the present study, bovine corneal endothelial (BCE) cells were grown in a high-aspect rotating-wall vessel (HARV) attached to collagen-coated Cytodex-3 beads as a representative monolayer culture to investigate factors during HARV cultivation which affect three-dimensional growth

  10. The mechanisms of uremic serum-induced expression of bone matrix proteins in bovine vascular smooth muscle cells

    Microsoft Academic Search

    N X Chen; D Duan; K D O'Neill; G O Wolisi; J J Koczman; R LaClair; S M Moe

    2006-01-01

    We have previously found that uremic human serum upregulates RUNX2 in vascular smooth muscle cells (VSMCs), and that RUNX2 is upregulated in areas of vascular calcification in vivo. To confirm the role of RUNX2, we transiently transfected a dominant-negative RUNX2 (?RUNX2) construct in bovine vascular smooth muscle cells (BVSMCs). Blocking RUNX2 transcriptional activity significantly decreased uremic serum induced alkaline phosphatase

  11. Modelling protein–protein interaction networks via a stickiness index

    PubMed Central

    Pržulj, Nataša; Higham, Desmond J

    2006-01-01

    What type of connectivity structure are we seeing in protein–protein interaction networks? A number of random graph models have been mooted. After fitting model parameters to real data, the models can be judged by their success in reproducing key network properties. Here, we propose a very simple random graph model that inserts a connection according to the degree, or ‘stickiness’, of the two proteins involved. This model can be regarded as a testable distillation of more sophisticated versions that attempt to account for the presence of interaction surfaces or binding domains. By computing a range of network similarity measures, including relative graphlet frequency distance, we find that our model outperforms other random graph classes. In particular, we show that given the underlying degree information, fitting a stickiness model produces better results than simply choosing a degree-matching graph uniformly at random. Therefore, the results lend support to the basic modelling methodology. PMID:16971339

  12. Reactivity of monoclonal antibodies against a tryptophan-riboflavin adduct toward irradiated and non-irradiated bovine-eye-lens protein fractions: an indicator of in vivo visible-light-mediated phototransformations.

    PubMed

    Mancini, M; Edwards, A M; Becker, M I; de Ioannes, A; Silva, E

    2000-03-01

    We describe here the reactivity toward the soluble protein of bovine eye lens of anti-tryptophan-riboflavin (anti-Trp-RF) adduct monoclonal antibodies, which recognize the hapten tryptophan-riboflavin generated by irradiation of a solution of bovine serum albumin in the presence of riboflavin. It is demonstrated that five different anti-Trp-RF adduct monoclonal antibodies, all belonging to the IgG1 isotype, react with the total soluble proteins of bovine eye lens. The components of the soluble protein are separated by Sephadex G-200 chromatography and the isolated fractions analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). All the separated protein fractions also react by a direct ELISA with the monoclonal antibodies; this reaction is more intense when the isolated fractions have been previously irradiated with visible light in the presence of riboflavin under an atmosphere of oxygen or nitrogen. Irradiation of the total soluble protein with visible light in the presence of riboflavin produces the appearance of new bands, corresponding to compounds of higher molecular weight. Riboflavin-sensitized irradiation of the protein fractions with visible light under an oxygen or nitrogen atmosphere is accompanied by a concomitant decrease of the tryptophan fluorescence. It is postulated that the action of visible light in the presence of either the endogenous riboflavin or its derivatives could be partly responsible for the protein aggregation observed during aging. PMID:10877061

  13. The development of a bovine interspecies model for the analysis of genomic imprinting in normal and nuclear transfer derived fetuses

    E-print Network

    Dindot, Scott Victor

    2004-11-15

    ..........................................................52 2.9 PipMaker dot plot schematic of the bovine and mouse XIST/Xist locus...............53 2.10 Schematic of CpG island prediction in exon 1 of the bovine and mouse XIST/Xist locus......................................................................................................55 2.11 Comparative sequence analysis of the ? 45 region of the XIST/Xist promoter between the mouse, bovine, human, horse and rabbit..........................................57 2.12 Amplification of the bovine XIST CpG island in exon 1...

  14. A proteomic study of the differential protein expression in MDBK cells after bovine herpesvirus type 1 infection (BHV-1) strain treatment

    PubMed Central

    Guo, Li; Yang, Yanling; Liu, Linna; Liao, Peng; Wen, Yongjun; Wu, Hua; Cheng, Shipeng

    2015-01-01

    Different BHV-1 strains, such as the virulent IBRV LN01/08 strains and the attenuated vaccine strain IBRV LNM, produces different clinical immune responses; however, the study of the differential protein expression in Madin-Darby bovine kidney (MDBK) cells after BHV-1-infection still remains unclear. Here, we applied a comparative proteomic strategy, based on 2D and MALDI-TOF/MS platforms, to examine the differential expression of proteins in MDBK cells that were treated and not treated with virulent IBRV LN01/08 and attenuated IBRV LNM strains. A total of eight differential proteins, including pyruvate kinase, heat shock protein (HSP) 90 (HSP90AA1 and HSP90AB1), annexin A, albumin (ALB), scinderin (SCIN), tubulin (alpha 1a) and vimentin (VIM), were identified. Among these proteins, pyruvate kinase, and HSP90 (HSP90AB1), tubulin and vimentin were identified in the virulent IBRV LN01/08 strain group, but were not identified in the attenuated IBRV LNM group. These results play an important role in tumor formation and development, cell migration, tumor cell line apoptosis, cell invasion and viral infection. The HSP90 (HSP90AA1) protein was identified in the control group and the attenuated IBRV LNM-infected group. Most studies have shown that HSP90 proteins were more of a cancer gene target, and inhibiting its function would result to oncogene degradation during cancer treatment. On the other hand, ALB is associated to cell differentiation, apoptosis, necrosis, cell death, viral infection, autophagy, interstitial tissue inflammation, and cell survival. These results provide a theoretical basis for the systematic understanding of BHV-1-infection mechanisms and BHV-1-induced immune responses.

  15. Impact of external sources of infection on the dynamics of bovine tuberculosis in modelled badger populations

    PubMed Central

    2012-01-01

    Background The persistence of bovine TB (bTB) in various countries throughout the world is enhanced by the existence of wildlife hosts for the infection. In Britain and Ireland, the principal wildlife host for bTB is the badger (Meles meles). The objective of our study was to examine the dynamics of bTB in badgers in relation to both badger-derived infection from within the population and externally-derived, trickle-type, infection, such as could occur from other species or environmental sources, using a spatial stochastic simulation model. Results The presence of external sources of infection can increase mean prevalence and reduce the threshold group size for disease persistence. Above the threshold equilibrium group size of 6–8 individuals predicted by the model for bTB persistence in badgers based on internal infection alone, external sources of infection have relatively little impact on the persistence or level of disease. However, within a critical range of group sizes just below this threshold level, external infection becomes much more important in determining disease dynamics. Within this critical range, external infection increases the ratio of intra- to inter-group infections due to the greater probability of external infections entering fully-susceptible groups. The effect is to enable bTB persistence and increase bTB prevalence in badger populations which would not be able to maintain bTB based on internal infection alone. Conclusions External sources of bTB infection can contribute to the persistence of bTB in badger populations. In high-density badger populations, internal badger-derived infections occur at a sufficient rate that the additional effect of external sources in exacerbating disease is minimal. However, in lower-density populations, external sources of infection are much more important in enhancing bTB prevalence and persistence. In such circumstances, it is particularly important that control strategies to reduce bTB in badgers include efforts to minimise such external sources of infection. PMID:22738118

  16. Chronic wasting disease and atypical forms of bovine spongiform encephalopathy and scrapie are not transmissible to mice expressing wild-type levels of human prion protein.

    PubMed

    Wilson, Rona; Plinston, Chris; Hunter, Nora; Casalone, Cristina; Corona, Cristiano; Tagliavini, Fabrizio; Suardi, Silvia; Ruggerone, Margherita; Moda, Fabio; Graziano, Silvia; Sbriccoli, Marco; Cardone, Franco; Pocchiari, Maurizio; Ingrosso, Loredana; Baron, Thierry; Richt, Juergen; Andreoletti, Olivier; Simmons, Marion; Lockey, Richard; Manson, Jean C; Barron, Rona M

    2012-07-01

    The association between bovine spongiform encephalopathy (BSE) and variant Creutzfeldt-Jakob disease (vCJD) has demonstrated that cattle transmissible spongiform encephalopathies (TSEs) can pose a risk to human health and raises the possibility that other ruminant TSEs may be transmissible to humans. In recent years, several novel TSEs in sheep, cattle and deer have been described and the risk posed to humans by these agents is currently unknown. In this study, we inoculated two forms of atypical BSE (BASE and H-type BSE), a chronic wasting disease (CWD) isolate and seven isolates of atypical scrapie into gene-targeted transgenic (Tg) mice expressing the human prion protein (PrP). Upon challenge with these ruminant TSEs, gene-targeted Tg mice expressing human PrP did not show any signs of disease pathology. These data strongly suggest the presence of a substantial transmission barrier between these recently identified ruminant TSEs and humans. PMID:22495232

  17. Imino sugars inhibit the formation and secretion of bovine viral diarrhea virus, a pestivirus model of hepatitis C virus: Implications for the development of broad spectrum anti-hepatitis virus agents

    PubMed Central

    Zitzmann, Nicole; Mehta, Anand S.; Carrouée, Sandra; Butters, Terry D.; Platt, Frances M.; McCauley, John; Blumberg, Baruch S.; Dwek, Raymond A.; Block, Timothy M.

    1999-01-01

    One function of N-linked glycans is to assist in the folding of glycoproteins by mediating interactions of the lectin-like chaperone proteins calnexin and calreticulin with nascent glycoproteins. These interactions can be prevented by inhibitors of the ?-glucosidases, such as N-butyl-deoxynojirimycin (NB-DNJ) and N-nonyl-DNJ (NN-DNJ), and this causes some proteins to be misfolded and retained within the endoplasmic reticulum (ER). We have shown previously that the NN-DNJ-induced misfolding of one of the hepatitis B virus (HBV) envelope glycoproteins prevents the formation and secretion of virus in vitro and that this inhibitor alters glycosylation and reduces the viral levels in an animal model of chronic HBV infection. This led us to investigate the effect of glucosidase inhibitors on another ER-budding virus, bovine viral diarrhea virus, a tissue culture surrogate of human hepatitis C virus (HCV). Here we show that in MDBK cells ?-glucosidase inhibitors prevented the formation and secretion of infectious bovine viral diarrhea virus. Data also are presented showing that NN-DNJ, compared with NB-DNJ, exhibits a prolonged retention in liver in vivo. Because viral secretion is selectively hypersensitive to glucosidase inhibition relative to the secretion of cellular proteins, the possibility that glucosidase inhibitors could be used as broad-based antiviral hepatitis agents is discussed. A single drug against HBV, HCV, and, possibly, HDV, which together chronically infect more than 400 million people worldwide, would be of great therapeutic value. PMID:10518544

  18. Measurements of bovine albumin as a model system for the development of a hand-held ellipsometer for ophthalmic applications

    NASA Astrophysics Data System (ADS)

    Miran Baygi, M. H.; Payne, P. A.

    2000-06-01

    Scattering ellipsometry is a measurement technique that can extract the characteristics of particles with relative ease. For some while, we have been interested in the use of scattering ellipsometry to examine the particles in the anterior chamber of the human eye. These particles are globular proteins, typically some 6 or 7 nm in diameter and measurements conducted at a wavelength of 670 nm confirm this. Particles or particle aggregates much greater in size will give rise to characteristic patterns in the ellipsometric results and analysis of these patterns could lead to a complete description of the physical properties of such particles. Examples of measurements of this nature made on latex particles are included. The results obtained from the measurements described confirm that bovine serum albumin is suitable as a calibrant for a scattering ellipsometer.

  19. Enhancement of Asynchronous and Train-Evoked Exocytosis in Bovine Adrenal Chromaffin Cells Infected with a Replication Deficient Adenovirus

    Microsoft Academic Search

    Ramachandran Thiagarajan; Jennifer Wilhelm; Teclemichael Tewolde; Yingjie Li; Mark M. Rich; Kathrin L. Engisch

    2005-01-01

    Bovine adrenal chromaffin cells share many characteristics with neurons and are often used as a simple model system to study ion channels and neurotransmitter release. We infected bovine adrenal chromaffin cells with a replication deficient adenovirus that induces expression of the common reporters beta-galactosidase and Green Fluorescent Protein via a bicistronic sequence. In perforated-patch recordings performed 48-h postinfection, peak calcium

  20. Brain protein kinase C phosphorylating poly(arginine,serine) or lamin B is stimulated by anions and by an activator purified from bovine serum albumin preparations.

    PubMed Central

    Abdel-Ghany, M; el-Gendy, K; Zhang, S; Raden, D; Racker, E

    1989-01-01

    The phosphorylation of histone by purified protein kinase C (PK-C) from rat brain is dependent on the presence of Ca2+ and lipids. Phosphorylation of a synthetic random polymer of arginine and serine (3:1) is only moderately enhanced by Ca2+ and lipids, but it is greatly enhanced in the absence of Ca2+ and lipids by a contaminant in crystalline bovine serum albumin or by heated cellular fractions. The phosphorylation ratio of histone to poly(arginine,serine) varies between different PK-C fractions from brains of rat, pig, or lamb. These variations are partly caused by a PK-C isozyme that prefers poly(arginine,serine) over histone as substrate. The kinase activator (KA) was partly purified from bovine serum albumin and from extracts of plasma membranes of human placenta. KA is also present in mitochondria, nuclei, and the cytosol. Sulfates and phosphates at 10 mM substitute for KA with poly(arginine,serine) as substrate. The phosphorylation of histone III in the presence of Ca2+ and lipids is moderately stimulated by KA, but the phosphorylation of lamin B and some other endogenous proteins is greatly enhanced by KA. With histones as substrates, inorganic anions do not stimulate phosphorylation. The phosphorylation of poly-(arginine,serine) is very sensitive to low concentrations of staurosporin and is inhibited by PK-C antibody, but, in contrast to histone phosphorylation, it is resistant to sphingosine and polymyxin B. The poly(arginine,serine) phosphorylating activity is more stable at 4 degrees C than the histone phosphorylating activity, but the latter is stabilized by 0.05% Triton X-100. Images PMID:2928301

  1. Enhanced Virulence of Sheep-Passaged Bovine Spongiform Encephalopathy Agent Is Revealed by Decreased Polymorphism Barriers in Prion Protein Conversion Studies

    PubMed Central

    Priem, Jan; Langeveld, Jan P. M.; van Keulen, Lucien J. M.; van Zijderveld, Fred G.; Andreoletti, Olivier

    2014-01-01

    ABSTRACT Bovine spongiform encephalopathy (BSE) can be efficiently transmitted to small ruminants (sheep and goats) with certain prion protein (PrP) genotypes. Polymorphisms in PrP of both the host and donor influence the transmission efficiency of transmissible spongiform encephalopathies (TSEs) in general. These polymorphisms in PrP also modulate the PrP conversion underlying TSE agent replication. Here we demonstrate that single-round protein misfolding cyclic amplification (PMCA) can be used to assess species and polymorphism barriers at the molecular level. We assessed those within and between the ovine and bovine species in vitro using a variety of natural scrapie and experimentally generated cross-species BSE agents. These BSE agents include ovBSE-ARQ isolates (BSE derived from sheep having the ARQ/ARQ PrP genotype), and two unique BSE-derived variants: BSE passaged in VRQ/VRQ sheep and a cow BSE agent isolate generated by back-transmission of ovBSE-ARQ into its original host. PMCA allowed us to quantitatively determine PrP conversion profiles that correlated with known in vivo transmissibility and susceptibility in the two ruminant species in which strain-specific molecular signatures, like its molecular weight after protease digestion, were maintained. Furthermore, both BSE agent isolates from ARQ and VRQ sheep demonstrated a surprising transmission profile in which efficient transmissions to both sheep and bovine variants was combined. Finally, all data support the notion that ARQ-derived sheep BSE points to a significant increase in virulence compared to all other tested scrapie- and BSE-derived variants reflected by the increased conversion efficiencies of previously inefficient convertible PrP variants (including the so-called “resistant” sheep ARR variant). IMPORTANCE Prion diseases such as scrapie in sheep and goats, BSE in cattle, and Creutzfeldt-Jakob disease (CJD) in humans are fatal neurodegenerative diseases caused by prions. BSE is known to be transmissible to a variety of hosts, including sheep and humans. Based on the typical BSE agent strain signatures and epidemiological data, the occurrence of a novel variant of CJD in humans was linked to BSE occurrence in the United Kingdom. Measures, including genetic selection of sheep toward less susceptible PrP genotypes, have been implemented to lower the risk of BSE transmission into sheep, since the disease could potentially spread into a natural reservoir. In this study, we demonstrated using molecular PrP conversion studies that when BSE is first transmitted through sheep, the host range is modified significantly and the PrP converting potency increased, allowing the ovine BSE to transmit more efficiently than cow BSE into supposedly less susceptible hosts. PMID:24371051

  2. Efficacy of a Novel Whey Protein Gel Complex to Increase the Unsaturated Fatty Acid Composition of Bovine Milk Fat

    Microsoft Academic Search

    S. M. Carroll; E. J. DePeters; M. Rosenberg

    2006-01-01

    A novel whey protein emulsion gel (WPEG) complex was developed to protect dietary unsaturated fatty acids from rumen biohydrogenation with the goal of modifying the fatty acid composition of milk fat. Three experiments were conducted with WPEG complexes made from either whey protein concentrate containing 80% crude protein, whey protein isolate, or whey pro- tein concentrate high-gel capacity. Each experiment

  3. Stochastic model for protein flexibility analysis

    NASA Astrophysics Data System (ADS)

    Xia, Kelin; Wei, Guo-Wei

    2013-12-01

    Protein flexibility is an intrinsic property and plays a fundamental role in protein functions. Computational analysis of protein flexibility is crucial to protein function prediction, macromolecular flexible docking, and rational drug design. Most current approaches for protein flexibility analysis are based on Hamiltonian mechanics. We introduce a stochastic model to study protein flexibility. The essential idea is to analyze the free induction decay of a perturbed protein structural probability, which satisfies the master equation. The transition probability matrix is constructed by using probability density estimators including monotonically decreasing radial basis functions. We show that the proposed stochastic model gives rise to some of the best predictions of Debye-Waller factors or B factors for three sets of protein data introduced in the literature.

  4. Technical note: a pilot study using a mouse mastitis model to study differences between bovine associated coagulase-negative staphylococci.

    PubMed

    Breyne, K; De Vliegher, S; De Visscher, A; Piepers, S; Meyer, E

    2015-02-01

    Coagulase-negative staphylococci (CNS) are a group of bacteria classified as either minor mastitis pathogens or commensal microbiota. Recent research suggests species- and even strain-related epidemiological and genetic differences within the large CNS group. The current pilot study investigated in 2 experiments whether a mouse mastitis model validated for bovine Staphylococcus aureus can be used to explore further differences between CNS species and strains. In a first dose titration experiment, a low inoculum dose of S. aureus Newbould 305 (positive control) was compared with increasing inoculum doses of a Staphylococcus chromogenes strain originating from a chronic bovine intramammary infection to a sham-inoculated mammary glands (negative control). In contrast to the high bacterial growth following inoculation with S. aureus, S. chromogenes was retrieved in very low levels at 24 h postinduction (p.i.). In a second experiment, the inflammation inflicted by 3 CNS strains was studied in mice. The host immune response induced by the S. chromogenes intramammary strain was compared with the one induced by a Staphylococcus fleurettii strain originating from cow bedding sawdust and by a S. chromogenes strain originating from a teat apex of a heifer. As expected, at 28 and 48 h p.i., low bacterial growth and local neutrophil influx in the mammary gland were induced by all CNS strains. As hypothesized, bacterial growth p.i. was the lowest for S. fleurettii compared with that induced by the 2 S. chromogenes strains, and the overall immune response established by the 3 CNS strains was less pronounced compared with the one induced by S. aureus. Proinflammatory cytokine profiling revealed that S. aureus locally induced IL-6 and IL-1? but not TNF-?, whereas, overall, CNS-inoculated glands lacked a strong cytokine host response but also induced IL-1? locally. Compared with both other CNS strains, S. chromogenes from the teat apex inflicted a more variable IL-1? response characterized by a more intense local reaction in several mice. This pilot study suggests that an intraductal mouse model can mimic bovine CNS mastitis and has potential as a complementary in vivo tool for future CNS mastitis research. Furthermore, it indicates that epidemiologically different bovine CNS species or strains induce a differential host innate immune response in the murine mammary gland. PMID:25497801

  5. The effect of the number of observations used for Fourier transform infrared model calibration for bovine milk fat composition on the estimated genetic parameters of the predicted data

    Microsoft Academic Search

    M. J. M. Rutten; H. Bovenhuis; Arendonk van J. A. M

    2010-01-01

    Fourier transform infrared spectroscopy is a suitable method to determine bovine milk fat composition. However, the determination of fat composition by gas chromatography, required for calibration of the infrared prediction model, is expensive and labor intensive. It has recently been shown that the number of calibration samples is strongly related to the model's validation r2 (i.e., accuracy of prediction). However,

  6. Relationships of the phase velocity with the microarchitectural parameters in bovine trabecular bone in vitro: Application of a stratified model

    NASA Astrophysics Data System (ADS)

    Lee, Kang Il

    2012-08-01

    The present study aims to provide insight into the relationships of the phase velocity with the microarchitectural parameters in bovine trabecular bone in vitro. The frequency-dependent phase velocity was measured in 22 bovine femoral trabecular bone samples by using a pair of transducers with a diameter of 25.4 mm and a center frequency of 0.5 MHz. The phase velocity exhibited positive correlation coefficients of 0.48 and 0.32 with the ratio of bone volume to total volume and the trabecular thickness, respectively, but a negative correlation coefficient of -0.62 with the trabecular separation. The best univariate predictor of the phase velocity was the trabecular separation, yielding an adjusted squared correlation coefficient of 0.36. The multivariate regression models yielded adjusted squared correlation coefficients of 0.21-0.36. The theoretical phase velocity predicted by using a stratified model for wave propagation in periodically stratified media consisting of alternating parallel solid-fluid layers showed reasonable agreements with the experimental measurements.

  7. Evolution of Bovine Respiratory Syncytial Virus

    Microsoft Academic Search

    JEAN-FRANCOIS VALARCHER; FRANCOIS SCHELCHER; H. Bourhy

    2000-01-01

    Until now, the analysis of the genetic diversity of bovine respiratory syncytial virus (BRSV) has been based on small numbers of field isolates. In this report, we determined the nucleotide and deduced amino acid sequences of regions of the nucleoprotein (N protein), fusion protein (F protein), and glycoprotein (G protein) of 54 European and North American isolates and compared them

  8. The Escherichia coli O157:H7 cattle immunoproteome includes outer membrane protein A (OmpA), a modulator of adherence to bovine rectoanal junction squamous epithelial (RSE) cells.

    PubMed

    Kudva, Indira T; Krastins, Bryan; Torres, Alfredo G; Griffin, Robert W; Sheng, Haiqing; Sarracino, David A; Hovde, Carolyn J; Calderwood, Stephen B; John, Manohar

    2015-06-01

    Building on previous studies, we defined the repertoire of proteins comprising the immunoproteome (IP) of Escherichia coli O157:H7 (O157) cultured in DMEM supplemented with norepinephrine (O157 IP), a ?-adrenergic hormone that regulates E. coli O157 gene expression in the gastrointestinal tract, using a variation of a novel proteomics-based platform proteome mining tool for antigen discovery, called "proteomics-based expression library screening" (PELS; Kudva et al., 2006). The E. coli O157 IP (O157-IP) comprised 91 proteins, and included those identified previously using proteomics-based expression library screening, and also proteins comprising DMEM and bovine rumen fluid proteomes. Outer membrane protein A (OmpA), a common component of the above proteomes, and reportedly a contributor to E. coli O157 adherence to cultured HEp-2 epithelial cells, was interestingly found to be a modulator rather than a contributor to E. coli O157 adherence to bovine rectoanal junction squamous epithelial cells. Our results point to a role for yet to be identified members of the O157-IP in E. coli O157 adherence to rectoanal junction squamous epithelial cells, and additionally implicate a possible role for the outer membrane protein A regulator, TdcA, in the expression of such adhesins. Our observations have implications for the development of efficacious vaccines for preventing E. coli O157 colonization of the bovine gastrointestinal tract. PMID:25643951

  9. Learning Generative Models for Protein Fold Families

    E-print Network

    Langmead, Christopher James

    to CJL. #12;Keywords: Structure Learning, Generative Models, Probabilistic Graphical Models, Proteins #12 the network of interactions within a protein is sparse, it is richer than previously believed. #12 into the constraints that govern structure, function, and dynamics. These constraints can reflect both position

  10. Identification of the bovine herpesvirus 1 circ protein, a myristylated and virion-associated polypeptide which is not essential for virus replication in cell culture.

    PubMed Central

    Fraefel, C; Ackermann, M; Schwyzer, M

    1994-01-01

    We have recently reported immediate-early (IE) transcription over covalently joined genome ends of bovine herpesvirus 1 (BHV-1). A spliced 1.5-kb IE RNA (IER1.5) is coterminal with an unspliced 1.1-kb late RNA (LR1.1) which is transcribed from the left end of the genome. Sequence analysis reveals an open reading frame common to IER1.5 and LR1.1 predicted to encode the 247-amino-acid circ polypeptide. This paper reports on the identification of circ as a protein. Using a rabbit antiserum raised against a synthetic oligopeptide representing the carboxy terminus of the predicted circ polypeptide for Western blot (immunoblot) analyses and immunofluorescence assays, we identified a 34-kDa virion-associated protein which accumulated in the cytoplasm of infected cells. To confirm that LR1.1 indeed encoded the 34-kDa polypeptide, we inserted a DNA fragment containing circ coding sequences into the Autographa californica baculovirus genome. A group of recombinant polypeptides with sizes of 32, 34, and 35 kDa were identified by their reactivity with the antipeptide serum. Chicken egg yolk antibodies raised against total proteins of insect cells infected with the recombinant baculovirus identified the 34-kDa circ protein specified by BHV-1. The recombinant circ polypeptides and the circ protein specified by BHV-1 were both myristylated, as determined by radiolabeling with [3H]myristic acid. It was noted that the circ gene could be deleted from the BHV-1 genome without impairing virus replication in cell culture. Images PMID:7966598

  11. Hidden Markov Models in Computational Biology Applications to Protein Modeling

    Microsoft Academic Search

    Anders Krogh; Michael Brown; I. Saira

    1994-01-01

    Ridden .Markov Models (HMMs) are applied t.0 the problems of statistical modeling, database searching and multiple sequence alignment of protein families and protein domains. These methods are demonstrated on the globin family, the protein kinase catalytic domain, and the EF-hand calcium binding motif. In each case the parameters of an HMM are estimated from a training set of unaligned sequences.

  12. A soft and transparent handleable protein model

    NASA Astrophysics Data System (ADS)

    Kawakami, Masaru

    2012-08-01

    The field of structural biology currently relies on computer-generated graphical representations of three-dimensional (3D) structures to conceptualize biomolecules. As the size and complexity of the molecular structure increases, model generation and peer discussions become more difficult. It is even more problematic when discussing protein-protein interactions wherein large surface area contact is considered. This report demonstrates the viability of a new handleable protein molecular model with a soft and transparent silicone body similar to the molecule's surface. A full-color printed main chain structure embedded in the silicone body enables users to simultaneously feel the molecular surface, view through the main chain structure, and manually simulate molecular docking. The interactive, hands-on experience deepens the user's intuitive understanding of the complicated 3D protein structure and elucidates ligand binding and protein-protein interactions. This model would be an effective discussion tool for the classroom or laboratory that stimulates inspired learning in this study field.

  13. Interaction between neurotrophin 4 and gonadotrophin in bovine oviducts.

    PubMed

    Sun, Yongfeng; Zhang, Jing; Li, Chunjin; Wang, Dong; Ma, Yonghe; Sun, Yanling; Liu, Zhuo; Wang, Chunqiang; Zhou, Xu

    2012-07-01

    The expression and localization of neurotrophin 4 (NT4) and its receptor, tyrosine kinase B (TRKB), in the bovine oviduct, and their interaction with gonadotrophins in bovine oviduct epithelial cells (BOECs), were examined. Transcripts for NT4 and TRKB were detected by reverse transcription polymerase chain reaction (RT-PCR) in bovine oviducts in the follicular and luteal phases, and their proteins were immunolocalized in BOECs. Based on real time PCR, NT4 mRNA did not differ significantly between the two phases of the cycle, although TRKB mRNA expression was higher (P < 0.05) in the luteal phase than that in follicular phase. The BOECs were treated with various concentrations of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in vitro; for NT4, mRNA and protein were higher (P < 0.05) than those in the control (based on real time PCR and enzyme-linked immunosorbent assay (ELISA) assays). The effects of NT4 and the TRKB inhibitor (K252a) on the expression of LH receptor (LHR) and FSH receptor (FSHR) in the oviduct epithelial cells were also studied using a monolayer culture model. Expression levels of LHR and FSHR mRNA in BOECs treated with various concentrations of NT4 were higher (P < 0.05) than those in the control. However, these expressions were blocked by treatment with K252?. We concluded that neurotrophin 4 may have a role in regulating the function of bovine oviducts by interacting with gonadotrophins. PMID:22460152

  14. Statistical models of protein-ligand interactions and protein geometry

    NASA Astrophysics Data System (ADS)

    Dewitte, Robert Simon

    In this thesis, several examples are presented wherein complexity-first, or statistical, modelling provides keen insight into the physics of complex molecular systems as well as technological advance. In the first part, where the system of interest is the general protein-ligand complex, coarse-graining of a knowledge-base is shown to be a successful approach to modelling the interaction free energy of binding. Specifically, when the coarse graining length scale corresponds to the correlation length of induced solvent order due to solute presence, the coarse-grained potential adequately incorporates solvent entropy changes upon binding into a simple, fast and accurate binding free energy estimate. This estimate is comparable to the best results of other approaches, but is computable in a fraction of the time. Moreover, the coarse-graining effectively smooths the interaction potential energy surface such that the space of possible ligands can be spanned by an efficient Metropolis Monte Carlo growth algorithm. This algorithm generates novel ligands de novo in a matter of seconds. The combination of algorithm and knowledge-based potential is demonstrated to be able to derive several interesting small molecule ligand candidates with minimal human intervention. Furthermore, the smoothed potential surface of interaction paves the way for the introduction of a novel, efficient and rigorous conformational search procedure that provides phenomenal speedup over simple grid search methods. In the second part, where the system of interest is the general protein global fold, the enumeration of restrained self-avoiding random walks demonstrates that very little residue-residue contact information is needed to specify a protein global fold. In particular, on the order of one restraint per monomer suffices to limit the number of protein global folds to of order one. Using a hierarchical model of inter-residue contacts, a general expression is derived for an upper bound for the number of constraints needed to specify the global fold uniquely. This result is discussed in light of protein structure reconstruction from Nuclear Overhauser Magnetic Resonance Experiments. In the third part, where the system of interest is the general problem of secondary structure selection in protein backbones, consideration of the abstract pseudodihedral angle (angle describing abstract torsions along the vector path connecting protein alpha-carbons) reveals an a priori angular selection depending on residue identity. This observation led to the development of a knowledge-based potential which has possible applications in secondary structure prediction, protein homology modelling (sequence and structure), as well as protein folding simulations. The pseudodihedral was also applied to criticism of various lattice models as reduced representations of proteins.

  15. Evaluation of protein-protein docking model structures using all-atom molecular dynamics simulations combined with the solution theory in the energy representation

    NASA Astrophysics Data System (ADS)

    Takemura, Kazuhiro; Guo, Hao; Sakuraba, Shun; Matubayasi, Nobuyuki; Kitao, Akio

    2012-12-01

    We propose a method to evaluate binding free energy differences among distinct protein-protein complex model structures through all-atom molecular dynamics simulations in explicit water using the solution theory in the energy representation. Complex model structures are generated from a pair of monomeric structures using the rigid-body docking program ZDOCK. After structure refinement by side chain optimization and all-atom molecular dynamics simulations in explicit water, complex models are evaluated based on the sum of their conformational and solvation free energies, the latter calculated from the energy distribution functions obtained from relatively short molecular dynamics simulations of the complex in water and of pure water based on the solution theory in the energy representation. We examined protein-protein complex model structures of two protein-protein complex systems, bovine trypsin/CMTI-1 squash inhibitor (PDB ID: 1PPE) and RNase SA/barstar (PDB ID: 1AY7), for which both complex and monomer structures were determined experimentally. For each system, we calculated the energies for the crystal complex structure and twelve generated model structures including the model most similar to the crystal structure and very different from it. In both systems, the sum of the conformational and solvation free energies tended to be lower for the structure similar to the crystal. We concluded that our energy calculation method is useful for selecting low energy complex models similar to the crystal structure from among a set of generated models.

  16. Evaluation of protein-protein docking model structures using all-atom molecular dynamics simulations combined with the solution theory in the energy representation.

    PubMed

    Takemura, Kazuhiro; Guo, Hao; Sakuraba, Shun; Matubayasi, Nobuyuki; Kitao, Akio

    2012-12-01

    We propose a method to evaluate binding free energy differences among distinct protein-protein complex model structures through all-atom molecular dynamics simulations in explicit water using the solution theory in the energy representation. Complex model structures are generated from a pair of monomeric structures using the rigid-body docking program ZDOCK. After structure refinement by side chain optimization and all-atom molecular dynamics simulations in explicit water, complex models are evaluated based on the sum of their conformational and solvation free energies, the latter calculated from the energy distribution functions obtained from relatively short molecular dynamics simulations of the complex in water and of pure water based on the solution theory in the energy representation. We examined protein-protein complex model structures of two protein-protein complex systems, bovine trypsin/CMTI-1 squash inhibitor (PDB ID: 1PPE) and RNase SA/barstar (PDB ID: 1AY7), for which both complex and monomer structures were determined experimentally. For each system, we calculated the energies for the crystal complex structure and twelve generated model structures including the model most similar to the crystal structure and very different from it. In both systems, the sum of the conformational and solvation free energies tended to be lower for the structure similar to the crystal. We concluded that our energy calculation method is useful for selecting low energy complex models similar to the crystal structure from among a set of generated models. PMID:23231264

  17. Analysis of a bovine herpesvirus 1 protein encoded by an alternatively spliced latency related (LR) RNA that is abundantly expressed in latently infected neurons.

    PubMed

    Sinani, Devis; Liu, Yilin; Jones, Clinton

    2014-09-01

    The bovine herpes virus 1 (BoHV-1) encoded latency-related RNA (LR-RNA) is abundantly expressed in latently infected sensory neurons. A LR mutant virus with three stop codons at the amino-terminus of ORF2 does not reactivate from latency or replicate efficiently in certain tissues. ORF2 inhibits apoptosis, interacts with Notch1 or Notch3, and interferes with Notch mediated signaling. Alternative splicing of LR-RNA in trigeminal ganglia yields transcripts that have the potential to encode a protein containing most of ORF2 sequences and parts of other coding sequences located within the LR gene. In this study, we determined that an ORF2 protein fused with reading frame B (15d ORF) was more stable in transfected cells. ORF2 and the 15d ORF stimulated neurite formation in mouse neuroblastoma cells, interfered with Notch3 mediated trans-activation, and had similar DNA binding properties. Increased stability of the 15d ORF is predicted to enhance the establishment of latency. PMID:25104616

  18. Models of globular proteins in aqueous solutions

    NASA Astrophysics Data System (ADS)

    Wentzel, Nathaniel James

    Protein crystallization is a continuing area of research. Currently, there is no universal theory for the conditions required to crystallize proteins. A better understanding of protein crystallization will be helpful in determining protein structure and preventing and treating certain diseases. In this thesis, we will extend the understanding of globular proteins in aqueous solutions by analyzing various models for protein interactions. Experiments have shown that the liquid-liquid phase separation curves for lysozyme in solution with salt depend on salt type and salt concentration. We analyze a simple square well model for this system whose well depth depends on salt type and salt concentration, to determine the phase coexistence surfaces from experimental data. The surfaces, calculated from a single Monte Carlo simulation and a simple scaling argument, are shown as a function of temperature, salt concentration and protein concentration for two typical salts. Urate Oxidase from Asperigillus flavus is a protein used for studying the effects of polymers on the crystallization of large proteins. Experiments have determined some aspects of the phase diagram. We use Monte Carlo techniques and perturbation theory to predict the phase diagram for a model of urate oxidase in solution with PEG. The model used includes an electrostatic interaction, van der Waals attraction, and a polymerinduced depletion interaction. The results agree quantitatively with experiments. Anisotropy plays a role in globular protein interactions, including the formation of hemoglobin fibers in sickle cell disease. Also, the solvent conditions have been shown to play a strong role in the phase behavior of some aqueous protein solutions. Each has previously been treated separately in theoretical studies. Here we propose and analyze a simple, combined model that treats both anisotropy and solvent effects. We find that this model qualitatively explains some phase behavior, including the existence of a lower critical point under certain conditions.

  19. Regeneration of bovine and octopus opsins in situ with natural and artificial retinals

    SciTech Connect

    Koutalos, Y.; Ebrey, T.G.; Tsuda, M.; Odashima, K.; Lien, T.; Park, M.H.; Shimizu, N.; Derguini, F.; Nakanishi, K.; Gilson, H.R.; Honig, B. (Univ. of Illinois, Urbana-Champaign (USA))

    1989-03-21

    The authors consider the problem of color regulation in visual pigments for both bovine rhodopsin and octopus rhodopsin. Both pigments have 11-cis-retinal as their chromophore. These rhodopsins were bleached in their native membranes, and the opsins were regenerated with natural and artificial chromophores. Both bovine and octopus opsins were regenerated with the 9-cis- and 11-cis-retinal isomers, but the octopus opsin was additionally regenerated with the 13-cis and all-trans isomers. Titration of the octopus opsin with 11-cis-retinal gave an extinction coefficient for octopus rhodopsin of 27,000 {plus minus} 3,000 M{sup {minus}1} cm{sup {minus}1} at 475 nm. The absorption maxima of bovine artificial pigments formed by regenerating opsin with the 11-cis dihydro series of chromophores support a color regulation model for bovine rhodopsin in which the chromophore-binding site of the protein has two negative charges: one directly hydrogen bonded to the Schiff base nitrogen and another near carbon-13. Formation of octopus artificial pigments with both all-trans and 11-cis dihydro chromophores leads to a similar model for octopus rhodopsin and metarhodopsin: there are two negative charges in the chromophore-binding site, one directly hydrogen bonded to the Schiff base nitrogen and a second near carbon-13. The interaction of this second charge with the chromophore in octopus rhodopsin is weaker than in bovine, while in metarhodopsin it is as strong as in bovine.

  20. SWISS-MODEL: an automated protein homology-modeling server

    Microsoft Academic Search

    Torsten Schwede; Jürgen Kopp; Nicolas Guex; Manuel C. Peitsch

    2003-01-01

    SWISS-MODEL (http:\\/\\/swissmodel.expasy.org) is a server for automated comparative modeling of three- dimensional (3D) protein structures. It pioneered the field of automated modeling starting in 1993 and is the most widely-used free web-based automated modeling facility today. In 2002 the server computed 120 000 user requests for 3D protein models. SWISS- MODEL provides several levels of user interaction through its World

  1. Changes in 131I-Labelled Immune Bovine gamma -globulin during Transmission to the Circulation after Oral Adminstration to the Young Rat

    Microsoft Academic Search

    F. W. R. Brambell; R. Halliday; W. A. Hemmings

    1961-01-01

    Bovine gamma -globulin, trace-labelled with 131I, reaches the circulation in substantial amounts although bacterial agglutinins from bovine antisera appear to be completely excluded. Thus when the bovine globulin is both isotopically labelled and immune, the labelled protein which reaches the circulation is still precipitable by antisera to bovine globulin but is without antibody activity. When the labelled bovine gamma -globulin

  2. Phase diagrams of a crystalline membrane protein, bovine heart cytochrome c oxidase, in the salting-in region

    NASA Astrophysics Data System (ADS)

    Ataka, Mitsuo; Shinzawa-Itoh, Kyoko; Yoshikawa, Shinya

    1992-08-01

    Phase diagrams were determined for cytochrome c oxidase crystals, based on the protein concentration changes with time in the supernatant due to crystal growth. The solubility of the protein decreases when the concentration of sodium phosphate buffer, pH 7.4, decreases in the range of 0.5mM-10mM, as well as when the concentration of Brij-35, a detergent stabilizing this enzyme, increases in the examined range (0%-14%). The precipitated protein was all crystalline without any amorphous materials. These results indicate a "salting-in" of the enzyme due to the phosphate ion at any fixed Brij-35 concentration. Thus, this protein is most readily crystallized either by addition of Brij-35 or by concentration of the protein solution without changing the buffer concentration (with an ultrafiltration apparatus). Nucleation and crystal growth rates were also determined. Instantaneous nucleation always occurred even when the initial protein concentration was only 24% higher than the solubility of the crystal. The present results suggest that the mechanism of crystal growth of large membrane proteins is significantly different from that of small water-soluble proteins.

  3. Stretching lattice models of protein folding

    PubMed Central

    Socci, Nicholas D.; Onuchic, José Nelson; Wolynes, Peter G.

    1999-01-01

    A new class of experiments that probe folding of individual protein domains uses mechanical stretching to cause the transition. We show how stretching forces can be incorporated in lattice models of folding. For fast folding proteins, the analysis suggests a complex relation between the force dependence and the reaction coordinate for folding. PMID:10051589

  4. Denaturation of bovine spleen galectin-1 in guanidine hydrochloride and fluoroalcohols: structural characterization and implications for protein folding.

    PubMed

    Mandal, Pritha; Molla, Anisur R; Mandal, Dipak K

    2013-12-01

    Guanidine hydrochloride (GdnHCl)-induced unfolding of bovine spleen galectin-1 (Gal-1) exhibits three-state mechanism involving exclusive, structured tertiary monomer in 0.5 M GdnHCl. Gal-1 has one tryptophan residue (Trp 68) per subunit. Phosphorescence spectra of both Gal-1 dimer and intermediate monomer at 77 K show single (0,0) band at 405.6 nm, characteristics of free tryptophan environment as of N-acetyl-l-tryptophanamide. Unfolded Gal-1 in 4 M GdnHCl gives (0,0) band at longer wavelength (408.6 nm) signifying that Trp 68 is less solvent exposed, being localized in an environment of residual structure. Trifluoroethanol (TFE)- and hexafluoroisopropanol (HFIP)-induced structural changes of Gal-1 dimer and monomer have been investigated by far-UV CD and FTIR. CD results show reversible nature of ?-sheet to ?-helix transition, with 30% helix in 80% TFE or 40% HFIP for Gal-1 dimer. Temperature-dependent studies show that induced helix entails reduced thermal stability. FTIR results reveal partial ?-sheet to ?-helix conversion but with quantitative yield. At intermediate TFE concentration, both Gal-1 dimer and monomer aggregate as evidenced by FTIR band at ?1617/cm, however, the onset of aggregation and stability of aggregates for monomer differ from those of dimer. The results may provide important insights into perturbed folding problem of Gal-1. PMID:24037640

  5. Monolith immuno-affinity enrichment liquid chromatography tandem mass spectrometry for quantitative protein analysis of recombinant bovine somatotropin in serum.

    PubMed

    Smits, Nathalie G E; Blokland, Marco H; Wubs, Klaas L; Nessen, Merel A; van Ginkel, Leen A; Nielen, Michel W F

    2015-08-01

    The use of recombinant bovine somatotropin (rbST) to enhance milk production is approved in several countries, but it is prohibited in the European Union. According to EU legislation, it is necessary to confirm positive screening results prior to enforcement. Although adequate screening assays are available nowadays, development of liquid chromatography tandem mass spectrometry (LC-MS/MS) confirmatory methods to detect low levels of rbST is still a challenge. Here, we present a novel approach using immuno-affinity enrichment on monolithic micro-columns in combination with state-of-the-art ultra-high pressure LC-MS/MS (UHPLC-MS/MS) detection. The developed approach enables detection and confirmation of rbST in serum at a decision limit (CC?) concentration of 0.8 ng mL(-1). Furthermore, the method is easy to handle, robust and reproducible. We successfully applied the confirmatory method to serum samples from rbST treated cows that were found suspect after immunoassay-based screening. The use of rbST could be confirmed over 1 week after treatment, and the developed method demonstrated the sensitivity needed for effective control. Graphical Abstract Graphical summary of the workflow, for serum preparation, enrichment with monolith microcolumns and LC-MS/MS measurement of rbST. PMID:26077745

  6. Quantitative thermodynamic model for globular protein folding

    NASA Astrophysics Data System (ADS)

    Yakubovich, Alexander V.; Solov'yov, Andrey V.

    2014-06-01

    We present a statistical mechanics formalism for theoretical description of the process of protein folding ? unfolding transition in water environment. The formalism is based on the construction of the partition function of a protein obeying two-stage-like folding kinetics. Using the statistical mechanics model of solvation of hydrophobic hydrocarbons we obtain the partition function of infinitely diluted solution of proteins in water environment. The calculated dependencies of the protein heat capacities upon temperature are compared with the corresponding results of experimental measurements for staphylococcal nuclease and metmyoglobin.

  7. Predicting protein structure using hidden Markov models

    E-print Network

    Karplus, Kevin

    Predicting protein structure using hidden Markov models Kevin Karplusy Kimmen Sjolanderz Christian Santa Cruz, CA 95064 USA abstract We discuss how methods based on hidden Markov models performed in the fold recogni- tion section of the CASP2 experiment. Hidden Markov models were built for a set of about

  8. MicroRNA regulation of bovine monocyte inflammatory and metabolic networks in an in vivo infection model

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine mastitis is an inflammation-driven disease of the bovine mammary gland that costs the global dairy industry several billion dollars per annum. Because disease susceptibility is a multi-factorial complex phenotype, a multi-omic integrative biology approach is required to dissect the multilayer...

  9. Cultured bovine brain capillary endothelial cells (BBCEC) - a blood-brain barrier model for studying the binding and internalization of insulin and insulin-like growth factor 1

    SciTech Connect

    Keller, B.T.; Borchardt, R.T.

    1987-05-01

    Cultured bovine brain capillary endothelial cells (BBCEC) have previously been reported by their laboratory as a working model for studying nutrient and drug transport and metabolism at the blood-brain barrier. In the present study, they have utilized this culture system to investigate the binding and internalization of (/sup 125/I)-labelled insulin (INS) and insulin-like growth factor 1(IGF-1) by BBCEC. After 2 hrs at 23/sup 0/C, the specific binding of INS and IGF-1 was 1.6% and 13.6%, respectively. At 37/sup 0/C, the maximum specific binding was 0.9% for INS and 5.8% for IGF-1. Using an acid-wash technique to assess peptide internalization, it was observed that, at 37/sup 0/C, approximately 60% of the bound INS rapidly became resistant to acid treatment, a value which was constant over 2 hr. With IGF-1, a similar proportion of the bound material, 62%, became resistant by 30 min, but subsequently decreased to 45% by 2 hr. Scatchard analysis of competitive binding studies indicated the presence of two binding sites for each protein, having K/sub d/'s of 0.82 nM and 19.2 nM for INS and 0.39 nM and 3.66 nM for IGF-1. Little change in the amount of INS binding was observed over a four-day interval as the cultures became a confluent monolayer. The present report of binding and internalization of these proteins suggests that the BBCEC may utilize a receptor-mediated process to internalize and/or transport (transcytosis) INS and IGF-1 from the circulation.

  10. Mathematical analysis of a model for the growth of the bovine corpus luteum.

    PubMed

    Prokopiou, Sotiris A; Byrne, Helen M; Jeffrey, Mike R; Robinson, Robert S; Mann, George E; Owen, Markus R

    2014-12-01

    The corpus luteum (CL) is an ovarian tissue that grows in the wound space created by follicular rupture. It produces the progesterone needed in the uterus to maintain pregnancy. Rapid growth of the CL and progesterone transport to the uterus require angiogenesis, the creation of new blood vessels from pre-existing ones, a process which is regulated by proteins that include fibroblast growth factor 2 (FGF2). In this paper we develop a system of time-dependent ordinary differential equations to model CL growth. The dependent variables represent FGF2, endothelial cells (ECs), luteal cells, and stromal cells (like pericytes), by assuming that the CL volume is a continuum of the three cell types. We assume that if the CL volume exceeds that of the ovulated follicle, then growth is inhibited. This threshold volume partitions the system dynamics into two regimes, so that the model may be classified as a Filippov (piecewise smooth) system. We show that normal CL growth requires an appropriate balance between the growth rates of luteal and stromal cells. We investigate how angiogenesis influences CL growth by considering how the system dynamics depend on the dimensionless EC proliferation rate, ??. We find that weak (low ??) or strong (high ??) angiogenesis leads to 'pathological' CL growth, since the loss of CL constituents compromises progesterone production or delivery. However, for intermediate values of ??, normal CL growth is predicted. The implications of these results for cow fertility are also discussed. For example, inadequate angiogenesis has been linked to infertility in dairy cows. PMID:24337679

  11. Cyperus rotundus suppresses AGE formation and protein oxidation in a model of fructose-mediated protein glycoxidation.

    PubMed

    Ardestani, Amin; Yazdanparast, Razieh

    2007-12-01

    Non-enzymatic glycation, as the chain reaction between reducing sugars and the free amino groups of proteins, has been shown to correlate with severity of diabetes and its complications. Cyperus rotundus (Cyperaceae) is used both as a food to promote health and as a drug to treat certain diseases. In this study, considering the antioxidative effects of C. rotundus, we examined whether C. rotundus also protects against protein oxidation and glycoxidation. The protein glycation inhibitory activity of hydroalcoholic extract of C. rotundus was evaluated in vitro using a model of fructose-mediated protein glycoxidation. The C. rotundus extract with glycation inhibitory activity also demonstrated antioxidant activity when a ferric reducing antioxidant power (FRAP) and Trolox equivalent antioxidant capacity (TEAC) assays as well as metal chelating activity were applied. Fructose (100mM) increased fluorescence intensity of glycated bovine serum albumin (BSA) in terms of total AGEs during 14 days of exposure. Moreover, fructose caused more protein carbonyl (PCO) formation and also oxidized thiol groups more in glycated than in native BSA. The extract of C. rotundus at different concentrations (25-250microg/ml) has significantly decreased the formation of AGEs in term of the fluorescence intensity of glycated BSA. Furthermore, we demonstrated the significant effect of C. rotundus extract on preventing oxidative protein damages including effect on PCO formation and thiol oxidation which are believed to form under the glycoxidation process. Our results highlight the protein glycation inhibitory and antioxidant activity of C. rotundus. These results might lead to the possibility of using the plant extract or its purified active components for targeting diabetic complications. PMID:17765965

  12. Identification and isolation of a platelet GPIb-like protein in human umbilical vein endothelial cells and bovine aortic smooth muscle cells.

    PubMed Central

    Asch, A S; Adelman, B; Fujimoto, M; Nachman, R L

    1988-01-01

    Glycoprotein Ib (GPIb) is an intrinsic platelet membrane protein that plays a major role in platelet adherence and mediates ristocetin-dependent platelet von Willebrand factor binding. Recent reports that the platelet membrane glycoprotein complex IIb/IIIa is expressed in several cell types prompted us to look for GPIb expression in other vascular cells. Immunoperoxidase staining of human stomach and skin histologic sections with polyclonal as well as monoclonal anti-GPIb antibody revealed the presence of GPIb in the endothelial cell and smooth muscle cell layers. Western blotting using monospecific polyclonal anti-GPIb antibodies confirmed the presence of immunoreactive GPIb in human umbilical vein endothelial and bovine aortic smooth muscle cell cultures. Fab fragments of a monoclonal anti-GPIb antibody were used to immunoprecipitate [3H]leucine labeled GPIb from metabolically labeled cells. The GPIb in these cells was functional as measured by ristocetin-dependent cell agglutination and by vWF binding. Endothelial cells as well as smooth muscle cells bound 125I-labeled vWF in a ristocetin-dependent manner, with a Kd of 7.9 nM. Images PMID:3284916

  13. Protection against bovine tuberculosis induced by oral vaccination of cattle with Mycobacterium bovis BCG is not enhanced by co-administration of mycobacterial protein vaccines.

    PubMed

    Wedlock, D Neil; Aldwell, Frank E; Vordermeier, H Martin; Hewinson, R Glyn; Buddle, Bryce M

    2011-12-15

    Mycobacterium bovis bacille Calmette-Guérin (BCG) delivered to calves by the oral route in a formulated lipid matrix has been previously shown to induce protection against bovine tuberculosis. A study was conducted in cattle to determine if a combination of a low dose of oral BCG and a protein vaccine could induce protective immunity to tuberculosis while not sensitising animals to tuberculin. Groups of calves (10 per group) were vaccinated by administering 2 × 10(7)colony forming units (CFU) of BCG orally or a combination of 2 × 10(7)CFU oral BCG and a protein vaccine comprised of M. bovis culture filtrate proteins (CFP) formulated with the adjuvants Chitin and Gel 01 and delivered by the intranasal route, or CFP formulated with Emulsigen and the TLR2 agonist Pam(3)CSK(4) and administered by the subcutaneous (s.c.) route. Two further groups were vaccinated with the CFP/Chitin/Gel 01 or CFP/Emulsigen/Pam(3)CSK(4) vaccines alone. Positive control groups were given 10(8)CFU oral BCG or 10(6)CFU s.c. BCG while a negative control group was non-vaccinated. All animals were challenged with M. bovis 15 weeks after vaccination and euthanized and necropsied at 16 weeks following challenge. Groups of cattle vaccinated with s.c. BCG, 10(8)CFU or 2 × 10(7)CFU oral BCG showed significant reductions in seven, three and four pathological or microbiological disease parameters, respectively, compared to the results for the non-vaccinated group. There was no evidence of protection in calves vaccinated with the combination of oral BCG and CFP/Emulsigen/Pam(3)CSK(4) or oral BCG and CFP/Chitin/Gel 01 or vaccinated with the protein vaccines alone. Positive responses in the comparative cervical skin test at 12 weeks after vaccination were only observed in animals vaccinated with s.c. BCG, 10(8)CFU oral BCG or a combination of 2 × 10(7)CFU oral BCG and CFP/Chitin/Gel 01. In conclusion, co-administration of a protein vaccine, administered by either systemic or mucosal routes with oral BCG did not enhance the protection conferred by administration of oral BCG alone. PMID:22005585

  14. Screening of Mycobacterium avium subsp. paratuberculosis mutants for attenuation in a bovine monocyte-derived macrophage model.

    PubMed

    Lamont, Elise A; Talaat, Adel M; Coussens, Paul M; Bannantine, John P; Grohn, Yrjo T; Katani, Robab; Li, Ling-ling; Kapur, Vivek; Sreevatsan, Srinand

    2014-01-01

    Vaccination remains a major tool for prevention and progression of Johne's disease, a chronic enteritis of ruminants worldwide. Currently there is only one licensed vaccine within the United States and two vaccines licensed internationally against Johne's disease. All licensed vaccines reduce fecal shedding of Mycobacterium avium subsp. paratuberculosis (MAP) and delay disease progression. However, there are no available vaccines that prevent disease onset. A joint effort by the Johne's Disease Integrated Program (JDIP), a USDA-funded consortium, and USDA-APHIS/VS sought to identify transposon insertion mutant strains as vaccine candidates in part of a three phase study. The focus of the Phase I study was to evaluate MAP mutant attenuation in a well-defined in vitro bovine monocyte-derived macrophage (MDM) model. Attenuation was determined by colony forming unit (CFUs) counts and slope estimates. Based on CFU counts alone, the MDM model did not identify any mutant that significantly differed from the wild-type control, MAP K-10. Slope estimates using mixed models approach identified six mutants as being attenuated. These were enrolled in protection studies involving murine and baby goat vaccination-challenge models. MDM based approach identified trends in attenuation but this did not correlate with protection in a natural host model. These results suggest the need for alternative strategies for Johne's disease vaccine candidate screening and evaluation. PMID:25072030

  15. Screening of Mycobacterium avium subsp. paratuberculosis mutants for attenuation in a bovine monocyte-derived macrophage model

    PubMed Central

    Lamont, Elise A.; Talaat, Adel M.; Coussens, Paul M.; Bannantine, John P.; Grohn, Yrjo T.; Katani, Robab; Li, Ling-ling; Kapur, Vivek; Sreevatsan, Srinand

    2014-01-01

    Vaccination remains a major tool for prevention and progression of Johne's disease, a chronic enteritis of ruminants worldwide. Currently there is only one licensed vaccine within the United States and two vaccines licensed internationally against Johne's disease. All licensed vaccines reduce fecal shedding of Mycobacterium avium subsp. paratuberculosis (MAP) and delay disease progression. However, there are no available vaccines that prevent disease onset. A joint effort by the Johne's Disease Integrated Program (JDIP), a USDA-funded consortium, and USDA—APHIS/VS sought to identify transposon insertion mutant strains as vaccine candidates in part of a three phase study. The focus of the Phase I study was to evaluate MAP mutant attenuation in a well-defined in vitro bovine monocyte-derived macrophage (MDM) model. Attenuation was determined by colony forming unit (CFUs) counts and slope estimates. Based on CFU counts alone, the MDM model did not identify any mutant that significantly differed from the wild-type control, MAP K-10. Slope estimates using mixed models approach identified six mutants as being attenuated. These were enrolled in protection studies involving murine and baby goat vaccination-challenge models. MDM based approach identified trends in attenuation but this did not correlate with protection in a natural host model. These results suggest the need for alternative strategies for Johne's disease vaccine candidate screening and evaluation. PMID:25072030

  16. Comparative spectroscopic studies on drug binding characteristics and protein surface hydrophobicity of native and modified forms of bovine serum albumin: Possible relevance to change in protein structure/function upon non-enzymatic glycation

    NASA Astrophysics Data System (ADS)

    Khodarahmi, Reza; Karimi, Seyyed Arash; Ashrafi Kooshk, Mohammad Reza; Ghadami, Seyyed Abolghasem; Ghobadi, Sirous; Amani, Mojtaba

    2012-04-01

    The interaction between serum albumin (SA) and drugs has provided an interesting ground for understanding of drug effects, especially in drug distribution and drug-drug interaction on SA, in the case of multi-drug therapy. Determination of the impact of various factors on drug-protein interaction is especially important upon significant binding of drug to albumin. In the present study, the interaction of two drugs (furosemide and indomethacin) with native and modified albumins were investigated by using various spectroscopic methods. Fluorescence data indicated that 1:1 binding of drugs to bovine serum albumin (BSA) is associated with quenching of albumin intrinsic fluorescence. The Job's plot also confirmed that drug binds to BSA via mentioned stoichiometry. Analysis of the quenching and thermodynamic parameters indicated that intermolecular interactions between drug and albumin may change upon protein modification. The theoretical analyses also suggested some conformational changes of interacting side chains in subdomain IIA binding site (at the vicinity of W237), which were in good agreement with experimental data. Decrease of protein surface hydrophobicity (PSH) was also observed upon both albumin modification and drug binding.

  17. Modelling the spread of bovine viral diarrhea virus (BVDV) in a beef cattle herd and its impact on herd productivity.

    PubMed

    Damman, Alix; Viet, Anne-France; Arnoux, Sandie; Guerrier-Chatellet, Marie-Claude; Petit, Etienne; Ezanno, Pauline

    2015-01-01

    Bovine viral diarrhea virus (BVDV) is a common pathogen of cattle herds that causes economic losses due to reproductive disorders in breeding cattle and increased morbidity and mortality amongst infected calves. Our objective was to evaluate the impact of BVDV spread on the productivity of a beef cow-calf herd using a stochastic model in discrete time that accounted for (1) the difference in transmission rates when animals are housed indoors versus grazing on pasture, (2) the external risk of disease introductions through fenceline contact with neighboring herds and the purchase of infected cattle, and (3) the risk of individual pregnant cattle generating persistently infected (PI) calves based on their stage in gestation. The model predicted the highest losses from BVDV during the first 3 years after disease was introduced into a naive herd. During the endemic phase, the impact of BVDV on the yearly herd productivity was much lower due to herd immunity. However, cumulative losses over 10 years in an endemic situation greatly surpassed the losses that occurred during the acute phase. A sensitivity analysis of key model parameters revealed that herd size, the duration of breeding, grazing, and selling periods, renewal rate of breeding females, and the level of numerical productivity expected by the farmer had a significant influence on the predicted losses. This model provides a valuable framework for evaluating the impact of BVDV and the efficacy of different control strategies in beef cow-calf herds. PMID:25828555

  18. Crystal structure of the cytochrome bc⁠complex from bovine heart mitochondria

    Microsoft Academic Search

    Di Xia; Hoeon Kim; J. Deisenhofer; Li Zhang

    1997-01-01

    On the basis of x-ray diffraction data to a resolution of 2.9 angstroms, atomic models of most protein components of the bovine cytochrome bc⁠complex were built, including core 1, core 2, cytochrome b, subunit 6, and subunit 7, a carboxyl-terminal fragment of cytochrome câ, and an amino-terminal fragment of the iron-sulfur protein. The positions of the four iron centers

  19. REMOVAL OF DECORIN CORE-PROTEIN FROM POWDERED BOVINE HIDES BY TREATMENTS USED TO PROCESS INTACT HIDES INTO LEATHER

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using a modification of a previously developed sandwich ELISA procedure, we determined the amount of decorin core-protein (DCP) content of raw powdered hide and powdered hide treated with the early steps of the tanning process. We found approximately 0.15 mg DCP/g hide in raw powdered hide. Treatm...

  20. Relationship between single nucleotide polymorphisms in the bovine heat shock protein 70 gene and milk characteristics of beef cows

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Heat shock proteins (HSPs) are induced by various stressors such as heat, cold, toxins, and oxygen deprivation. Our objective was to determine the genetic diversity in a segment of the HSP-70 gene of cattle. Genomic DNA was collected from 157 cows. The cows were Angus (n = 42), Brahman (n = 41), and...

  1. Divergent antibody isotype responses induced in mice by systemic exposure to proteins: a comparison of ovalbumin with bovine serum albumin

    Microsoft Academic Search

    R. J Dearman; H Caddick; D. A Basketter; I Kimber

    2000-01-01

    Whereas many foreign proteins are immunogenic, only a proportion is associated commonly with allergy, having the potential to induce the quality of immune response necessary for IgE antibody production and the development of immediate type hypersensitivity reactions in the gastrointestinal and\\/or respiratory tracts. In the context of toxicological evaluations there is a need to identify those properties that confer on

  2. THE MYCOBACTERIUM AVIUM SUPSPECIES PARATUBERCULOSIS 35 KDA PROTEIN PLAYS A ROLE IN INVASION OF BOVINE EPITHELIAL CELLS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) enters intestinal epithelial cells of cattle and other ruminants via a mechanism that remains to be fully elucidated. In this study, we observed that a gene encoding the M. paratuberculosis 35-kDa major membrane protein (MMP) is ...

  3. The Use of Bovine Serum Protein as an Oral Support Therapy Following Coronavirus Challenge in Calves1

    Microsoft Academic Search

    J. D. Arthington; C. A. Jaynes; H. D. Tyler; S. Kapil; J. D. Quigley III

    2002-01-01

    The objective of this experiment was to investigate thetherapeuticefficacyofasupplementalbovineserum protein blend fed to calves challenged with virulent coronavirus. Twelve Holstein bull calves (approxi- mately3wkofage)wereallocatedbyinitialbodyweight to Control (n = 5) and treated (n = 7) groups. On d 0, all calves were orally challenged with 1 × 107 plaque- forming units of virulent coronavirus isolate. Infection was allowed to progress for

  4. Learning generative models for protein fold families.

    PubMed

    Balakrishnan, Sivaraman; Kamisetty, Hetunandan; Carbonell, Jaime G; Lee, Su-In; Langmead, Christopher James

    2011-04-01

    We introduce a new approach to learning statistical models from multiple sequence alignments (MSA) of proteins. Our method, called GREMLIN (Generative REgularized ModeLs of proteINs), learns an undirected probabilistic graphical model of the amino acid composition within the MSA. The resulting model encodes both the position-specific conservation statistics and the correlated mutation statistics between sequential and long-range pairs of residues. Existing techniques for learning graphical models from MSA either make strong, and often inappropriate assumptions about the conditional independencies within the MSA (e.g., Hidden Markov Models), or else use suboptimal algorithms to learn the parameters of the model. In contrast, GREMLIN makes no a priori assumptions about the conditional independencies within the MSA. We formulate and solve a convex optimization problem, thus guaranteeing that we find a globally optimal model at convergence. The resulting model is also generative, allowing for the design of new protein sequences that have the same statistical properties as those in the MSA. We perform a detailed analysis of covariation statistics on the extensively studied WW and PDZ domains and show that our method out-performs an existing algorithm for learning undirected probabilistic graphical models from MSA. We then apply our approach to 71 additional families from the PFAM database and demonstrate that the resulting models significantly out-perform Hidden Markov Models in terms of predictive accuracy. PMID:21268112

  5. Non-classical mechanisms of steroid sensing in the ovary: lessons from the bovine oxytocin model.

    PubMed

    Ivell, Richard; Dai, Yanzhenzi; Mann, Navdeep; Anand-Ivell, Ravinder

    2014-01-25

    Steroidogenic tissues such as the ovary, testes or adrenal glands are paradoxical in that they often indicate actions of steroid hormones within a dynamic range of ligand concentration in a high nanomolar or even micromolar level, i.e. at the natural concentrations existing within those organs. Yet ligand-activated nuclear steroid receptors act classically by direct interaction with DNA in the picomolar or low nanomolar range. Moreover, global genomic studies suggest that less than 40% of steroid-regulated genes involve classical responsive elements in gene promoter regions. The bovine oxytocin gene is a key element in the maternal recognition of pregnancy in ruminants and is regulated via an SF1 site in its proximal promoter. This gene is also regulated by steroids acting in a non-classical manner, involving nuclear receptors which do not interact directly with DNA. Dose-response relationships for these actions are in the high nanomolar range. Similar 'steroid sensing' mechanisms may prevail for other SF1-regulated genes and predict alternative pathways by which environmental endocrine disruptors might influence the functioning of steroid-producing organs and hence indirectly the steroid-dependent control of physiology and development. PMID:23632104

  6. Rigidity Analysis for Modeling Protein Motion 

    E-print Network

    Thomas, Shawna L.

    2010-07-14

    of freedom computed by rigidity analysis, and (f) RMSD to both target states. ........................... 60 19 Example turn protein fragment (FANGST) where hydrogen bonds are indicated with dashed lines and hydrophobic interactions with dotted lines... helical protein fragement (E-V-L-A-Y-A-D-T) where hy- drogen bonds are indicated with dashed lines and hydrophobic interactions with dotted lines. ...................... 70 23 Single-body, C? model example of the helical segment in Fig. 22. .. 71 24 Two...

  7. Generation of a Persistently Infected MDBK Cell Line with Natural Bovine Spongiform Encephalopathy (BSE)

    PubMed Central

    Tark, Dongseob; Kim, Hyojin; Neale, Michael H.; Kim, Minjeong; Sohn, Hyunjoo; Lee, Yoonhee; Cho, Insoo; Joo, Yiseok; Windl, Otto

    2015-01-01

    Bovine spongiform encephalopathy (BSE) is a zoonotic transmissible spongiform encephalopathy (TSE) thought to be caused by the same prion strain as variant Creutzfeldt-Jakob disease (vCJD). Unlike scrapie and chronic wasting disease there is no cell culture model allowing the replication of proteinase K resistant BSE (PrPBSE) and the further in vitro study of this disease. We have generated a cell line based on the Madin-Darby Bovine Kidney (MDBK) cell line over-expressing the bovine prion protein. After exposure to naturally BSE-infected bovine brain homogenate this cell line has shown to replicate and accumulate PrPBSE and maintain infection up to passage 83 after initial challenge. Collectively, we demonstrate, for the first time, that the BSE agent can infect cell lines over-expressing the bovine prion protein similar to other prion diseases. These BSE infected cells will provide a useful tool to facilitate the study of potential therapeutic agents and the diagnosis of BSE. PMID:25647616

  8. A soft and transparent handleable protein model.

    PubMed

    Kawakami, Masaru

    2012-08-01

    The field of structural biology currently relies on computer-generated graphical representations of three-dimensional (3D) structures to conceptualize biomolecules. As the size and complexity of the molecular structure increases, model generation and peer discussions become more difficult. It is even more problematic when discussing protein-protein interactions wherein large surface area contact is considered. This report demonstrates the viability of a new handleable protein molecular model with a soft and transparent silicone body similar to the molecule's surface. A full-color printed main chain structure embedded in the silicone body enables users to simultaneously feel the molecular surface, view through the main chain structure, and manually simulate molecular docking. The interactive, hands-on experience deepens the user's intuitive understanding of the complicated 3D protein structure and elucidates ligand binding and protein-protein interactions. This model would be an effective discussion tool for the classroom or laboratory that stimulates inspired learning in this study field. PMID:22938316

  9. Diagnosis of tropical bovine theileriosis by ELISA with recombinant merozoite surface protein of Theileria annulata (Tams1).

    PubMed

    Rajendran, C; Ray, D D

    2014-03-01

    Theileria annulata merozoite surface protein (Tams1) and native soluble piroplasm antigen of an Indian isolate of T. annulata were used to optimize an enzyme linked immunosorbent assay (ELISA) to detect anti-Theileria antibodies in cross-bred cattle experimentally infected with sporozoites or immunized with macroschizonts of the homologous strain of the parasite. The recombinant protein has showed similar antibody titre when compared with the sonicated merozoite/piroplasm antigen in an ELISA. IFAT was used as a reference test. In 56 sera of apparently healthy cattle collected from endemic area, ELISA showed positive antibody response in 29 samples with Tams1 whereas the soluble piroplasm antigen reacted positively with 34 samples and also with sera of cattle positive for Babesia bigemina. Microscopical detection of piroplasms was possible in the blood smears of 17 animals which were serologically positive in both native and recombinant antigens. PMID:24505176

  10. The effect of myofibrillar protein interaction on the tenderness of bovine muscle subjected to cold-shortening and postmortem conditioning 

    E-print Network

    Pollard, Michele Andrea

    1983-01-01

    have been due to the covering up of gap filaments by myosin tails in the contracted sarcomeres (Locker, 1977). ACKNOWLEDGMENTS The author wishes to express her sincere apprecia- tion to Dr. T. R. Dutson, Dr. G. C. Smith and Dr. F. A. Gardner... suggest that the gap filaments, a myofibrillar protein component which spans the gap between myosin filaments in adjacent sarcomeres, play a dominant role in determining the tensile strength and therefore the toughness of the myofibril in both raw...

  11. Protein modeling using hidden Markov models: analysis of globins

    Microsoft Academic Search

    D. Haussler; A. Krogh; I. Saira Mian; Kimmen Sjolander

    1993-01-01

    The authors apply hidden Markov models to the problem of statistical modeling and multiple sequence alignment of protein families. A variant of the expectation maximization algorithm known as the Viterbi algorithm is used to obtain the statistical model from the unaligned sequences. In a detailed series of experiments, they have taken 400 unaligned globin sequences, and produced a statistical model

  12. Color modeling of protein optical probes.

    PubMed

    Murugan, N Arul; Kongsted, Jacob; Rinkevicius, Zilvinas; Ågren, Hans

    2012-01-21

    We present a strategy for modeling optical probes within heterogeneous environments of restricted dimension. The method is based on a multiphysics approach comprising sequential structure modeling by means of hybrid Car-Parrinello molecular dynamics and property modeling by means of quantum mechanics/molecular mechanics response theory. For demonstration we address the structural and optical properties of nile red within the ?-lacto globulin protein. We consider the cases with the probe situated on the surface or within the cavity of the protein, or embedded in a water solvent. We find the absorption properties of the probe to be highly dependent on its position relative to the protein. Structural rearrangements of the optical probe are found to contribute significantly to these environmental effects. PMID:22134524

  13. Primary cell cultures from fetal bovine hypothalamus and cerebral cortex: A reliable model to study P450 Arom and ? and ? estrogen receptors in vitro

    Microsoft Academic Search

    Antonella Peruffo; Genny Buson; Bruno Cozzi; Cristina Ballarin

    2008-01-01

    Estrogens synthesized by neural P450 aromatase (P450Arom) are implicated in many aspects of mammalian brain development and particularly in sexual differentiation of the central nervous system (CNS). This study analyzes the usefulness of an in vitro model based on bovine primary cell cultures from the hypothalamus and frontal cortex to investigate the role of P450Arom and estrogen receptors (ERs) in

  14. Protein Structure Modeling With MODELLER Narayanan Eswar$

    E-print Network

    Sali, Andrej

    sequences. Key Words: Comparative modeling, fold assignment, sequence-structure alignment, model assessment resonance (NMR) spectroscopy. In the absence of experimentally determined structures, computationally template structure (see Section on Materials for definitions of these terms); (ii) alignment of the target

  15. Higher-order structure of bovine mitochondrial tRNA(SerUGA): chemical modification and computer modeling.

    PubMed Central

    Watanabe, Y; Kawai, G; Yokogawa, T; Hayashi, N; Kumazawa, Y; Ueda, T; Nishikawa, K; Hirao, I; Miura, K; Watanabe, K

    1994-01-01

    On the basis of enzymatic probing and phylogenetic comparison, we have previously proposed that mammalian mitochondrial tRNA(sSer) (anticodon UGA) possess a slightly altered cloverleaf structure in which only one nucleotide exists between the acceptor stem and D stem (usually two nucleotides) and the anticodon stem consists of six base pairs (usually five base pairs) [Yokogawa et al. (1991) Nucleic Acids Res. 19, 6101-6105]. To ascertain whether such tRNA(sSer) can be folded into a normal L-shaped tertiary structure, the higher-order structure of bovine mitochondrial tRNA(SerUGA) was examined by chemical probing using dimethylsulfate and diethylpyrocarbonate, and on the basis of the results a tertiary structure model was obtained by computer modeling. It was found that a one-base-pair elongation in the anticodon stem was compensated for by multiple-base deletions in the D and extra loop regions of the tRNA(SerUGA), which resulted in preservation of an L-shaped tertiary structure similar to that of conventional tRNAs. By summarizing the findings, the general structural requirements of mitochondrial tRNAs necessary for their functioning in the mitochondrial translation system are considered. Images PMID:7529407

  16. A Three-dimensional Cell Culture Model for Bovine Endometrium: Regeneration of a Multicellular Spheroid Using Ascorbate

    Microsoft Academic Search

    N Yamauchi; O Yamada; T Takahashi; K Imai; T Sato; A Ito; K Hashizume

    2003-01-01

    The development of a multicellular spheroid comprising bovine endometrial epithelial cells (BEE) and bovine endometrial stromal cells (BES) is described in this study. The BES were cultured to confluence in medium with l -ascorbic acid phosphate magnesium salt n -hydrate (AsA-P) which stimulates collagen synthesis in BES. The BEE were co-cultured on a BES cell-sheet for 24h before detachment of

  17. Fast loop modeling for protein structures

    NASA Astrophysics Data System (ADS)

    Zhang, Jiong; Nguyen, Son; Shang, Yi; Xu, Dong; Kosztin, Ioan

    2015-03-01

    X-ray crystallography is the main method for determining 3D protein structures. In many cases, however, flexible loop regions of proteins cannot be resolved by this approach. This leads to incomplete structures in the protein data bank, preventing further computational study and analysis of these proteins. For instance, all-atom molecular dynamics (MD) simulation studies of structure-function relationship require complete protein structures. To address this shortcoming, we have developed and implemented an efficient computational method for building missing protein loops. The method is database driven and uses deep learning and multi-dimensional scaling algorithms. We have implemented the method as a simple stand-alone program, which can also be used as a plugin in existing molecular modeling software, e.g., VMD. The quality and stability of the generated structures are assessed and tested via energy scoring functions and by equilibrium MD simulations. The proposed method can also be used in template-based protein structure prediction. Work supported by the National Institutes of Health [R01 GM100701]. Computer time was provided by the University of Missouri Bioinformatics Consortium.

  18. Modelling of DNA-protein recognition

    NASA Technical Reports Server (NTRS)

    Rein, R.; Garduno, R.; Colombano, S.; Nir, S.; Haydock, K.; Macelroy, R. D.

    1980-01-01

    Computer model-building procedures using stereochemical principles together with theoretical energy calculations appear to be, at this stage, the most promising route toward the elucidation of DNA-protein binding schemes and recognition principles. A review of models and bonding principles is conducted and approaches to modeling are considered, taking into account possible di-hydrogen-bonding schemes between a peptide and a base (or a base pair) of a double-stranded nucleic acid in the major groove, aspects of computer graphic modeling, and a search for isogeometric helices. The energetics of recognition complexes is discussed and several models for peptide DNA recognition are presented.

  19. Enrofloxacin and Macrolides Alone or in Combination with Rifampicin as Antimicrobial Treatment in a Bovine Model of Acute Chlamydia psittaci Infection

    PubMed Central

    Prohl, Annette; Lohr, Markus; Ostermann, Carola; Liebler-Tenorio, Elisabeth; Berndt, Angela; Schroedl, Wieland; Rothe, Michael; Schubert, Evelyn; Sachse, Konrad; Reinhold, Petra

    2015-01-01

    Chlamydia psittaci is a zoonotic bacterium with a wide host range that can cause respiratory disease in humans and cattle. In the present study, effects of treatment with macrolides and quinolones applied alone or in combination with rifampicin were tested in a previously established bovine model of respiratory C. psittaci infection. Fifty animals were inoculated intrabronchially at the age of 6–8 weeks. Seven served as untreated controls, the others were assigned to seven treatment groups: (i) rifampicin, (ii) enrofloxacin, (iii) enrofloxacin + rifampicin, (iv) azithromycin, (v) azithromycin + rifampicin, (vi) erythromycin, and (vii) erythromycin + rifampicin. Treatment started 30 hours after inoculation and continued until 14 days after inoculation (dpi), when all animals were necropsied. The infection was successful in all animals and sufficient antibiotic levels were detected in blood plasma and tissue of the treated animals. Reisolation of the pathogen was achieved more often from untreated animals than from other groups. Nevertheless, pathogen detection by PCR was possible to the same extent in all animals and there were no significant differences between treated and untreated animals in terms of local (i.e. cell count and differentiation of BALF-cells) and systemic inflammation (i.e. white blood cells and concentration of acute phase protein LBP), clinical signs, and pathological findings at necropsy. Regardless of the reduced reisolation rate in treated animals, the treatment of experimentally induced respiratory C. psittaci infection with enrofloxacin, azithromycin or erythromycin alone or in combination with rifampicin was without obvious benefit for the host, since no significant differences in clinical and pathological findings or inflammatory parameters were detected and all animals recovered clinically within two weeks. PMID:25768665

  20. Lysozyme: a model protein for amyloid research.

    PubMed

    Swaminathan, Rajaram; Ravi, Vijay Kumar; Kumar, Satish; Kumar, Mattaparthi Venkata Satish; Chandra, Nividh

    2011-01-01

    Ever since lysozyme was discovered by Fleming in 1922, this protein has emerged as a model for investigations on protein structure and function. Over the years, several high-resolution structures have yielded a wealth of structural data on this protein. Extensive studies on folding of lysozyme have shown how different regions of this protein dynamically interact with one another. Data is also available from numerous biotechnological studies wherein lysozyme has been employed as a model protein for recovering active recombinant protein from inclusion bodies using small molecules like l-arginine. A variety of conditions have been developed in vitro to induce fibrillation in hen lysozyme. They include (a) acidic pH at elevated temperature, (b) concentrated solutions of ethanol, (c) moderate concentrations of guanidinium hydrochloride at moderate temperature, and (d) alkaline pH at room temperature. This review aims to bring together similarities and differences in aggregation mechanisms, morphology of aggregates, and related issues that arise using the different conditions mentioned above to improve our understanding. The alkaline pH condition (pH 12.2), discovered and studied extensively in our lab, shall receive special attention. More than a decade ago, it was revealed that mutations in human lysozyme can cause accumulation of large quantities of amyloid in liver, kidney, and other regions of gastrointestinal tract. Understanding the mechanism of lysozyme aggregation will probably have therapeutic implications for the treatment of systemic nonneuropathic amyloidosis. Numerous studies have begun to focus attention on inhibition of lysozyme aggregation using antibody or small molecules. The enzymatic activity of lysozyme presents a convenient handle to quantify the native population of lysozyme in a sample where aggregation has been inhibited. The rich information available on lysozyme coupled with the multiple conditions that have been successful in inducing/inhibiting its aggregation in vitro makes lysozyme an ideal model protein to investigate amyloidogenesis. PMID:21846563

  1. Estradiol and Progesterone Exhibit Similar Patterns of Hepatic Gene Expression Regulation in the Bovine Model

    PubMed Central

    Piccinato, Carla A.; Rosa, Guilherme J. M.; N’Jai, Alhaji U.; Jefcoate, Colin R.; Wiltbank, Milo C.

    2013-01-01

    Female sex steroid hormones, estradiol-17? (E2-17?) and progesterone (P4) regulate reproductive function and gene expression in a broad range of tissues. Given the central role of the liver in regulating homeostasis including steroid hormone metabolism, we sought to understand how E2-17? and P4 interact to affect global gene expression in liver. Ovariectomized cows (n?=?8) were randomly assigned to 4 treatment groups applied in a replicated Latin Square design: 1) No hormone supplementation, 2) E2-17? treatment (ear implant), 3) P4 treatment (intravaginal inserts), and 4) E2-17? combined with P4. After 14 d of treatment, liver biopsies were collected, allowing 28 d intervals between periods. Changes in gene expression in the liver biopsies were monitored using bovine-specific arrays. Treatment with E2-17? altered expression of 479 genes, P4 472 genes, and combined treatment significantly altered expression of 468 genes. In total, 578 genes exhibited altered expression including a remarkable number (346 genes) that responded similarly to E2-17?, P4, or combined treatment. Additional evidence for similar gene expression actions of E2-17ß and/or P4 were: principal component analysis placed almost every treatment array at a substantial distance from controls; Venn diagrams indicated overall treatment effects for most regulated genes; clustering analysis indicated the two major clusters had all treatments up-regulating (172 genes) or down-regulating (173 genes) expression. Thus, unexpectedly, common biological pathways were regulated by E2-17? and/or P4 in liver. This indicates that the mechanism of action of these steroid hormones in the liver might be either indirect or might occur through non-genomic pathways. This unusual pattern of gene expression in response to steroid hormones is consistent with the idea that there are classical and non-classical tissue-specific responses to steroid hormone actions. Future studies are needed to elucidate putative mechanism(s) responsible for overlapping actions of E2-17? and P4 on the liver transcriptome. PMID:24069207

  2. A Three-Pore Model Describes Transport Properties of Bovine Retinal Endothelial Cells in Normal and Elevated Glucose

    PubMed Central

    Lopez-Quintero, Sandra V.; Ji, Xin-Ying; Antonetti, David A.

    2011-01-01

    Purpose. Changes in blood vessel barrier properties contribute to retinal edema in diabetic retinopathy (DR). However, limited data are available to describe the routes of transport for fluids and solutes across the inner blood–retinal barrier (iBRB). In this study, a three-pore model was developed to characterize such routes in normal and elevated glucose levels. Methods. Diffusive and apparent permeabilities to TAMRA (467 Da), dextran (70 kDa), and LDL (2000 kDa), as well as hydraulic conductivity, were measured across bovine retinal endothelial cell (BREC) monolayers after exposure to normal- and high-glucose media for 6 days. The data were used to develop a model of transport dynamics. Claudin 5 and eNOS Western blot analysis were used to measure changes in expression and phosphorylation. Immunolocalization of ZO-1 and VE-cadherin demonstrated organization of the junctional complex. Apoptosis was measured by TUNEL assay. Results. A three-pore model describes the fractional transport of water and molecular tracers across the retinal endothelial barrier. No change in permeability or hydraulic conductivity was observed after exposure to high glucose, whereas VEGF increased permeability in both normal- and high-glucose environments. The transport results were consistent with ZO-1 and VE-cadherin immunocytochemistry and expression of claudin-5, which were all unaltered by high glucose. Conclusions. The data describe, for the first time, a model for transport of various size solutes and fluids across endothelial cells of the iBRB. Further, the results support the existence of an indirect pathway by which iBRB permeability is increased through the upregulation of retinal VEGF in response to hyperglycemia. PMID:21357410

  3. Molecular modeling and spectroscopic studies on the interaction of the chiral drug venlafaxine hydrochloride with bovine serum albumin.

    PubMed

    Shahabadi, Nahid; Hadidi, Saba

    2014-03-25

    This study was designed to examine the interaction of racemic antidepressant drug "S,R-venlafaxine hydrochloride (VEN)" with bovine serum albumin (BSA) under physiological conditions. The mechanism of interaction was studied by spectroscopic techniques combination with molecular modeling. Stern-Volmer analysis of fluorescence quenching data shows the presence of the static quenching mechanism. The thermodynamic parameters indicated that the hydrogen bonding and weak van der Waals interactions are the predominant intermolecular forces stabilizing the complex. The number of binding sites (n) was calculated. Through the site marker competitive experiment, VEN was confirmed to be located in subdomain IIIA of BSA. The binding distance (r=4.93 nm) between the donor BSA and acceptor VEN was obtained according to Förster's non-radiative energy transfer theory. According to UV-vis spectra and CD data binding of VEN leaded to conformational changes of BSA. Molecular docking simulations of S and R-VEN revealed that both isomers have similar interaction and the same binding sites, from this point of view S and R isomers are equal. PMID:24299981

  4. Evaluation of processed bovine cancellous bone matrix seeded with syngenic osteoblasts in a critical size calvarial defect rat model

    PubMed Central

    Kneser, U; Stangenberg, L; Ohnolz, J; Buettner, O; Stern-Straeter, J; Möbest, D; Horch, R E; Stark, G B; Schaefer, D J

    2006-01-01

    Introduction: Biologic bone substitutes may offer alternatives to bone grafting procedures. The aim of this study was to evaluate a preformed bone substitute based on processed bovine cancellous bone (PBCB) with or without osteogenic cells in a critical size calvarial defect rat model. Methods: Discs of PBCB (Tutobone®) were seeded with second passage fibrin gel-immobilized syngenic osteoblasts (group A, n = 40). Cell-free matrices (group B, n = 28) and untreated defects (group C; n=28) served as controls. Specimens were explanted between day 0 and 4 months after implantation and were subjected to histological and morphometric evaluation. Results: At 1 month, bone formation was limited to small peripheral areas. At 2 and 4 months, significant bone formation, matrix resorption as well as integration of the implants was evident in groups A and B. In group C no significant regeneration of the defects was observed. Morphometric analysis did not disclose differences in bone formation in matrices from groups A and B. Carboxyfluorescine-Diacetate-Succinimidylester (CFDA) labeling demonstrated low survival rates of transplanted cells. Discussion: Osteoblasts seeded into PBCB matrix display a differentiated phenotype following a 14 days cell culture period. Lack of initial vascularization may explain the absence of added osteogenicity in constructs from group A in comparison to group B. PBCB is well integrated and represents even without osteogenic cells a promising biomaterial for reconstruction of critical size calvarial bone defects. PMID:16989729

  5. X-ray fluorescence-based differentiation of neck tissues in a bovine model: implications for potential intraoperative use.

    PubMed

    Lahav, G; Shilstein, S; Shchemelinin, S; Ikher, S; Halperin, D; Chechik, R; Breskin, A

    2015-05-01

    This study explores the possibility of using X-ray fluorescence (XRF)-based trace-element analysis for differentiation of various bovine neck tissues. It is motivated by the requirement for an intra-operative in-vivo method for identifying parathyroid glands, particularly beneficial in surgery in the central neck-compartment. Using a dedicated X-ray spectral analysis, we examined ex-vivo XRF spectra from various histologically verified fresh neck tissues from cow, which was chosen as the animal model; these tissues included fat, muscle, thyroid, parathyroid, lymph nodes, thymus and salivary gland. The data for six trace elements K, Fe, Zn, Br, Rb and I, provided the basis for tissue identification by using multi-parameter analysis of the recorded XRF spectra. It is shown that the combination of XRF signals from these elements is sufficient for a reliable tissue differentiation. The average total abundance of these trace elements was evaluated in each tissue type, including parathyroid and salivary gland for the first time. It is shown that some tissues can unequivocally be identified on the basis of the abundance of a single element, for example, iodine and zinc for the identification of thyroid gland and muscle, respectively. PMID:25677045

  6. Animal models for protein respiratory sensitizers

    Microsoft Academic Search

    Marsha D. W. Ward; MaryJane K. Selgrade

    2007-01-01

    Protein induced respiratory hypersensitivity, particularly atopic disease in general, and allergic asthma in particular, has increased dramatically over the last several decades in the US and other industrialized nations as a result of ill-defined changes in living conditions in modern western society. In addition, work-related asthma has become the most frequently diagnosed occupational respiratory illness. Animal models have demonstrated great

  7. Extracting Protein-Protein Interactions from MEDLINE using the Hidden Vector State model

    Microsoft Academic Search

    Deyu Zhou; Yulan He; Chee Keong Kwoh

    2008-01-01

    A major challenge in text mining for biomedicine is automatically extracting protein-protein interactions from the vast amount of biomedical literature. We have constructed an information extraction system based on the Hidden Vector State (HVS) model for protein-protein interactions. The HVS model can be trained using only lightly annotated data whilst simultaneously retaining sufficient ability to capture the hierarchical structure. When

  8. Fold assessment for comparative protein structure modeling.

    PubMed

    Melo, Francisco; Sali, Andrej

    2007-11-01

    Accurate and automated assessment of both geometrical errors and incompleteness of comparative protein structure models is necessary for an adequate use of the models. Here, we describe a composite score for discriminating between models with the correct and incorrect fold. To find an accurate composite score, we designed and applied a genetic algorithm method that searched for a most informative subset of 21 input model features as well as their optimized nonlinear transformation into the composite score. The 21 input features included various statistical potential scores, stereochemistry quality descriptors, sequence alignment scores, geometrical descriptors, and measures of protein packing. The optimized composite score was found to depend on (1) a statistical potential z-score for residue accessibilities and distances, (2) model compactness, and (3) percentage sequence identity of the alignment used to build the model. The accuracy of the composite score was compared with the accuracy of assessment by single and combined features as well as by other commonly used assessment methods. The testing set was representative of models produced by automated comparative modeling on a genomic scale. The composite score performed better than any other tested score in terms of the maximum correct classification rate (i.e., 3.3% false positives and 2.5% false negatives) as well as the sensitivity and specificity across the whole range of thresholds. The composite score was implemented in our program MODELLER-8 and was used to assess models in the MODBASE database that contains comparative models for domains in approximately 1.3 million protein sequences. PMID:17905832

  9. Differences in proteinase K resistance and neuronal deposition of abnormal prion proteins characterize bovine spongiform encephalopathy (BSE) and scrapie strains.

    PubMed Central

    Kuczius, T.; Groschup, M. H.

    1999-01-01

    Prion diseases are associated with the accumulation of an abnormal isoform of host-encoded prion protein (PrP(Sc)). A number of prion strains can be distinguished by "glycotyping" analysis of the respective deposited PrP(Sc) compound. In this study, the long-term proteinase K resistance, the molecular mass, and the localization of PrP(Sc) deposits derived from conventional and transgenic mice inoculated with 11 different BSE and scrapie strains or isolates were examined. Differences were found in the long-term proteinase K resistance (50 microg/ml at 37 degrees C) of PrP(Sc). For example, scrapie strain Chandler or PrP(Sc) derived from field BSE isolates were destroyed after 6 hr of exposure, whereas PrP(Sc) of strains 87V and ME7 and of the Hessen1 isolate were extremely resistant to proteolytic cleavage. Nonglycosylated, proteinase K-treated PrP(Sc) of BSE isolates and of scrapie strain 87V exhibited a 1-2 kD lower molecular mass than PrP(Sc) derived from all other scrapie strains and isolates. With the exception of strain 87V, PrP(Sc) was generally deposited in the cerebrum, cerebellum, and brain stem of different mouse lines at comparable levels. Long-term proteinase resistance, molecular mass, and the analysis of PrP(Sc) deposition therefore provide useful criteria in discriminating prion strains and isolates (e.g., BSE and 87V) that are otherwise indistinguishable by the PrP(Sc) "glycotyping" technique. Images Fig. 2 Fig. 4 Fig. 6 PMID:10415165

  10. ISOLATION AND PROPERTIES OF BOVINE COLOSTRAL TRYPSIN INHIBITOR

    E-print Network

    Paris-Sud XI, Université de

    COLOSTRUM DES BOVINS. ― L'inhibiteur de la trypsine du colostrum des bovins a été isolé par chromato colostrum (CTI) is a heat- and acid-resistant protein, which was originally isolated by Laskowski the dynamics of the secretion of CTI and those of colostral immunoglobulins. Materials and Methods 1. COLOSTRUM

  11. A study of intrusion characteristics of low viscosity cement Simplex-P and Palacos cements in a bovine cancellous bone model.

    PubMed

    Rey, R M; Paiement, G D; McGann, W M; Jasty, M; Harrigan, T P; Burke, D W; Harris, W H

    1987-02-01

    Aseptic loosening is the most common long-term complication of cemented total hip arthroplasties (THA). The functional longevity of these implants depends on the bone-cement interface. The influence of cement injection pressure, type of cement, ambient temperature, chilling of the monomer, and centrifugation of cement-on-cement intrusion depth was investigated in specimens of bovine cancellous bone. In order to validate the bovine model for comparative purposes relative to use in man, a linear relationship between human and bovine cancellous bone was first demonstrated for various porosities and cement intrusion depth. Three cements (Low Viscosity Cement [LVC], Simplex-P, and Palacos) were intruded at three different pressures (20, 40, and 60 PSI) at the same ambient temperature and relative humidity into commercially prepared plugs of bovine cancellous bone. Cement intrusion depth was proportional to injection pressure for all three cements, but was significantly different for each cement at a given pressure. At 20, 40, and 60 PSI, Palacos had a cement intrusion depth of 1.4, 2.4, and 2.8 mm respectively, while the figures for Simplex-P were 2.2, 4.2, and 5.0 mm, and for LVC were 8.0, 12.0, and 14.6 mm. Ambient temperature had an inverse relationship with cement intrusion depth for all three cements given the same experimental conditions. Chilling the monomer increased the intrusion of Simplex-P to 5.8, 8.2, and 12.7 mm at 20, 40, and 60 PSI injection pressure respectively. Simplex-P intrusion depth was not modified by cement centrifugation at any of the three injection pressures tested.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3802646

  12. Activation of Adenosine Monophosphate-Activated Protein Kinase Is an Additional Mechanism That Participates in Mediating Inhibitory Actions of Prostaglandin F2Alpha in Mature, but Not Developing, Bovine Corpora Lutea.

    PubMed

    Bowdridge, Elizabeth C; Goravanahally, Madhusudan P; Inskeep, E Keith; Flores, Jorge A

    2015-07-01

    Elevated cytosolic calcium and protein kinase C are well-established mediators of luteolytic actions of prostaglandin F2alpha (PGF2alpha). The objectives of this study were to determine 1) if calcium/calmodulin-dependent kinase kinase 2 (CAMKK2) participates in mediating PGF2alpha actions in developing (Day [d]-4) and mature (d-10) bovine corpus luteum (CL), 2) distal targets of CAMKK2, 3) developmental expression of adenosine monophosphate-activated protein kinase (AMPK), and 4) effects of AMPK activation on progesterone (P4) production. Expression of AMPK increased as the CL matured. Activation of the prostaglandin receptor (FP) induced rapid phosphorylation of AMPK, which was blocked by a CAMKK2 inhibitor. Changes in basal P4 secretion in vitro were determined in response to AMPK activation via metformin (met) or 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) in d-4 and d-10 CL. Production of P4 in d-10 CL decreased with met or AICAR compared to control, similar to activation by PGF2alpha. Therefore, potential distal targets of AMPK in d-10 CL were examined during induced functional regression via exogenous PGF2alpha. Serum and luteal P4 decreased at 2 and 4?h after administration of PGF2alpha. Protein expression of LDLR decreased at 2 and 4 h, while those of ACAT1 and STAR increased 4 h after PGF2alpha. During induced regression, alterations of cholesterol transport proteins contributed to decreased luteal and serum P4. Therefore, developmental differences in signal transduction associated with FP, specifically CAMKK2 and AMPK, partially contribute to differences in the ability of PGF2alpha to induce regression in mature, but not developing, bovine CL. Multiple cholesterol transport proteins, including LDLR, were altered by PGF2alpha and could be potential AMPK targets. PMID:25972015

  13. Analysis of osmotic pressure data for aqueous protein solutions via a multicomponent model

    NASA Astrophysics Data System (ADS)

    Druchok, M.; Kalyuzhnyi, Yu.; Reš?i?, J.; Vlachy, V.

    2006-03-01

    Integral equation theories and Monte Carlo simulations were used to study the Donnan equilibrium, which is established by an equilibrium distribution of a simple electrolyte between an aqueous protein-electrolyte mixture and an aqueous solution of the same simple electrolyte, when these two phases are separated by a semipermeable membrane. In order to describe the unusually low osmotic pressure found in many experiments we assumed that protein molecules can form dimers. The model solution contains proteins in a monomeric form, represented as charged hard spheres, or in a dimerized form, modeled as fused charged hard spheres. The counterions and coions were also modeled as charged hard spheres but of a much smaller size. The associative mean spherical and hypernetted-chain approximations were applied to this model. In addition, Monte Carlo computer simulations were performed for the same model system mimicking a lysozyme solution in the presence of 0.1M sodium chloride. Theory and simulations were found to be in reasonably good agreement for the thermodynamic properties such as chemical potential and osmotic pressure under these conditions. Using the theoretical approaches mentioned above, we analyzed the experimental data for the osmotic pressure of bovine serum albumin in 0.15M sodium chloride, human serum albumin solution (HSA) in 0.1M phosphate buffer, and lysozyme in sulphate and phosphate buffers. The theoretically determined osmotic coefficients were fitted to the existing experimental data in order to obtain the fraction of dimers in solution. Our analysis indicated that there was relatively small self-association of protein molecules for bovine serum albumin solutions at pH =5.4 and 7.3, with the fraction of dimers smaller than 10%, while at pH =4.5 the dimer fraction was equal to 50%. In the case of HSA solutions, strong negative deviations from the ideal value were found and at pH =8.0 a reasonably good agreement between the theory and experiment is obtained by assuming full dimerization. For HSA solution at pH =5.4, the best fit to the experimental results was obtained for a fraction of dimers equal to 80%.

  14. Microwave radiation can alter protein conformation without bulk heating

    Microsoft Academic Search

    David I. de Pomerai; Brette Smith; Adam Dawe; Kate North; Tim Smith; David B. Archer; Ian R. Duce; Donald Jones; E. Peter M. Candido

    2003-01-01

    Exposure to microwave radiation enhances the aggregation of bovine serum albumin in vitro in a time- and temperature-dependent manner. Microwave radiation also promotes amyloid fibril formation by bovine insulin at 60°C. These alterations in protein conformation are not accompanied by measurable temperature changes, consistent with estimates from field modelling of the specific absorbed radiation (15–20 mW kg?1). Limited denaturation of

  15. DNA vaccine-derived human IgG produced in transchromosomal bovines protect in lethal models of hantavirus pulmonary syndrome.

    PubMed

    Hooper, Jay W; Brocato, Rebecca L; Kwilas, Steven A; Hammerbeck, Christopher D; Josleyn, Matthew D; Royals, Michael; Ballantyne, John; Wu, Hua; Jiao, Jin-an; Matsushita, Hiroaki; Sullivan, Eddie J

    2014-11-26

    Polyclonal immunoglobulin-based medical products have been used successfully to treat diseases caused by viruses for more than a century. We demonstrate the use of DNA vaccine technology and transchromosomal bovines (TcBs) to produce fully human polyclonal immunoglobulins (IgG) with potent antiviral neutralizing activity. Specifically, two hantavirus DNA vaccines [Andes virus (ANDV) DNA vaccine and Sin Nombre virus (SNV) DNA vaccine] were used to produce a candidate immunoglobulin product for the prevention and treatment of hantavirus pulmonary syndrome (HPS). A needle-free jet injection device was used to vaccinate TcB, and high-titer neutralizing antibodies (titers >1000) against both viruses were produced within 1 month. Plasma collected at day 10 after the fourth vaccination was used to produce purified ?-HPS TcB human IgG. Treatment with 20,000 neutralizing antibody units (NAU)/kg starting 5 days after challenge with ANDV protected seven of eight animals, whereas zero of eight animals treated with the same dose of normal TcB human IgG survived. Likewise, treatment with 20,000 NAU/kg starting 5 days after challenge with SNV protected immunocompromised hamsters from lethal HPS, protecting five of eight animals. Our findings that the ?-HPS TcB human IgG is capable of protecting in animal models of lethal HPS when administered after exposure provides proof of concept that this approach can be used to develop candidate next-generation polyclonal immunoglobulin-based medical products without the need for human donors, despeciation protocols, or inactivated/attenuated vaccine antigen. PMID:25429055

  16. Interaction of a potential chloride channel blocker with a model transport protein: a spectroscopic and molecular docking investigation.

    PubMed

    Ganguly, Aniruddha; Paul, Bijan Kumar; Ghosh, Soumen; Dalapati, Sasanka; Guchhait, Nikhil

    2014-05-14

    The present work demonstrates a detailed characterization of the interaction of a potential chloride channel blocker, 9-methyl anthroate (9-MA), with a model transport protein, Bovine Serum Albumin (BSA). The modulated photophysical properties of the emissive drug molecule within the microheterogeneous bio-environment of the protein have been exploited spectroscopically to monitor the probe-protein binding interaction. Apart from evaluating the binding constant, the probable location of the neutral molecule within the protein cavity (subdomain IB) is explored by an AutoDock-based blind docking simulation. The absence of the Red-Edge Effect has been corroborated by the enhanced lifetime of the probe, being substantially greater than the solvent reorientation time. A dip-and-rise characteristic of the rotational relaxation profile of the drug within the protein has been argued to originate from a significant difference in the lifetime as well as amplitude of the free and protein-bound drug molecule. Unfolding of the protein in the presence of the drug molecule has been probed by the decrease of the ?-helical content, obtained via circular dichroism (CD) spectroscopy, which is also supported by the gradual loss of the esterase activity of the protein in the presence of the drug molecule. PMID:24668158

  17. Identification of polymorphisms in the promoter region of the bovine heat shock protein gene and associations with bull calf weaning weight

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Our objective was to evaluate the relationship between genotypic variation of the bovine HSP-70 promoter and bull calf weaning weights and serum concentrations of HSP-70 at weaning. Blood samples were collected from 33 crossbred bull calves. Calves were sired by Angus bulls and had Brahman-cross dam...

  18. Identification of single nucleotide polymorphisms within the promoter region of the bovine heat shock protein 70 gene and associations with pregnancy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Objectives were: 1) determine genetic diversity in a promoter segment of the bovine HSP-70 gene, and 2) determine if the identified single nucleotide polymorphisms (SNPs) were related to pregnancy rates. Genomic DNA was collected from 107 Bos taurus/Bos indicus crossbred cows. Specific primers (HSP-...

  19. Two state model for a constant disease hazard in paratuberculosis (and other bovine diseases).

    PubMed

    Louzoun, Yoram; Mitchell, Rebecca; Behar, Hilla; Schukken, Ynte

    2015-01-01

    Many diseases are characterized by a long and varying sub-clinical period. Two main mechanisms can explain such periods: a slow progress toward disease or a sudden transition from a healthy state to a disease state induced by internal or external events. We here survey epidemiological features of the amount of bacteria shed during Mycobacterium Avium Paratuberculosis (MAP) infection to test which of these two models, slow progression or sudden transition (or a combination of the two), better explains the transition from intermittent and low shedding to high shedding. Often, but not always, high shedding is associated with the occurrence of clinical signs. In the case of MAP, the clinical signs include diarrhea, low milk production, poor fertility and eventually emaciation and death. We propose a generic model containing bacterial growth, immune control and fluctuations. This proposed generic model can represent the two hypothesized types of transitions in different parameter regimes. The results show that the sudden transition model provides a simpler explanation of the data, but also suffers from some limitations. We discuss the different immunological mechanism that can explain and support the sudden transition model and the interpretation of each term in the studied model. These conclusions are applicable to a wide variety of diseases, and MAP serves as a good test case based on the large scale measurements of single cow longitudinal profiles in this disease. PMID:26092587

  20. CSAW: a dynamical model of protein folding

    E-print Network

    Kerson Huang

    2006-01-12

    CSAW (conditioned self-avoiding walk) is a model of protein folding that combines SAW (self-avoiding walk) with Monte-Carlo. It simulates the Brownian motion of a chain molecule in the presence of interactions, both among chain residues, and with the environment. In a first model that includes the hydrophobic effect and hydrogen bonding, a chain of 30 residues folds into a native state with stable secondary and tertiary structures. The process starts with a rapid collapse into an intermediate "molten globule", which slowly decays into the native state afer a relatively long quiescent period. The behavior of the radius of gyration mimics experimental data.

  1. Protein viscoelastic dynamics: a model system

    E-print Network

    Craig Fogle; Joseph Rudnick; David Jasnow

    2015-02-02

    A model system inspired by recent experiments on the dynamics of a folded protein under the influence of a sinusoidal force is investigated and found to replicate many of the response characteristics of such a system. The essence of the model is a strongly over-damped oscillator described by a harmonic restoring force for small displacements that reversibly yields to stress under sufficiently large displacement. This simple dynamical system also reveals unexpectedly rich behavior, exhibiting a series of dynamical transitions and analogies with equilibrium thermodynamic phase transitions. The effects of noise and of inertia are briefly considered and described.

  2. Fibrous proteins—silk as a model system

    Microsoft Academic Search

    David L. Kaplan

    1998-01-01

    The repetitive nature of fibrous proteins leads to regular secondary and higher order structures. This attribute suggests that these polymers are useful models to gain insight into structure-function relationships. Genetic variants of these proteins can be constructed from precisely defined synthetic oligonucleotides to encode the specific sized protein of interest. The biosynthesis and assembly of this type of protein is

  3. Reconstitution of Membrane Proteins into Model Membranes: Seeking Better Ways to Retain Protein Activities

    PubMed Central

    Shen, Hsin-Hui; Lithgow, Trevor; Martin, Lisandra L.

    2013-01-01

    The function of any given biological membrane is determined largely by the specific set of integral membrane proteins embedded in it, and the peripheral membrane proteins attached to the membrane surface. The activity of these proteins, in turn, can be modulated by the phospholipid composition of the membrane. The reconstitution of membrane proteins into a model membrane allows investigation of individual features and activities of a given cell membrane component. However, the activity of membrane proteins is often difficult to sustain following reconstitution, since the composition of the model phospholipid bilayer differs from that of the native cell membrane. This review will discuss the reconstitution of membrane protein activities in four different types of model membrane—monolayers, supported lipid bilayers, liposomes and nanodiscs, comparing their advantages in membrane protein reconstitution. Variation in the surrounding model environments for these four different types of membrane layer can affect the three-dimensional structure of reconstituted proteins and may possibly lead to loss of the proteins activity. We also discuss examples where the same membrane proteins have been successfully reconstituted into two or more model membrane systems with comparison of the observed activity in each system. Understanding of the behavioral changes for proteins in model membrane systems after membrane reconstitution is often a prerequisite to protein research. It is essential to find better solutions for retaining membrane protein activities for measurement and characterization in vitro. PMID:23344058

  4. Knowledge-based model building of proteins: concepts and examples.

    PubMed Central

    Bajorath, J.; Stenkamp, R.; Aruffo, A.

    1993-01-01

    We describe how to build protein models from structural templates. Methods to identify structural similarities between proteins in cases of significant, moderate to low, or virtually absent sequence similarity are discussed. The detection and evaluation of structural relationships is emphasized as a central aspect of protein modeling, distinct from the more technical aspects of model building. Computational techniques to generate and complement comparative protein models are also reviewed. Two examples, P-selectin and gp39, are presented to illustrate the derivation of protein model structures and their use in experimental studies. PMID:7505680

  5. Homology Modeling and Analysis of Structure Predictions of the Bovine Rhinitis B Virus RNA Dependent RNA Polymerase (RdRp)

    PubMed Central

    Rai, Devendra K.; Rieder, Elizabeth

    2012-01-01

    Bovine Rhinitis B Virus (BRBV) is a picornavirus responsible for mild respiratory infection of cattle. It is probably the least characterized among the aphthoviruses. BRBV is the closest relative known to Foot and Mouth Disease virus (FMDV) with a ~43% identical polyprotein sequence and as much as 67% identical sequence for the RNA dependent RNA polymerase (RdRp), which is also known as 3D polymerase (3Dpol). In the present study we carried out phylogenetic analysis, structure based sequence alignment and prediction of three-dimensional structure of BRBV 3Dpol using a combination of different computational tools. Model structures of BRBV 3Dpol were verified for their stereochemical quality and accuracy. The BRBV 3Dpol structure predicted by SWISS-MODEL exhibited highest scores in terms of stereochemical quality and accuracy, which were in the range of 2Å resolution crystal structures. The active site, nucleic acid binding site and overall structure were observed to be in agreement with the crystal structure of unliganded as well as template/primer (T/P), nucleotide tri-phosphate (NTP) and pyrophosphate (PPi) bound FMDV 3Dpol (PDB, 1U09 and 2E9Z). The closest proximity of BRBV and FMDV 3Dpol as compared to human rhinovirus type 16 (HRV-16) and rabbit hemorrhagic disease virus (RHDV) 3Dpols is also substantiated by phylogeny analysis and root-mean square deviation (RMSD) between C-? traces of the polymerase structures. The absence of positively charged ?-helix at C terminal, significant differences in non-covalent interactions especially salt bridges and CH-pi interactions around T/P channel of BRBV 3Dpol compared to FMDV 3Dpol, indicate that despite a very high homology to FMDV 3Dpol, BRBV 3Dpol may adopt a different mechanism for handling its substrates and adapting to physiological requirements. Our findings will be valuable in the design of structure-function interventions and identification of molecular targets for drug design applicable to Aphthovirus RdRps. PMID:22942748

  6. Evaluation of pulmonary dysfunctions and acid–base imbalances induced by Chlamydia psittaci in a bovine model of respiratory infection

    PubMed Central

    2014-01-01

    Background Chlamydia psittaci (Cp) is a respiratory pathogen capable of inducing acute pulmonary zoonotic disease (psittacosis) or persistent infection. To elucidate the pathogenesis of this infection, a translational large animal model was recently introduced by our group. This study aims at quantifying and differentiating pulmonary dysfunction and acid–base imbalances induced by Cp. Methods Forty-two calves were grouped in (i) animals inoculated with Cp (n?=?21) and (ii) controls sham-inoculated with uninfected cell culture (n?=?21). For pulmonary function testing, impulse oscillometry, capnography, and FRC (functional residual capacity) measurement were applied to spontaneously breathing animals. Variables of acid–base status were assessed in venous blood using both (i) traditional Henderson-Hasselbalch and (ii) strong ion approach. Results Both obstructive and restrictive pulmonary disorders were induced in calves experimentally inoculated with Cp. Although disorders in respiratory mechanics lasted for 8–11 days, the pattern of spontaneous breathing was mainly altered in the period of acute illness (until 4 days post inoculation, dpi). Expiration was more impaired than inspiration, resulting in elevated FRC. Ventilation was characterised by a reduction in tidal volume (?25%) combined with an increased percentage of dead space volume and a significant reduction of alveolar volume by 10%. Minute ventilation increased significantly (+50%) due to a compensatory doubling of respiratory rate. Hyperventilatory hypocapnia at 2–3 dpi resulted in slightly increased blood pH at 2 dpi. However, the acid–base equilibrium was additionally influenced by metabolic components, i.e. the systemic inflammatory response, all of which were detected with help of the strong ion theory. Decreased concentrations of albumin (2–10 dpi), a negative acute-phase marker, resulted in a decrease in the sum of non-volatile weak acids (Atot), revealing an alkalotic effect. This was counterbalanced by acidic effects of decreased strong ion difference (SID), mediated by the interplay between hypochloraemia (alkalotic effect) and hyponatraemia (acidic effect). Conclusions This bovine model was found to be suitable for studying pathophysiology of respiratory Cp infection and may help elucidating functional host-pathogen interactions in the mammalian lung. PMID:24517577

  7. Estrogen Receptor Expression in Bovine and Rat Retinas

    Microsoft Academic Search

    Kaori Kobayashi; Hiroshi Kobayashi; Masamichi Ueda; Yoshihito Honda

    PURPOSE. TO investigate the expression and distribution of estrogen receptor protein and mRNA in bovine and rat retinas. METHODS. Western blot analysis with an antiestrogen receptor monoclonal antibody (mAb) was used to detect estrogen receptor protein in the bovine retina. Immunohistochemistry with an antiestrogen receptor mAb and in situ hybridization with an oligodeoxynucleotide sequence coding for estrogen receptor were applied

  8. Expression and localization of progesterone receptor membrane component 1 and 2 and serpine mRNA binding protein 1 in the bovine corpus luteum during the estrous cycle and the first trimester of pregnancy.

    PubMed

    Kowalik, Magdalena K; Rekawiecki, Robert; Kotwica, Jan

    2014-11-01

    The aim of this study was to evaluate the mRNA and protein expression and the localization of progesterone receptor membrane component 1 (PGRMC1), PGRMC2, and the PGRMC1 partner serpine mRNA binding protein 1 (SERBP1) in the bovine CL on Days 2 to 5, 6 to 10, 11 to 16, and 17 to 20 of the estrous cycle as well as during Weeks 3 to 5, 6 to 8, and 9 to 12 of pregnancy (n = 5-6 per each period). The highest levels of PGRMC1 and PGRMC2 mRNA expression were found on Days 6 to 16 (P < 0.05) and 11 to 16, respectively, of the estrous cycle and during pregnancy (P < 0.001). The level of PGRMC1 protein was the highest (P < 0.05) on Days 11 to 16 of the estrous cycle compared with the other stages of the estrous cycle and pregnancy, whereas PGRMC2 protein expression (P < 0.001) was the highest on Days 17 to 20 and also during pregnancy. The mRNA expression of SERBP1 was increased (P < 0.05) on Days 11 to 16, whereas the level of its protein product was decreased (P < 0.05) on Days 6 to 10 of the estrous cycle and was at its lowest (P < 0.001) on Days 17 to 20. In pregnant cows, the patterns of SERBP1 mRNA and protein expression remained constant and were comparable with those observed during the estrous cycle. Progesterone receptor membrane component 1 and PGRMC2 localized to both large and small luteal cells, whereas SERBP1 was observed mainly in small luteal cells and much less frequently in large luteal cells. All proteins were also localized in the endothelial cells of blood vessels. The data obtained indicate the variable expression of PGRMC1, PGRMC2, and SERBP1 mRNA and protein in the bovine CL and suggest that progesterone may regulate CL function via its membrane receptors during both the estrous cycle and pregnancy. PMID:25168721

  9. In bovine binucleate trophoblast giant cells, pregnancy-associated glycoproteins and placental prolactin-related protein-I are conjugated to asparagine-linked N -acetylgalactosaminyl glycans

    Microsoft Academic Search

    Karl Klisch; Rudolf Leiser

    2003-01-01

    Bovine binucleate trophoblast giant cells (BNCs) produce large amounts of PAS-positive cytoplasmic granules. After fusion of BNCs with uterine epithelial cells, the contents of these granules are released into the maternal stroma which underlies the uterine epithelium. Histochemically, the granules can be labeled with N-acetylgalactosamine-specific lectins (Dolichos biflorus, Vicia villosa, and Wisteria floribunda agglutinins) and with Phaseolus vulgaris leucoagglutinin. In

  10. Proteolysis of Specific Muscle Structural Proteins by m-Calpain at Low pH and Temperature Is Similar to Degradation in Postmortem Bovine Muscle1,2

    Microsoft Academic Search

    Elisabeth Huff-Lonergan; Tomiko Mitsuhashi; Dirk D. Beekman; F. C. Parrish; Dennis G. Olson; Richard M. Robson

    2010-01-01

    Postmortem (PM) and m-calpain- induced degradation of specific skeletal muscle pro- teins was monitored by SDS-PAGE and Western blotting. Samples were removed from bovine longissi- mus thoracis (LT) at approximately 45 min PM for the preparation of at-death (0-d) myofibrils (MF). The LT was excised at 1 d PM, vacuum-packaged, and stored at 2°C. Samples were removed for Warner- Bratzler

  11. Modeling Protein Folding and Applying It to a Relevant Activity

    ERIC Educational Resources Information Center

    Nelson, Allan; Goetze, Jim

    2004-01-01

    The different levels of protein structure that can be easily understood by creating a model that simulates protein folding, which can then be evaluated by applying it to a relevant activity, is presented. The materials required and the procedure for constructing a protein folding model are mentioned.

  12. Thermodynamics of Protein Folding from Coarse-Grained Models' Perspectives

    E-print Network

    Janke, Wolfhard

    8 Thermodynamics of Protein Folding from Coarse-Grained Models' Perspectives Michael Bachmann applications. In this lecture, we focus on the anal- ysis of mesoscopic models for protein folding, aggregation for a more universal description of the notoriously difficult problem of protein fold- ing. In this approach

  13. Modeling Protein Folding Pathways Christopher Bystroff, Yu Shao

    E-print Network

    Bystroff, Chris

    Modeling Protein Folding Pathways Christopher Bystroff, Yu Shao Dept of Biology Rensselaer Polytechnic Institute, Troy, NY. e-mail:{bystrc, shaoy}@rpi.edu Summary Proteins fold through a series of intermediate states called a pathway. Protein folding pathways have been modeled using either simulations

  14. Protein Structure Prediction by Comparative Modeling: An Analysis of Methodology

    E-print Network

    Protein Structure Prediction by Comparative Modeling: An Analysis of Methodology Jennifer Wang, Biochemistry 218 Submitted December 11, 2009 1. Introduction Protein structure determination has become an important area of research in molecular biology and structural genomics. Understanding the tertiary

  15. Modeling Intrinsically Disordered Proteins with Bayesian Statistics

    PubMed Central

    2010-01-01

    The characterization of intrinsically disordered proteins is challenging because accurate models of these systems require a description of both their thermally accessible conformers and the associated relative stabilities or weights. These structures and weights are typically chosen such that calculated ensemble averages agree with some set of prespecified experimental measurements; however, the large number of degrees of freedom in these systems typically leads to multiple conformational ensembles that are degenerate with respect to any given set of experimental observables. In this work we demonstrate that estimates of the relative stabilities of conformers within an ensemble are often incorrect when one does not account for the underlying uncertainty in the estimates themselves. Therefore, we present a method for modeling the conformational properties of disordered proteins that estimates the uncertainty in the weights of each conformer. The Bayesian weighting (BW) formalism incorporates information from both experimental data and theoretical predictions to calculate a probability density over all possible ways of weighting the conformers in the ensemble. This probability density is then used to estimate the values of the weights. A unique and powerful feature of the approach is that it provides a built-in error measure that allows one to assess the accuracy of the ensemble. We validate the approach using reference ensembles constructed from the five-residue peptide met-enkephalin and then apply the BW method to construct an ensemble of the K18 isoform of the tau protein. Using this ensemble, we indentify a specific pattern of long-range contacts in K18 that correlates with the known aggregation properties of the sequence. PMID:20925316

  16. SR proteins Asf/SF2 and 9G8 interact to activate enhancer-dependent intron D splicing of bovine growth hormone pre-mRNA in vitro.

    PubMed Central

    Li, X; Shambaugh, M E; Rottman, F M; Bokar, J A

    2000-01-01

    The alternative splicing of the last intron (intron D) of bovine growth hormone (bGH) pre-mRNA requires a down-stream exonic splicing enhancer (FP/ESE). The presence of at least one SR protein has been shown to be essential for FP/ESE function and splicing of intron D in in vitro splicing assays. However, in vitro reconstitution of splicing using individual purified SR proteins may not accurately reflect the true complexity of alternative splicing in an intact nucleus, where multiple SR proteins in varying amounts are likely to be available simultaneously. Here, a panel of recombinant baculovirus-expressed SR proteins was produced and tested for the ability to activate FP/ESE-dependent splicing. Individual recombinant SR proteins differed significantly in their activity in promoting intron D splicing. Among the recombinant SR proteins tested, SRp55 was the most active, SC35 showed very little activity, and ASF/SF2 and 9G8 individually had intermediate activity. At least one SR protein (ASF/SF2) bound to the FP/ESE with characteristics of a cooperative interaction. Most interestingly, low concentrations of ASF/SF2 and 9G8 acted synergistically to activate intron D splicing. This was due in part to synergistic binding to the FP/ESE. Splicing of bGH intron D is inherently complex, and is likely controlled by an interaction of the FP/ESE with several trans-acting protein factors acting both independently and cooperatively. This level of complexity may be required for precise control of alternative splicing by an exon sequence, which simultaneously is constrained to maintain translational integrity of the mature mRNA. PMID:11142383

  17. Unlocking the bovine genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The draft genome sequence of cattle (Bos taurus) has now been analyzed by the Bovine Genome Sequencing and Analysis Consortium and the Bovine HapMap Consortium, which together represent an extensive collaboration involving more than 300 scientists from 25 different countries. ...

  18. IDENTIFICATION OF BOVINE MICRORNAS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs are small ~22 nucleotides-long non-coding RNAs capable of controlling gene expression by inhibiting translation or targeting messenger RNA for degradation. Bovine genome sequence is not yet annotated for the microRNAs and there are currently no bovine miRNAs reported in the miRBase. Alignm...

  19. FLUORESCEIN-CONJUGATED BOVINE ALBUMIN

    PubMed Central

    Schiller, Alfred A.; Schayer, Richard W.; Hess, E. L.

    1953-01-01

    Fluorescein-bovine albumin conjugates have been prepared and found not to differ appreciably in size, shape, and homogeneity from the precursor, bovine serum albumin. Fluorescein has also been conjugated to rat plasma proteins. Their disappearance rates from the circulation of rats correspond with those obtained from the use of isotope labeling. Their sites of localization in rat tissues were shown to be in the cytoplasm but not in the nuclei of Kupffer cells, fixed macrophages, granulocytes, and proximal renal tubules. Adsorption to endothelium was a characteristic finding. Extracellular localizations were predominantly in the lumina of blood vessels and proximal renal tubules (but never in the lumina of collecting tubules), and the interstitial fluid of skeletal and cardiac muscle (but not that of glandular organs such as the adrenals, liver, and spleen). BAC absorption from the skin of rabbits requires days whereas sodium fluorescein absorption is measured in hours, attesting to the persistence of the colloidal state of BAC in vivo. Fluorescein conjugates have been used to visualize the transcapillary passage of circulating proteins in the mesenteric circulation of frogs and rats by direct microscopic observation and found to diffuse slowly in the manner predicted for plasma proteins. The normal cutaneous vessels of the rat are impermeable in the gross to the labeled proteins; second degree burn promptly increases the permeability of these vessels rendering the presence of the label detectable in the gross in the skin. The process of labeling does not render guinea pig albumin antigenic, although slight antigenicity results from labeling whole plasma protein. It is believed that sufficient biological evidence is presented to support the conclusion that fluorescein-conjugated plasma proteins, particularly albumin, behave in vivo like their native precursors. PMID:13035065

  20. The Fibronectin-Binding Proteins of Staphylococcus aureus May Promote Mammary Gland Colonization in a Lactating Mouse Model of Mastitis

    Microsoft Academic Search

    Eric Brouillette; Brian G. Talbot; Francois Malouin

    2003-01-01

    The fibronectin-binding proteins (FnBPs) of Staphylococcus aureus are believed to be implicated in the pathogen's adherence to and colonization of bovine mammary glands, thus leading to infectious mastitis. In vitro studies have shown that FnBPs help the adhesion of the pathogen to bovine mammary epithelial cells. However, the importance of FnBPs for the infection of mammary glands has never been

  1. Structural libraries of protein models for multiple species to understand evolution of the renin-angiotensin system.

    PubMed

    Prokop, Jeremy W; Petri, Victoria; Shimoyama, Mary E; Watanabe, Ingrid K M; Casarini, Dulce E; Leeper, Thomas C; Bilinovich, Stephanie M; Jacob, Howard J; Santos, Robson A S; Martins, Almir S; Araujo, Fabiano C; Reis, Fernando M; Milsted, Amy

    2015-05-01

    The details of protein pathways at a structural level provides a bridge between genetics/molecular biology and physiology. The renin-angiotensin system is involved in many physiological pathways with informative structural details in multiple components. Few studies have been performed assessing structural knowledge across the system. This assessment allows use of bioinformatics tools to fill in missing structural voids. In this paper we detail known structures of the renin-angiotensin system and use computational approaches to estimate and model components that do not have their protein structures defined. With the subsequent large library of protein structures, we then created a species specific protein library for human, mouse, rat, bovine, zebrafish, and chicken for the system. The rat structural system allowed for rapid screening of genetic variants from 51 commonly used rat strains, identifying amino acid variants in angiotensinogen, ACE2, and AT1b that are in contact positions with other macromolecules. We believe the structural map will be of value for other researchers to understand their experimental data in the context of an environment for multiple proteins, providing pdb files of proteins for the renin-angiotensin system in six species. With detailed structural descriptions of each protein, it is easier to assess a species for use in translating human diseases with animal models. Additionally, as whole genome sequencing continues to decrease in cost, tools such as molecular modeling will gain use as an initial step in designing efficient hypothesis driven research, addressing potential functional outcomes of genetic variants with precompiled protein libraries aiding in rapid characterizations. PMID:25260253

  2. Tactile Teaching: Exploring Protein Structure/Function Using Physical Models

    ERIC Educational Resources Information Center

    Herman, Tim; Morris, Jennifer; Colton, Shannon; Batiza, Ann; Patrick, Michael; Franzen, Margaret; Goodsell, David S.

    2006-01-01

    The technology now exists to construct physical models of proteins based on atomic coordinates of solved structures. We review here our recent experiences in using physical models to teach concepts of protein structure and function at both the high school and the undergraduate levels. At the high school level, physical models are used in a…

  3. MATHEMATICAL MODELS OF PROTEIN FOLDING Daniel B. Dix

    E-print Network

    Dix, Daniel B.

    MATHEMATICAL MODELS OF PROTEIN FOLDING Daniel B. Dix Department of Mathematics University of South Carolina Abstract. We present an elementary introduction to the protein folding problem directed toward, and biological problem, protein folding can also be precisely formulated as a set of mathematics problems. We

  4. Mechanical Modeling and Computer Simulation of Protein Folding

    ERIC Educational Resources Information Center

    Prigozhin, Maxim B.; Scott, Gregory E.; Denos, Sharlene

    2014-01-01

    In this activity, science education and modern technology are bridged to teach students at the high school and undergraduate levels about protein folding and to strengthen their model building skills. Students are guided from a textbook picture of a protein as a rigid crystal structure to a more realistic view: proteins are highly dynamic…

  5. Exploiting Models of Molecular Evolution to Efficiently Direct Protein Engineering

    E-print Network

    Gaucher, Eric

    Exploiting Models of Molecular Evolution to Efficiently Direct Protein Engineering Megan F. Cole Ó Springer Science+Business Media, LLC 2010 Abstract Directed evolution and protein engineering evolution and protein engineering that exploits phylogenetic and sequence analyses to identify amino acid

  6. Bayesian Models and Algorithms for Protein Beta-Sheet Prediction

    E-print Network

    Erdogan, Hakan

    0 Bayesian Models and Algorithms for Protein Beta-Sheet Prediction Zafer Aydin, Student Member-parallel), and -residue interactions (or contact maps). We introduce a Bayesian approach for proteins with six or less effectively. For proteins with more than six -strands, we first compute -strand pairings using the Beta

  7. Completion of autobuilt protein models using a database of protein fragments

    SciTech Connect

    Cowtan, Kevin, E-mail: cowtan@ysbl.york.ac.uk [Department of Chemistry, University of York, Heslington, York YO10 5DD (United Kingdom)

    2012-04-01

    Two developments in the process of automated protein model building in the Buccaneer software are described: the use of a database of protein fragments in improving the model completeness and the assembly of disconnected chain fragments into complete molecules. Two developments in the process of automated protein model building in the Buccaneer software are presented. A general-purpose library for protein fragments of arbitrary size is described, with a highly optimized search method allowing the use of a larger database than in previous work. The problem of assembling an autobuilt model into complete chains is discussed. This involves the assembly of disconnected chain fragments into complete molecules and the use of the database of protein fragments in improving the model completeness. Assembly of fragments into molecules is a standard step in existing model-building software, but the methods have not received detailed discussion in the literature.

  8. Mechanisms of m-cresol induced protein aggregation studied using a model protein cytochrome c†

    PubMed Central

    Singh, Surinder M.; Hutchings, Regina L.; Mallela, Krishna M.G.

    2014-01-01

    Multi-dose protein formulations require an effective antimicrobial preservative (AP) to inhibit microbial growth during long-term storage of unused formulations. m-cresol is one such AP, but has been shown to cause protein aggregation. However, the fundamental physical mechanisms underlying such AP-induced protein aggregation are not understood. In this study, we used a model protein cytochrome c to identify the protein unfolding that triggers protein aggregation. m-cresol induced cytochrome c aggregation at preservative concentrations that are commonly used to inhibit microbial growth. Addition of m-cresol decreased the temperature at which the protein aggregated and increased the aggregation rate. However, m-cresol did not perturb the tertiary or secondary structure of cytochrome c. Instead, it populated an “invisible” partially unfolded intermediate where a local protein region around the methionine residue at position 80 was unfolded. Stabilizing the Met80 region drastically decreased the protein aggregation, which conclusively shows that this local protein region acts as an aggregation “hot-spot”. Based on these results, we propose that APs induce protein aggregation by partial rather than global unfolding. Because of the availability of site-specific probes to monitor different levels of protein unfolding, cytochrome c provided a unique advantage in characterizing the partial protein unfolding that triggers protein aggregation. PMID:21229618

  9. Identification and isolation of two pentraxins from bovine serum.

    PubMed

    Maudsley, S; Rowe, I F; de Beer, F C; Munn, E A; Herbert, J; Feinstein, A; Pepys, M B

    1987-03-01

    Two distinct pentraxin proteins were isolated from bovine serum by calcium-dependent affinity chromatography on high pyruvate agarose gel. One of these proteins cross-reacted specifically with certain rabbit anti-human CRP antisera and was therefore designated as bovine CRP. The other cross-reacted specifically with a sheep anti-human SAP antiserum and was therefore designated as bovine SAP. Although the mixture of these two pentraxins was not resolved by gel filtration chromatography, they were separated by solid phase absorption of the CRP with immobilized rabbit anti-human CRP antibodies. Their identity as pentraxins was confirmed by their electron microscopic appearance. Bovine CRP was composed of a single type of non-glycosylated subunit whilst bovine SAP contained two major types of glycosylated subunits and a minor polypeptide, the glycosylation of which was not determined. Serum concentrations were in the range of 5-40 mg/l and neither protein behaved as an acute phase reactant. No bovine serum protein undergoing calcium-dependent binding to phosphoryl choline or pneumococcal C-polysaccharide was obtained. PMID:3608235

  10. Accuracy of Protein-Protein Binding Sites in High-Throughput Template-Based Modeling

    E-print Network

    Kundrotas, Petras J.; Vakser, Ilya A.

    2010-04-01

    ranked 1 by a simple sequence identity criteria had RMSD<5 Å, the accuracy suitable for low-resolution template free docking. Such models corresponded to multi-domain target proteins, whereas for single-domain proteins the best models had 5 Å

  11. Human activated protein C variants in a rat model of arterial thrombosis

    PubMed Central

    Malm, Karl; Arnljots, Björn; Dahlbäck, Björn

    2008-01-01

    Background Activated protein C (APC) inhibits coagulation by degrading activated factor V (FVa) and factor VIII (FVIIIa), protein S (PS) functioning as a cofactor to APC. Methods By mutagenesis of the vitamin K-dependent Gla domain of APC, we have recently created an APC variant having enhanced anticoagulant activity due to increased affinity for negatively charged phospholipid membranes. In the present study, the potential antithrombotic effects of this APC variant, and of a variant APC that is additionally mutated in the serine protease domain, have been evaluated in a blind randomized study in a rat model of arterial thrombosis. In this model, we have previously found the combination of bovine APC and PS to be highly antithrombotic. Four treatment groups each containing 10 rats were, in a blind random fashion, given intravenous bolus injections of wild-type or mutant variants of APC (0.8 mg/kg) together with human PS (0.6 mg/kg) or human PS (0.6 mg/kg) alone. A control group with 20 animals where given vehicle only. Results A trend to increased patency rates was noted in a group receiving one of the APC variants, but it did not reach statistical significance. Conclusion In conclusion, administration of human APC variants having enhanced anticoagulant efficacy together with human PS in a rat model of arterial thrombosis did not give an efficient antithrombotic effect. The lack of effect may be due to species-specific differences between the human protein C system and the rat hemostatic system. PMID:18957140

  12. Infectious Bovine Rhinotracheitis 

    E-print Network

    Sprott, L. R.

    1998-11-30

    Infectious bovine rhinotracheitis (IBR) is a complex of disease syndromes occuring throughout the United States and the other major cattle-producing areas of the world. It affects cattle and some wild ruminants. This publication describes...

  13. Protection of the bovine fetus from bovine viral diarrhoea virus by means of a new inactivated vaccine

    Microsoft Academic Search

    J Brownlie; MC Clarke; LB Hooper; GD Bell

    1995-01-01

    A model is described for the validation of vaccines designed to protect the bovine fetus from bovine viral diarrhoea virus (BVDV). The fetopathic nature of the challenge strain of virus was confirmed and the method used to test a commercial vaccine (Bovidec) developed from a Compton prototype. Heifers were vaccinated two or three times about the time of impregnation and

  14. Virtual Ligand Screening Against Comparative Protein Structure Models

    E-print Network

    Sali, Andrej

    Chapter 8 Virtual Ligand Screening Against Comparative Protein Structure Models Hao Fan, John J. Irwin, and Andrej Sali Abstract Virtual ligand screening uses computation to discover new ligands. Comparative protein structure modeling extends the applicability of virtual screening beyond the atomic

  15. Spectral deciphering of the interaction between an intramolecular hydrogen bonded ESIPT drug, 3,5-dichlorosalicylic acid, and a model transport protein.

    PubMed

    Paul, Bijan Kumar; Ray, Debarati; Guchhait, Nikhil

    2012-07-01

    The present work demonstrates a detailed characterization of the interaction of a bio-active drug molecule 3,5-dichlorosalicyclic acid (3,5DCSA) with a model transport protein Bovine Serum Albumin (BSA). The drug molecule is a potential candidate exhibiting Excited-State Intramolecular Proton Transfer (ESIPT) reaction and the modulation of ESIPT photophysics within the bio-environment of the protein has been exploited spectroscopically to monitor the drug-protein binding interaction. Apart from evaluating the binding constant (K (±10%) = 394 M(-1)) the probable location of the neutral drug molecule within the protein cavity (hydrophobic subdomain IIA) is explored by AutoDock-based blind docking simulation. The rotational relaxation dynamics of the drug within the protein has been interpreted on the lexicon of the two-step and wobbling-in-cone model. Circular dichroism (CD) spectroscopy delineates the effect of drug binding on the protein secondary structure in terms of decrease of ?-helical content of BSA with increasing drug concentration. Also the esterase activity of the drug:protein conjugate system is found to be reduced in comparison to the native protein. PMID:22307316

  16. On the role of surface composition and curvature on biointerface formation and colloidal stability of nanoparticles in a protein-rich model system.

    PubMed

    Orts-Gil, Guillermo; Natte, Kishore; Thiermann, Raphael; Girod, Matthias; Rades, Steffi; Kalbe, Henryk; Thünemann, Andreas F; Maskos, Michael; Österle, Werner

    2013-08-01

    The need for a better understanding of nanoparticle-protein interactions and the mechanisms governing the resulting colloidal stability has been emphasised in recent years. In the present contribution, the short and long term colloidal stability of silica nanoparticles (SNPs) and silica-poly(ethylene glycol) nanohybrids (Sil-PEG) have been scrutinised in a protein model system. Well-defined silica nanoparticles are rapidly covered by bovine serum albumin (BSA) and form small clusters after 20min while large agglomerates are detected after 10h depending on both particle size and nanoparticle-protein ratio. Oppositely, Sil-PEG hybrids present suppressive protein adsorption and enhanced short and long term colloidal stability in protein solution. No critical agglomeration was found for either system in the absence of protein, proving that instability found for SNPs must arise as a consequence of protein adsorption and not to high ionic environment. Analysis of the small angle X-ray scattering (SAXS) structure factor indicates a short-range attractive potential between particles in the silica-BSA system, which is in good agreement with a protein bridging agglomeration mechanism. The results presented here point out the importance of the nanoparticle surface properties on the ability to adsorb proteins and how the induced or depressed adsorption may potentially drive the resulting colloidal stability. PMID:23528607

  17. Role of G protein-coupled estrogen receptor-1, matrix metalloproteinases 2 and 9, and heparin binding epidermal growth factor-like growth factor in estradiol-17?-stimulated bovine satellite cell proliferation.

    PubMed

    Kamanga-Sollo, E; Thornton, K J; White, M E; Dayton, W R

    2014-10-01

    In feedlot steers, estradiol-17? (E2) and combined E2 and trenbolone acetate (a testosterone analog) implants enhance rate and efficiency of muscle growth; and, consequently, these compounds are widely used as growth promoters. Although the positive effects of E2 on rate and efficiency of bovine muscle growth are well established, the mechanisms involved in these effects are not well understood. Combined E2 and trenbolone acetate implants result in significantly increased muscle satellite cell number in feedlot steers. Additionally, E2 treatment stimulates proliferation of cultured bovine satellite cells (BSC). Studies in nonmuscle cells have shown that binding of E2 to G protein-coupled estrogen receptor (GPER)-1 results in activation of matrix metalloproteinases 2 and 9 (MMP2/9) resulting in proteolytic release of heparin binding epidermal growth factor-like growth factor (hbEGF) from the cell surface. Released hbEGF binds to and activates the epidermal growth factor receptor resulting in increased proliferation. To assess if GPER-1, MMP2/9, and/or hbEGF are involved in the mechanism of E2-stimulated BSC proliferation, we have examined the effects of G36 (a specific inhibitor of GPER-1), CRM197 (a specific inhibitor of hbEGF), and MMP-2/MMP-9 Inhibitor II (an inhibitor of MMP2/9 activity) on E2-stimulated BSC proliferation. Inhibition of GPER-1, MMP2/9, or hbEGF suppresses E2-stimulated BSC proliferation (P < 0.001) suggesting that all these are required in order for E2 to stimulate BSC proliferation. These results strongly suggest that E2 may stimulate BSC proliferation by binding to GPER-1 resulting in MMP2/9-catalyzed release of cell membrane-bound hbEGF and subsequent activation of epidermal growth factor receptor by binding of released hbEGF. PMID:25010024

  18. Fibroblast Growth Factor2 Is Expressed by the Bovine Uterus and Stimulates Interferon Production in Bovine Trophectoderm

    Microsoft Academic Search

    Donna D. Michael; Idania M. Alvarez; Olga M. Ocon; Anne M. Powell; Neil C. Talbot; Sally E. Johnson; Alan D. Ealy

    2010-01-01

    Uterine-derived factors are essential for conceptus develop- ment and secretion of the maternal recognition-of-pregnancy factor, interferon- (IFNT), in ruminant species. The objec- tives of this study were to determine whether fibroblast growth factor-2 (FGF-2) is expressed in the bovine uterus dur- ing early pregnancy in cattle and to determine whether FGF-2 supplementation affects IFNT mRNA and protein abundance in bovine

  19. Fertility control as a means of controlling bovine tuberculosis in badger (Meles meles) populations in south-west England: predictions from a spatial stochastic simulation model.

    PubMed Central

    White, P C; Lewis, A J; Harris, S

    1997-01-01

    A spatial stochastic simulation model was used to assess the potential of fertility control, based on a yet-to-be-developed oral bait-delivered contraceptive directed at females, for the control of bovine tuberculosis in badger populations in south-west England. The contraceptive had a lifelong effect so that females rendered sterile in any particular year remained so for the rest of their lives. The efficacy of fertility control alone repeated annually for varying periods of time was compared with a single culling operation and integrated control involving an initial single cull followed by annually repeated fertility control. With fertility control alone, in no instance was the disease eradicated completely while a viable badger population (mean group size of at least one individual) was still maintained. Near eradication of the disease (less than 1% prevalence) combined with the survival of a minimum viable badger population was only achieved under a very limited set of conditions, either with high efficiency of control (95%) over a short time period (1-3 years) or a low efficiency of control (20%) over an intermediate time period (10-20 years). Under these conditions, it took more than 20 years for the disease to decline to such low levels. A single cull of 80% efficiency succeeded in near eradication of the disease (below 1% prevalence) after a period of 6-8 years, while still maintaining a viable badger population. Integrated strategies reduced disease prevalence more rapidly and to lower levels than culling alone, although the mean badger group size following the onset of control was smaller. Under certain integrated strategies, principally where a high initial cull (80%) was followed by fertility control over a short (1-3 year) time period, the disease could be completely eradicated while a viable badger population was maintained. However, even under the most favourable conditions of integrated control, it took on average more than 12 years following the onset of control for the disease to disappear completely from the badger population. These results show that whilst fertility control would not be a successful strategy for the control of bovine tuberculosis in badgers if used alone, it could be effective if used with culling as part of an integrated strategy. This type of integrated strategy is likely to be more effective in terms of disease eradication than a strategy employing culling alone. However, the high cost of developing a suitable fertility control agent, combined with the welfare and conservation implications, are significant factors which should be taken into account when considering its possible use as a means of controlling bovine tuberculosis in badger populations in the UK. PMID:9447730

  20. Assessment of Bdellovibrio bacteriovorus 109J killing of Moraxella bovis in an in vitro model of infectious bovine keratoconjunctivitis.

    PubMed

    Boileau, Mélanie J; Clinkenbeard, Kenneth D; Iandolo, John J

    2011-10-01

    The objective of this study was to determine the potential of Bdellovibrio bacteriovorus 109J as an alternative non-chemotherapeutic treatment of infectious bovine keratoconjunctivitis (IBK). To accomplish this, various parameters of B. bacteriovorus predation of Moraxella bovis were determined in vitro. Initial passage of B. bacteriovorus using M. bovis as prey required 10 d for active cultures to develop compared with 2 d for culture on normal Escherichia coli prey; however by the 5th passage, time to active predatory morphology was reduced to 2 d. This high passage B. bacteriovorus culture [1 × 10(10) plaque forming units (PFU)/mL] killed 76% of M. bovis [1 × 10(7) colony forming units (CFU)/mL] present in suspension broth in a 4 h assay. The minimal level of M. bovis supporting B. bacteriovorus predation was 1 × 10(4) CFU/mL. To assess the ability of B. bacteriovorus to kill M. bovis on an epithelial surface mimicking IBK, an in vitro assay with Madin-Darby bovine kidney (MDBK) cells inoculated with 4 × 10(7) CFU/mL M. bovis was used. Treatment with a B. bacteriovorus suspension (1.6 × 10(11) PFU/mL) decreased adherence of M. bovis to MDBK cells by 6-fold at 12 h of treatment, as well as decreased the number of unattached M. bovis cells by 1.4-fold. This study demonstrates that B. bacteriovorus has potential as an effective biological control of M. bovis at levels likely present in IBK-infected corneal epithelia and ocular secretions. PMID:22468026

  1. MicroRNA regulation of bovine monocyte inflammatory and metabolic networks in an in vivo infection model.

    PubMed

    Lawless, Nathan; Reinhardt, Timothy A; Bryan, Kenneth; Baker, Mike; Pesch, Bruce; Zimmerman, Duane; Zuelke, Kurt; Sonstegard, Tad; O'Farrelly, Cliona; Lippolis, John D; Lynn, David J

    2014-06-01

    Bovine mastitis is an inflammation-driven disease of the bovine mammary gland that costs the global dairy industry several billion dollars per year. Because disease susceptibility is a multifactorial complex phenotype, an integrative biology approach is required to dissect the molecular networks involved. Here, we report such an approach using next-generation sequencing combined with advanced network and pathway biology methods to simultaneously profile mRNA and miRNA expression at multiple time points (0, 12, 24, 36 and 48 hr) in milk and blood FACS-isolated CD14(+) monocytes from animals infected in vivo with Streptococcus uberis. More than 3700 differentially expressed (DE) genes were identified in milk-isolated monocytes (MIMs), a key immune cell recruited to the site of infection during mastitis. Upregulated genes were significantly enriched for inflammatory pathways, whereas downregulated genes were enriched for nonglycolytic metabolic pathways. Monocyte transcriptional changes in the blood, however, were more subtle but highlighted the impact of this infection systemically. Genes upregulated in blood-isolated monocytes (BIMs) showed a significant association with interferon and chemokine signaling. Furthermore, 26 miRNAs were DE in MIMs and three were DE in BIMs. Pathway analysis revealed that predicted targets of downregulated miRNAs were highly enriched for roles in innate immunity (FDR < 3.4E-8), particularly TLR signaling, whereas upregulated miRNAs preferentially targeted genes involved in metabolism. We conclude that during S. uberis infection miRNAs are key amplifiers of monocyte inflammatory response networks and repressors of several metabolic pathways. PMID:24470219

  2. CHARACTERIZATION OF INTERACTIONS WITH MODEL PROTEINS

    E-print Network

    Lebendiker, Mario

    in PBS, pH7.4 on PPA HyperCel sorbent. Elution at pH 5.0, 4.0 and 2.6. Analysis by ELISA (test developed industrially relevant proteins. Their unique selectivity, illustrated by the separation of protein or peptide

  3. Homology-Based Modeling of Protein Structure

    NASA Astrophysics Data System (ADS)

    Xiang, Zhexin

    The human genome project has already discovered millions of proteins (http://www.swissprot.com). The potential of the genome project can only be fully realized once we can assign, understand, manipulate, and predict the function of these new proteins (Sanchez and Sali, 1997; Frishman et al., 2000; Domingues et al., 2000). Predicting protein function generally requires knowledge of protein three-dimensional structure (Blundell et al., 1978;Weber, 1990), which is ultimately determined by protein sequence (Anfinsen, 1973). Protein structure determination using experimental methods such as X-ray crystallography or NMR spectroscopy is very time consuming (Johnson et al. 1994). To date, fewer than 2% of the known proteins have had their structures solved experimentally. In 2004, more than half a million new proteins were sequenced that almost doubled the efforts in the previous year, but only 5300 structures were solved. Although the rate of experimental structure determination will continue to increase, the number of newly discovered sequences grows much faster than the number of structures solved (see Fig. 10.1).

  4. High-order fluorescence fluctuation analysis of model protein clusters.

    PubMed Central

    Palmer, A G; Thompson, N L

    1989-01-01

    The technique of high-order fluorescence fluctuation autocorrelation for detecting and characterizing protein oligomers was applied to solutions containing two fluorescent proteins in which the more fluorescent proteins were analogues for clusters of the less fluorescent ones. The results show that the model protein clusters can be detected for average numbers of observed subunits (free monomers plus monomers in oligomers) equal to 10-100 and for relative fluorescent yields that correspond to oligomers as small as trimers. High-order fluorescent fluctuation analysis may therefore be applicable to cell surface receptor clusters in natural or model membranes. PMID:2548201

  5. Global motions exhibited by proteins in micro- to milliseconds simulations concur with anisotropic network model predictions

    NASA Astrophysics Data System (ADS)

    Gur, M.; Zomot, E.; Bahar, I.

    2013-09-01

    The Anton supercomputing technology recently developed for efficient molecular dynamics simulations permits us to examine micro- to milli-second events at full atomic resolution for proteins in explicit water and lipid bilayer. It also permits us to investigate to what extent the collective motions predicted by network models (that have found broad use in molecular biophysics) agree with those exhibited by full-atomic long simulations. The present study focuses on Anton trajectories generated for two systems: the bovine pancreatic trypsin inhibitor, and an archaeal aspartate transporter, GltPh. The former, a thoroughly studied system, helps benchmark the method of comparative analysis, and the latter provides new insights into the mechanism of function of glutamate transporters. The principal modes of motion derived from both simulations closely overlap with those predicted for each system by the anisotropic network model (ANM). Notably, the ANM modes define the collective mechanisms, or the pathways on conformational energy landscape, that underlie the passage between the crystal structure and substates visited in simulations. In particular, the lowest frequency ANM modes facilitate the conversion between the most probable substates, lending support to the view that easy access to functional substates is a robust determinant of evolutionarily selected native contact topology.

  6. trans activation by the full-length E2 proteins of human papillomavirus type 16 and bovine papillomavirus type 1 in vitro and in vivo: cooperation with activation domains of cellular transcription factors.

    PubMed Central

    Ushikai, M; Lace, M J; Yamakawa, Y; Kono, M; Anson, J; Ishiji, T; Parkkinen, S; Wicker, N; Valentine, M E; Davidson, I

    1994-01-01

    Papillomaviral E2 genes encode proteins that regulate viral transcription. While the full-length bovine papillomavirus type 1 (BPV-1) E2 peptide is a strong trans activator, the homologous full-length E2 product of human papillomavirus type 16 (HPV-16) appeared to vary in function in previous studies. Here we show that when expressed from comparable constructs, the full-length E2 products of HPV-16 and BPV-1 trans activate a simple E2- and Sp1-dependent promoter up to approximately 100-fold in human keratinocytes and other epithelial cells as well as human and animal fibroblasts. Vaccinia virus-expressed, purified full-length HPV-16 and BPV-1 E2 proteins bound a consensus E2 site with high specific affinities (Kd = approximately 10(-9) M) and stimulated in vitro transcription up to six- to eightfold. In vivo and in vitro trans activation by either E2 protein required cooperation with another activator, such as Sp1, or other factors that interact with papillomavirus promoters, such as AP-1, Oct-1, nuclear factor 1/CTF, transcriptional enhancer factor 1, or USF. The glutamine-rich domain B of Sp1 or the mutually unrelated activation domains of other transcription factors were necessary and sufficient for cooperation with either E2 factor. We conclude that like BPV-1 E2, the HPV-16 E2 protein has the potential to function as a strong activator of viral gene expression in cooperation with cellular transcription factors. Images PMID:8083999

  7. Infection of Calves with Bovine Norovirus GIII.1 Strain Jena Virus: an Experimental Model To Study the Pathogenesis of Norovirus Infection ?

    PubMed Central

    Otto, Peter H.; Clarke, Ian N.; Lambden, Paul R.; Salim, Omar; Reetz, Jochen; Liebler-Tenorio, Elisabeth M.

    2011-01-01

    The experimental infection of newborn calves with bovine norovirus was used as a homologous large animal model to study the pathogenesis of norovirus infection and to determine target cells for viral replication. Six newborn calves were inoculated orally with Jena virus (JV), a bovine norovirus GIII.1 strain, and six calves served as mock-inoculated controls. Following infection, calves were euthanized before the onset of diarrhea (12 h postinoculation [hpi]), shortly after the onset of diarrhea (18 to 21 hpi), and postconvalescence (4 days pi [dpi]). Calves inoculated with JV developed severe watery diarrhea at 14 to 16 hpi, and this symptom lasted for 53.5 to 67.0 h. Intestinal lesions were characterized by severe villus atrophy together with loss and attenuation of villus epithelium. Viral capsid antigen (JV antigen) was detected by immunohistochemistry in the cytoplasm of epithelial cells on villi. In addition, granular material positive for JV antigen was detected in the lamina propria of villi. Lesions first appeared at 12 hpi and were most extensive at 18 to 19 hpi, extending from midjejunum to ileum. The intestinal mucosa had completely recovered at 4 dpi. There was no indication of systemic infection as described for norovirus infection in mice. JV was found in intestinal contents by reverse transcription-PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) as early as 12 hpi. Fecal shedding of the virus started at 13 hpi and stopped at 23 hpi or at necropsy (4 dpi), respectively. Throughout the trial, none of the control calves tested positive for JV by ELISA or RT-PCR. PMID:21880760

  8. From Genomes to Protein Models and Back

    NASA Astrophysics Data System (ADS)

    Tramontano, Anna; Giorgetti, Alejandro; Orsini, Massimiliano; Raimondo, Domenico

    2007-12-01

    The alternative splicing mechanism allows genes to generate more than one product. When the splicing events occur within protein coding regions they can modify the biological function of the protein. Alternative splicing has been suggested as one way for explaining the discrepancy between the number of human genes and functional complexity. We analysed the putative structure of the alternatively spliced gene products annotated in the ENCODE pilot project and discovered that many of the potential alternative gene products will be unlikely to produce stable functional proteins.

  9. Prespermatogenesis and spermatogoniogenesis in the bovine testis

    Microsoft Academic Search

    Karl-Heinz Wrobel

    2000-01-01

    The bovine male germ cell population was studied over the entire period from testicular differentiation in the embryo through\\u000a onset of spermatogenesis in the pubertal calf. Germ cells were identified by protein gene product 9.5 immunohistochemistry\\u000a and characterized by their ultrastructure. The proliferation pattern of germ cells was studied with immunohistochemical anti-Ki\\u000a 67 and anti-proliferating cell nuclear antigen reactions. Germ

  10. Developing algorithms for predicting protein-protein interactions of homology modeled proteins.

    SciTech Connect

    Martin, Shawn Bryan; Sale, Kenneth L.; Faulon, Jean-Loup Michel; Roe, Diana C.

    2006-01-01

    The goal of this project was to examine the protein-protein docking problem, especially as it relates to homology-based structures, identify the key bottlenecks in current software tools, and evaluate and prototype new algorithms that may be developed to improve these bottlenecks. This report describes the current challenges in the protein-protein docking problem: correctly predicting the binding site for the protein-protein interaction and correctly placing the sidechains. Two different and complementary approaches are taken that can help with the protein-protein docking problem. The first approach is to predict interaction sites prior to docking, and uses bioinformatics studies of protein-protein interactions to predict theses interaction site. The second approach is to improve validation of predicted complexes after docking, and uses an improved scoring function for evaluating proposed docked poses, incorporating a solvation term. This scoring function demonstrates significant improvement over current state-of-the art functions. Initial studies on both these approaches are promising, and argue for full development of these algorithms.

  11. Mutational robustness emerges in a microscopic model of protein evolution

    NASA Astrophysics Data System (ADS)

    Zeldovich, Konstantin; Shakhnovich, Eugene

    2009-03-01

    The ability to absorb mutations while retaining structure and function, or mutational robustness, is a remarkable property of natural proteins. We use a computational model of organismic evolution [Zeldovich et al, PLOS Comp Biol 3(7):e139 (2007)], which explicitly couples protein physics and population dynamics, to study mutational robustness of evolved model proteins. We compare evolved sequences with the ones designed to fold into the same native structures and having the same thermodynamic stability, and find that evolved sequences are more robust against point mutations, being less likely to be destabilized, and more likely to increase stability upon a point mutation. These results point to sequence evolution as an important method of protein engineering if mutational robustness of the artificially developed proteins is desired. On the biological side, mutational robustness of proteins appears to be a natural consequence of the divergence-mutation- selection evolutionary process.

  12. Inferring functional modules of protein families with probabilistic topic models

    PubMed Central

    2011-01-01

    Background Genome and metagenome studies have identified thousands of protein families whose functions are poorly understood and for which techniques for functional characterization provide only partial information. For such proteins, the genome context can give further information about their functional context. Results We describe a Bayesian method, based on a probabilistic topic model, which directly identifies functional modules of protein families. The method explores the co-occurrence patterns of protein families across a collection of sequence samples to infer a probabilistic model of arbitrarily-sized functional modules. Conclusions We show that our method identifies protein modules - some of which correspond to well-known biological processes - that are tightly interconnected with known functional interactions and are different from the interactions identified by pairwise co-occurrence. The modules are not specific to any given organism and may combine different realizations of a protein complex or pathway within different taxa. PMID:21554720

  13. Ensemble modeling of [beta]-sheet proteins

    E-print Network

    O'Donnell, Charles William

    2011-01-01

    Our ability to characterize protein structure and dynamics is vastly outpaced by the speed of modern genetic sequencing, creating a growing divide between our knowledge of biological sequence and structure. Structural ...

  14. Functional clustering of immunoglobulin superfamily proteins with protein-protein interaction information calibrated Hidden Markov model sequence-profiles

    PubMed Central

    Yap, Eng-Hui; Rosche, Tyler; Almo, Steve; Fiser, Andras

    2014-01-01

    Secreted and cell surface-localized members of the immunoglobulin superfamily (IgSF) play central roles in regulating adaptive and innate immune responses, and are prime targets for the development of protein-based therapeutics. An essential activity of the ectodomains of these proteins is the specific recognition of cognate ligands, which are often other members of the IgSF. In this work we provide functional insight for this important class of proteins through the development of a clustering algorithm that groups together extracellular domains of the IgSF with similar binding preferences. Information from hidden Markov model-based sequence profiles and domain structure is calibrated against manually curated protein interaction data to define functional families of IgSF proteins. The method is able to assign 82% of the 477 extracellular IgSF protein to a functional family, while the rest are either single proteins with unique function or proteins that could not be assigned with the current technology. The functional clustering of IgSF proteins generates hypotheses regarding the identification of new cognate receptor:ligand pairs and reduces the pool of possible interacting partners to a manageable level for experimental validation. PMID:24246499

  15. Time-resolved small-angle neutron scattering during heat-induced fibril formation from bovine ?-lactoglobulin

    NASA Astrophysics Data System (ADS)

    Arnaudov, Luben N.; de Vries, Renko; Cohen Stuart, Martien A.

    2006-02-01

    We study in situ the kinetics of heat-induced fibrilar aggregation of bovine ?-lactoglobulin at pH 2.0 and 80°C for the first time by time-resolved small-angle neutron scattering. A simple model for the scattering from a mixture of monodisperse charged spheres (monomeric ?-lactoglobulin) interacting via a screened electrostatic repulsion and noninteracting long cylinders (protein fibrils) is used to describe the data. The experimental data are fitted to the model and the concentration of the monomeric protein and the protein incorporated in fibrils are obtained as adjustable parameters. Thus, a simple physical model allows the determination of realistic kinetic parameters during fibrilar protein aggregation. This result constitutes an important step in understanding the process of irreversible fibrilar aggregation of proteins.

  16. Immunogenicity of Therapeutic Proteins: The Use of Animal Models

    Microsoft Academic Search

    Vera Brinks; Wim Jiskoot; Huub Schellekens

    Immunogenicity of therapeutic proteins lowers patient well-being and drastically increases therapeutic costs. Preventing immunogenicity\\u000a is an important issue to consider when developing novel therapeutic proteins and applying them in the clinic. Animal models\\u000a are increasingly used to study immunogenicity of therapeutic proteins. They are employed as predictive tools to assess different\\u000a aspects of immunogenicity during drug development and have become

  17. Raw bovine milk improves gut responses to feeding relative to infant formula in preterm piglets.

    PubMed

    Li, Yanqi; Jensen, Mikkel L; Chatterton, Dereck E W; Jensen, Bent B; Thymann, Thomas; Kvistgaard, Anne S; Sangild, Per T

    2014-01-01

    For preterm neonates, the quality of the first milk is crucial for intestinal maturation and resistance to necrotizing enterocolitis (NEC). Among other factors, milk quality is determined by the stage of lactation and processing. We hypothesized that unprocessed mature bovine milk (BM; raw bovine milk) would have less bioactivity than corresponding bovine colostrum (BC) in a preterm pig model, but have improved bioactivity relative to its homogenized, pasteurized, spray-dried equivalent, whole milk powder (WMP), or a bovine milk protein-based infant formula (IF). For 5 days, newborn preterm pigs received parenteral and enteral nutrition consisting of IF (n = 13), BM (n = 13), or BC (n = 14). In a second study, WMP (n = 15) was compared with IF (n = 10) and BM (n = 9). Compared with pigs fed IF, pigs that were fed BM had significantly improved intestinal structure (mucosal weight, villus height) and function (increased nutrient absorption and enzyme activities, decreased gut permeability, nutrient fermentation, and NEC severity). BC further improved these effects relative to BM (lactase activity, lactose absorption, plasma citrulline, and tissue interleukin-8). WMP induced similar effects as BM, except for lactase activity and lactose absorption. In conclusion, the maturational and protective effects on the immature intestine decreased in the order BC>BM>WMP, but all three intact bovine milk diets were markedly better than IF. The stage of lactation (colostrum vs. mature milk) and milk processing (e.g., homogenization, fractionation, pasteurization, spray-drying) are important factors in determining milk quality during the early postnatal period of preterm neonates. PMID:24157971

  18. Bovine Milk Proteome in the First 9 Days: Protein Interactions in Maturation of the Immune and Digestive System of the Newborn

    PubMed Central

    Zhang, Lina; Boeren, Sjef; Hageman, Jos A.; van Hooijdonk, Toon; Vervoort, Jacques; Hettinga, Kasper

    2015-01-01

    In order to better understand the milk proteome and its changes from colostrum to mature milk, samples taken at seven time points in the first 9 days from 4 individual cows were analyzed using proteomic techniques. Both the similarity in changes from day 0 to day 9 in the quantitative milk proteome, and the differences in specific protein abundance, were observed among four cows. One third of the quantified proteins showed a significant decrease in concentration over the first 9 days after calving, especially in the immune proteins (as much as 40 fold). Three relative high abundant enzymes (XDH, LPL, and RNASE1) and cell division and proliferation protein (CREG1) may be involved in the maturation of the gastro-intestinal tract. In addition, high correlations between proteins involved in complement and blood coagulation cascades illustrates the complex nature of biological interrelationships between milk proteins. The linear decrease of protease inhibitors and proteins involved in innate and adaptive immune system implies a protective role for protease inhibitor against degradation. In conclusion, the results found in this study not only improve our understanding of the role of colostrum in both host defense and development of the newborn calf but also provides guidance for the improvement of infant formula through better understanding of the complex interactions between milk proteins. PMID:25693162

  19. Modeling of protein side-chain conformations with RASP.

    PubMed

    Miao, Zhichao; Cao, Yang; Jiang, Taijiao

    2014-01-01

    Modeling of side-chain conformations on a fixed protein backbone, also called side-chain packing, plays an important role in protein structure prediction, protein design, molecular docking, and functional analysis. RASP, or RApid Side-chain Predictor, is a recently developed program that can model protein side-chain conformations with both high accuracy and high speed. Moreover, it can generate structures with few atomic clashes. This chapter first provides a brief introduction to the principle and performances of the RASP package. Then details on how to use RASP programs to predict protein side-chain conformations are elaborated. Finally, it describes case studies for structure refinement in homology modeling and residue substitution. PMID:24573473

  20. Weak self-interactions of globular proteins studied by small-angle X-ray scattering and structure-based modeling.

    PubMed

    Kaieda, Shuji; Lund, Mikael; Plivelic, Tomás S; Halle, Bertil

    2014-08-28

    We investigate protein-protein interactions in solution by small-angle X-ray scattering (SAXS) and theoretical modeling. The structure factor for solutions of bovine pancreatic trypsin inhibitor (BPTI), myoglobin (Mb), and intestinal fatty acid-binding protein (IFABP) is determined from SAXS measurements at multiple concentrations, from Monte Carlo simulations with a coarse-grained structure-based interaction model, and from analytic approximate solutions of two idealized colloidal interaction models without adjustable parameters. By combining these approaches, we find that the structure factor is essentially determined by hard-core and screened electrostatic interactions. Other soft short-ranged interactions (van der Waals and solvation-related) are either individually insignificant or tend to cancel out. The structure factor is also not significantly affected by charge fluctuations. For Mb and IFABP, with a small net charge and relatively symmetric charge distribution, the structure factor is well described by a hard-sphere model. For BPTI, with a larger net charge, screened electrostatic repulsion is also important, but the asymmetry of the charge distribution reduces the repulsion from that predicted by a charged hard-sphere model with the same net charge. Such charge asymmetry may also amplify the effect of shape asymmetry on the protein-protein potential of mean force. PMID:25117055

  1. Modelling the human rhesus proteins: implications for structure and function

    E-print Network

    Merrick, Mike

    Modelling the human rhesus proteins: implications for structure and function The human erythrocyte (Mouro et al, 1993). Humans express two other glycosylated non-erythroid Rh proteins, RhBG and Rh in erythrocytes (Ripoche et al, 2004), in yeast (Marini et al, 2000; Westhoff et al, 2004) and Xenopus oocytes

  2. Bayesian hierarchical reconstruction of protein profiles including a digestion model

    E-print Network

    Paris-Sud XI, Université de

    Bayesian hierarchical reconstruction of protein profiles including a digestion model Pierre to recover the protein biomarkers content in a robust way. We will focus on the digestion step since and each branch to a molecular processing such as digestion, ionisation and LC-MS separation

  3. Bayesian Model of Protein Primary Sequence for Secondary Structure Prediction

    E-print Network

    Vannucci, Marina

    improving secondary structure reduction given the primary structure, we propose a Bayesian model based made obtaining protein sequences relatively cheap, accurate and fast, in comparison to the costly design of protein structure [4] and enzymatic function [5] as well as in drug development [6]. Depending

  4. Coarse-Grained Model for Colloidal Protein Interactions, B22, and Protein Cluster Formation

    PubMed Central

    Blanco, Marco A.; Sahin, Eric; Robinson, Anne S.; Roberts, Christopher J.

    2014-01-01

    Reversible protein cluster formation is an important initial step in the processes of native and non-native protein aggregation, but involves relatively long time and length scales for detailed atomistic simulations and extensive mapping of free energy landscapes. A coarse-grained (CG) model is presented to semi-quantitatively characterize the thermodynamics and key configurations involved in the landscape for protein oligomerization, as well as experimental measures of interactions such as the osmotic second virial coefficient (B22). Based on earlier work, this CG model treats proteins as rigid bodies composed of one bead per amino acid, with each amino acid having specific parameters for its size, hydrophobicity, and charge. The net interactions are a combination of steric repulsions, short-range attractions, and screened long-range charge-charge interactions. Model parametrization was done by fitting simulation results against experimental values of the B22 as a function of solution ionic strength for ?-chymotrypsinogen A and ?D-crystallin (gD-Crys). The CG model is applied to characterize the pairwise interactions and dimerization of gD-Crys and the dependance on temperature, protein concentration, and ionic strength. The results illustrate that at experimentally relevant conditions where stable dimers do not form, the entropic contributions are predominant in the free-energy of protein cluster formation and colloidal protein interactions, arguing against interpretations that treat B22 primarily from energetic considerations alone. Additionally, the results suggest that electrostatic interactions help to modulate the population of the different stable configurations for protein nearest-neighbor pairs, while short-range attractions determine the relative orientations of proteins within these configurations. Finally, simulation results are combined with Principal Component Analysis to identify those amino-acids / surface patches that form inter-protein contacts at conditions that favor dimerization of gD-Crys. The resulting regions agree with previously found aggregation-prone sites, as well as suggesting new ones that may be important. PMID:24289039

  5. A physical map of the bovine genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background Cattle are important agriculturally and relevant as a model organism. Previously described genetic and radiation hybrid (RH) maps of the bovine genome have been used to identify genomic regions and genes affecting specific traits. Application of these maps to identify influential geneti...

  6. Bovine Mandibular Fractures

    PubMed Central

    Trent, A. M.; Ferguson, J. G.

    1985-01-01

    A retrospective study of bovine mandibular fractures was conducted. An increased incidence in males, beef breeds and animals less than one year of age was identified when the study group of 17 was compared to the total bovine case load. Manipulation during dystocia was the most common cause of fractures. Four neonates had rostral mandibular fractures. Fractures caudal to or involving the premolars were restricted to animals over a year of age. Treatment was attempted in ten of the seventeen cases, with euthanasia or slaughter elected in the remaining seven cases. Of the five cases treated by internal fixation, all four neonates died from conditions related to septicemia. PMID:17422602

  7. Molecular modeling of protein materials: case study of elastin

    NASA Astrophysics Data System (ADS)

    Tarakanova, Anna; Buehler, Markus J.

    2013-09-01

    Molecular modeling of protein materials is a quickly growing area of research that has produced numerous contributions in fields ranging from structural engineering to medicine and biology. We review here the history and methods commonly employed in molecular modeling of protein materials, emphasizing the advantages for using modeling as a complement to experimental work. We then consider a case study of the protein elastin, a critically important ‘mechanical protein’ to exemplify the approach in an area where molecular modeling has made a significant impact. We outline the progression of computational modeling studies that have considerably enhanced our understanding of this important protein which endows elasticity and recoil to the tissues it is found in, including the skin, lungs, arteries and the heart. A vast collection of literature has been directed at studying the structure and function of this protein for over half a century, the first molecular dynamics study of elastin being reported in the 1980s. We review the pivotal computational works that have considerably enhanced our fundamental understanding of elastin's atomistic structure and its extraordinary qualities—focusing on two in particular: elastin's superb elasticity and the inverse temperature transition—the remarkable ability of elastin to take on a more structured conformation at higher temperatures, suggesting its effectiveness as a biomolecular switch. Our hope is to showcase these methods as both complementary and enriching to experimental approaches that have thus far dominated the study of most protein-based materials.

  8. Downstream Processing of Recombinant Proteins from Transgenic Plant Systems: Phenolic Compounds Removal from Monoclonal Antibody Expressing Lemna minor and Purification of Recombinant Bovine Lysozyme from Sugarcane 

    E-print Network

    Barros, Georgia

    2012-07-16

    Transgenic plant systems have been proposed as bioreactors in the production of pharmaceutical and industrial proteins. The economic benefits of inexpensive plant production systems could be erased if the downstream ...

  9. Bayesian statistical modelling of human protein interaction network incorporating protein disorder information

    PubMed Central

    2010-01-01

    Background We present a statistical method of analysis of biological networks based on the exponential random graph model, namely p2-model, as opposed to previous descriptive approaches. The model is capable to capture generic and structural properties of a network as emergent from local interdependencies and uses a limited number of parameters. Here, we consider one global parameter capturing the density of edges in the network, and local parameters representing each node's contribution to the formation of edges in the network. The modelling suggests a novel definition of important nodes in the network, namely social, as revealed based on the local sociality parameters of the model. Moreover, the sociality parameters help to reveal organizational principles of the network. An inherent advantage of our approach is the possibility of hypotheses testing: a priori knowledge about biological properties of the nodes can be incorporated into the statistical model to investigate its influence on the structure of the network. Results We applied the statistical modelling to the human protein interaction network obtained with Y2H experiments. Bayesian approach for the estimation of the parameters was employed. We deduced social proteins, essential for the formation of the network, while incorporating into the model information on protein disorder. Intrinsically disordered are proteins which lack a well-defined three-dimensional structure under physiological conditions. We predicted the fold group (ordered or disordered) of proteins in the network from their primary sequences. The network analysis indicated that protein disorder has a positive effect on the connectivity of proteins in the network, but do not fully explains the interactivity. Conclusions The approach opens a perspective to study effects of biological properties of individual entities on the structure of biological networks. PMID:20100321

  10. A mesoscopic model for protein enzymatic dynamics in solution.

    PubMed

    Echeverria, Carlos; Togashi, Yuichi; Mikhailov, Alexander S; Kapral, Raymond

    2011-06-14

    A multi-scale, coarse-grained description of protein conformational dynamics in a solvent is presented. The focus of the paper is on the description of the conformational motions that may accompany enzyme catalysis as the enzyme executes a catalytic cycle, starting with substrate binding and ending with product release and return to the original unbound enzyme. The protein is modeled by a network of beads representing amino acid residues, the solvent is described by multiparticle collision dynamics, and substrate binding and unbinding events are modeled stochastically by conformation-dependent transitions that modify the bonding in the network to correspond to the different binding states of the protein. The solvent dynamics is coupled to that of the protein and hydrodynamic interactions, which are important for the large-scale protein motions, are taken into account. The multi-scale model is used to study the dynamics of the adenylate kinase enzyme in solution. A potential function that describes the different binding and conformational states of the protein and accounts for partial unfolding during the catalytic cycle is constructed as a network built from elastic network and soft potential links. The conformational dynamics of the protein as it undergoes cyclic enzymatic dynamics, as well as its translational diffusion and orientational motion, are investigated using both multiparticle collision dynamics and dynamics that suppresses hydrodynamic coupling. Hydrodynamic interactions are found to have important effects on the large scale conformational motions of the protein and significantly affect the translational diffusion coefficients and orientational correlation times. PMID:21442113

  11. Scoring predictive models using a reduced representation of proteins: model and energy definition

    Microsoft Academic Search

    Federico Fogolari; Lidia Pieri; Agostino Dovier; Luca Bortolussi; Gilberto Giugliarelli; Alessandra Corazza; Gennaro Esposito; Paolo Viglino

    2007-01-01

    BACKGROUND: Reduced representations of proteins have been playing a keyrole in the study of protein folding. Many such models are available, with different representation detail. Although the usefulness of many such models for structural bioinformatics applications has been demonstrated in recent years, there are few intermediate resolution models endowed with an energy model capable, for instance, of detecting native or

  12. Local protein structure prediction using discriminative models

    Microsoft Academic Search

    Oliver Sander; Ingolf Sommer; Thomas Lengauer

    2006-01-01

    Background: In recent years protein structure prediction methods using local structure information have shown promising improvements. The quality of new fold predictions has risen significantly and in fold recognition incorporation of local structure predictions led to improvements in the accuracy of results. We developed a local structure prediction method to be integrated into either fold recognition or new fold prediction

  13. ANIMAL MODELS FOR PROTEIN RESPIRATORY SENSITIZERS

    EPA Science Inventory

    Protein induced respiratory hypersensitivity, particularly atopic disease in general, and allergic asthma in particular, has increased dramatically over the last several decades in the U.S. and other industrialized nations as a result of ill-defined changes in living conditions i...

  14. Predicting protein structure using hidden Markov models Kevin Karplusy

    E-print Network

    Karplus, Kevin

    Predicting protein structure using hidden Markov models Kevin Karplusy Computer Engineering U, Supplement 1, 1997 Abstract We discuss how methods based on hidden Markov models performed in the fold-recognition section of the CASP2 experiment. Hidden Markov models were built for a representative set of just over one

  15. A model of sequence-dependent protein diffusion along DNA.

    PubMed

    Barbi, Maria; Place, Christophe; Popkov, Vladislav; Salerno, Mario

    2004-09-01

    We introduce a probabilistic model for protein sliding motion along DNA during the search of a target sequence. The model accounts for possible effects due to sequence-dependent interaction between the nonspecific DNA and the protein. Hydrogen bonds formed at the target site are used as the main sequence-dependent interaction between protein and DNA. The resulting dynamical properties and the possibility of an experimental verification are discussed in details. We show that, while at large times the process reaches a linear diffusion regime, it initially displays a sub-diffusive behavior. The sub-diffusive regime can last sufficiently long to be of biological interest. PMID:23345869

  16. Bovine Spongiform Encephalopathy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine spongiform encephalopathy (BSE), also referred to as “mad cow disease” is a chronic, non-febrile, neuro-degenerative disease affecting the central nervous system. The transmissible spongiform encephalopathies (TSEs) of domestic animals, of which BSE is a member includes scrapie of sheep...

  17. BOVINE VIRAL DIARRHEA VIRUSES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine viral diarrhea virus (BVDV) is an umbrella term for two species of viruses, BVDV1 and BVDV2, within the Pestivirus genus of the Flavivirus family. BVDV viruses are further subclassified as cytopathic and noncytopathic based on their activity in cultured epithelial cells. Noncytopathic BVDV p...

  18. BRUCELLOSE BOVINE EXPRIMENTALE XII. —

    E-print Network

    Paris-Sud XI, Université de

    BRUCELLOSE BOVINE EXPÉRIMENTALE XII. — PERSISTANCE A L'ÂGE ADULTE DE L'INFECTION CONGÉNITALE Brucella par culture a été pratiquée dans le mucus vaginal avant et après le part, dans la sécrétion, analogue à la souche 544, à partir du mucus vaginal, de la sécrétion mammaire, ante et post pavtum, et des

  19. Bovine besnoitiosis in Germany.

    PubMed

    Rostaher, Ana; Mueller, Ralf S; Majzoub, Monir; Schares, Gereon; Gollnick, Nicole S

    2010-08-01

    This paper reports a case of natural occurring bovine besnoitiosis in Germany. The skin lesions consisted of multifocal hypotrichosis and alopecia, lichenification, erythema and seborrhoea. Histopathologic findings revealed characteristic cysts of Besnoitia spp. The diagnosis was confirmed by serology and the species Besnoitia besnoiti was identified by polymerase chain reaction (PCR). PMID:20230585

  20. DIAGNOSIS OF BOVINE NEOSPOROSIS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The protozoan parasite Neospora caninum is a major cause of abortion in cattle. The diagnosis of neosporosis-associated mortality and abortion in cattle is difficult. In the present papers we review histologic, serologic, immunohistochemical, and molecular methods for dignosis of bovine neosporosis....

  1. Genotyping bovine coronaviruses.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine coronaviruses (BoCV) are enveloped, single-stranded, positive-sense RNA viruses of the Coronaviridae family. Infection is associated with enteritis and pneumonia in calves and Winter Dysentery in adult cattle. Strains, isolated more than 50 years ago, are used in vaccines and as laboratory ...

  2. A Computational Model of the LGI1 Protein Suggests a Common Binding Site for ADAM Proteins

    PubMed Central

    Leonardi, Emanuela; Andreazza, Simonetta; Vanin, Stefano; Busolin, Giorgia; Nobile, Carlo; Tosatto, Silvio C. E.

    2011-01-01

    Mutations of human leucine-rich glioma inactivated (LGI1) gene encoding the epitempin protein cause autosomal dominant temporal lateral epilepsy (ADTLE), a rare familial partial epileptic syndrome. The LGI1 gene seems to have a role on the transmission of neuronal messages but the exact molecular mechanism remains unclear. In contrast to other genes involved in epileptic disorders, epitempin shows no homology with known ion channel genes but contains two domains, composed of repeated structural units, known to mediate protein-protein interactions. A three dimensional in silico model of the two epitempin domains was built to predict the structure-function relationship and propose a functional model integrating previous experimental findings. Conserved and electrostatic charged regions of the model surface suggest a possible arrangement between the two domains and identifies a possible ADAM protein binding site in the ?-propeller domain and another protein binding site in the leucine-rich repeat domain. The functional model indicates that epitempin could mediate the interaction between proteins localized to different synaptic sides in a static way, by forming a dimer, or in a dynamic way, by binding proteins at different times. The model was also used to predict effects of known disease-causing missense mutations. Most of the variants are predicted to alter protein folding while several other map to functional surface regions. In agreement with experimental evidence, this suggests that non-secreted LGI1 mutants could be retained within the cell by quality control mechanisms or by altering interactions required for the secretion process. PMID:21479274

  3. Immunogenicity of synthetic peptide sequences S81 and S82 (residues 68-83 and 65-83) of bovine myelin basic protein. Time-course of antibody responses in rats and rabbits.

    PubMed

    Day, E D; Hashim, G A; Varitek, V A; Paterson, P Y

    1981-06-01

    The timing and intensity of the antibody responses to the marker determinants of synthetic peptide S81 and S82 sequences of bovine myelin basic protein (MBP) (residues 68-83 and 65-83, respectively) were studied in 20 Lewis rats and 6 rabbits. All rats immunized with either peptide in CFA responded with antibody development. All rabbits immunized with S82 and CFA developed both antibodies and experimental allergic encephalomyelitis. In contrast only one rabbit developed antibodies against S81 and none of the S81-challenged rabbits developed disease. On the basis of extrapolation of linear time-response curves to zero activity, the time of appearance of anti-peptide antibody activity in the Lewis rats was 15.1 +/- 1.7 days after a single immunization, a week longer than the normal latent period before appearance of anti-MBP antibodies. The time of appearance of anti-S82 antibody activity in rabbits exhibiting linear response curves was 18 days, 4 days after a booster immunization with S82 in incomplete Freund's adjuvant. The development of clinical signs of experimental allergic encephalomyelitis occurred within 4 weeks after initial challenge (a few days after boosting) and continued for 8--13 days in all S82-immunized rabbits. PMID:6175660

  4. Expression levels of mRNA for insulin-like growth factors 1 and 2, IGF receptors and IGF binding proteins in in vivo and in vitro grown bovine follicles.

    PubMed

    Rebouças, Emanuela L; Costa, José J N; Passos, Maria J; Silva, Anderson W B; Rossi, Rodrigo O D S; van den Hurk, Robert; Silva, José R V

    2014-11-01

    This study investigated mRNA levels for insulin-like growth factors (IGFs) IGF1 (IGF-I) and IGF2 (IGF-II), IGF receptors (IGF1R and IGF2R), and binding proteins (IGFBP-1, IGFBP-2. IGFBP-3, IGFBP-4, IGFBP-5 and IGFBP-6) in bovine follicles of 0.2, 0.5 or 1.0 mm in diameter. mRNA expression levels in in vitro cultured follicles that reached approximately 0.5 mm were compared with that of in vivo grown follicles. IGF1R and IGF2R expression levels in 0.5 mm in vivo follicles were higher than in 1.0 or 0.2 mm follicles, respectively. IGFBP-1, IGFBP-2. IGFBP-3, IGFBP-4, IGFBP-5 and IGFBP-6 showed variable expression in the follicular size classes analyzed. In vitro grown follicles had significantly reduced expression levels for IGF1, IGF1R, IGFBP-3, IGFBP-5 and IGFBP-6 mRNA when compared with 0.2 mm follicles, but, when compared with in vivo grown follicles (0.5 mm), only IGFBP-1, IGFBP-2, IGFBP-3 and IGFBP-6 showed a reduction in their expression. In conclusion, IGFs, their receptors and IGFBPs showed variable expression of mRNA levels in the follicular size classes analyzed. PMID:23659735

  5. Protein hydrogen exchange: testing current models.

    PubMed

    Skinner, John J; Lim, Woon K; Bédard, Sabrina; Black, Ben E; Englander, S Walter

    2012-07-01

    To investigate the determinants of protein hydrogen exchange (HX), HX rates of most of the backbone amide hydrogens of Staphylococcal nuclease were measured by NMR methods. A modified analysis was used to improve accuracy for the faster hydrogens. HX rates of both near surface and well buried hydrogens are spread over more than 7 orders of magnitude. These results were compared with previous hypotheses for HX rate determination. Contrary to a common assumption, proximity to the surface of the native protein does not usually produce fast exchange. The slow HX rates for unprotected surface hydrogens are not well explained by local electrostatic field. The ability of buried hydrogens to exchange is not explained by a solvent penetration mechanism. The exchange rates of structurally protected hydrogens are not well predicted by algorithms that depend only on local interactions or only on transient unfolding reactions. These observations identify some of the present difficulties of HX rate prediction and suggest the need for returning to a detailed hydrogen by hydrogen analysis to examine the bases of structure-rate relationships, as described in the companion paper (Skinner et al., Protein Sci 2012;21:996-1005). PMID:22544567

  6. Protein carbonylation, protein aggregation and neuronal cell death in a murine model of multiple sclerosis

    NASA Astrophysics Data System (ADS)

    Dasgupta, Anushka

    Many studies have suggested that oxidative stress plays an important role in the pathophysiology of both multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). Yet, the mechanism by which oxidative stress leads to tissue damage in these disorders is unclear. Recent work from our laboratory has revealed that protein carbonylation, a major oxidative modification caused by severe and/or chronic oxidative stress conditions, is elevated in MS and EAE. Furthermore, protein carbonylation has been shown to alter protein structure leading to misfolding/aggregation. These findings prompted me to hypothesize that carbonylated proteins, formed as a consequence of oxidative stress and/or decreased proteasomal activity, promote protein aggregation to mediate neuronal apoptosis in vitro and in EAE. To test this novel hypothesis, I first characterized protein carbonylation, protein aggregation and apoptosis along the spinal cord during the course of myelin-oligodendrocyte glycoprotein (MOG)35-55 peptide-induced EAE in C57BL/6 mice [Chapter 2]. The results show that carbonylated proteins accumulate throughout the course of the disease, albeit by different mechanisms: increased oxidative stress in acute EAE and decreased proteasomal activity in chronic EAE. I discovered not only that there is a temporal correlation between protein carbonylation and apoptosis but also that carbonyl levels are significantly higher in apoptotic cells. A high number of juxta-nuclear and cytoplasmic protein aggregates containing the majority of the oxidized proteins are also present during the course of EAE, which seems to be due to reduced autophagy. In chapter 3, I show that when gluthathione levels are reduced to those in EAE spinal cord, both neuron-like PC12 (nPC12) cells and primary neuronal cultures accumulate carbonylated proteins and undergo cell death (both by necrosis and apoptosis). Immunocytochemical and biochemical studies also revealed a temporal/spatial relationship between carbonylation, protein aggregation and cellular apoptosis. Furthermore, the effectiveness of the carbonyl scavenger hydralazine, histidine hydrazide and methoxylamine at preventing cell death identifies protein carbonyls as the toxic species. Experiments using well-characterized apoptosis inhibitors place protein carbonylation downstream of the mitochondrial transition pore opening and upstream of caspase activation. These in vitro studies demonstrate for the first time a causal relationship between carbonylation, protein aggregation and apoptosis of neurons undergoing oxidative damage. This relationship was further strengthened with the experiments carried out in chapter 4, which show that inhibition of protein aggregation with congo red (CR) or 2-hydroxypropyl beta-cyclodextrin (HPCD) significantly reduced neuronal cell death without affecting the levels of oxidized proteins. Interestingly, large, juxta-nuclear aggregates are not formed upon GSH depletion, suggesting that the small protein aggregates are the cytotoxic species. Together, our data suggest that protein carbonylation causes protein aggregation to mediate neuronal apoptosis in vitro and that a similar mechanism might be contributing to neuronal/glial apoptosis in EAE. These studies provide the basis for testing protein carbonylation scavengers and protein aggregation inhibitors for the treatment of inflammatory demyelinating disorders.

  7. RECURSIVE PROTEIN MODELING: A DIVIDE AND CONQUER STRATEGY FOR PROTEIN STRUCTURE PREDICTION AND ITS CASE STUDY IN CASP9

    PubMed Central

    CHENG, JIANLIN; EICKHOLT, JESSE; WANG, ZHENG; DENG, XIN

    2013-01-01

    After decades of research, protein structure prediction remains a very challenging problem. In order to address the different levels of complexity of structural modeling, two types of modeling techniques — template-based modeling and template-free modeling — have been developed. Template-based modeling can often generate a moderate- to high-resolution model when a similar, homologous template structure is found for a query protein but fails if no template or only incorrect templates are found. Template-free modeling, such as fragment-based assembly, may generate models of moderate resolution for small proteins of low topological complexity. Seldom have the two techniques been integrated together to improve protein modeling. Here we develop a recursive protein modeling approach to selectively and collaboratively apply template-based and template-free modeling methods to model template-covered (i.e. certain) and template-free (i.e. uncertain) regions of a protein. A preliminary implementation of the approach was tested on a number of hard modeling cases during the 9th Critical Assessment of Techniques for Protein Structure Prediction (CASP9) and successfully improved the quality of modeling in most of these cases. Recursive modeling can signicantly reduce the complexity of protein structure modeling and integrate template-based and template-free modeling to improve the quality and efficiency of protein structure prediction. PMID:22809379

  8. Towards accurate structural characterization of metal centres in protein crystals: the structures of Ni and Cu T{sub 6} bovine insulin derivatives

    SciTech Connect

    Frankaer, Christian Grundahl; Mossin, Susanne; Ståhl, Kenny; Harris, Pernille, E-mail: ph@kemi.dtu.dk [Technical University of Denmark, Kemitorvet 207, DK-2800 Kgs. Lyngby (Denmark)

    2014-01-01

    The level of structural detail around the metal sites in Ni{sup 2+} and Cu{sup 2+} T{sub 6} insulin derivatives was significantly improved by using a combination of single-crystal X-ray crystallography and X-ray absorption spectroscopy. Photoreduction and subsequent radiation damage of the Cu{sup 2+} sites in Cu insulin was followed by XANES spectroscopy. Using synchrotron radiation (SR), the crystal structures of T{sub 6} bovine insulin complexed with Ni{sup 2+} and Cu{sup 2+} were solved to 1.50 and 1.45 Å resolution, respectively. The level of detail around the metal centres in these structures was highly limited, and the coordination of water in Cu site II of the copper insulin derivative was deteriorated as a consequence of radiation damage. To provide more detail, X-ray absorption spectroscopy (XAS) was used to improve the information level about metal coordination in each derivative. The nickel derivative contains hexacoordinated Ni{sup 2+} with trigonal symmetry, whereas the copper derivative contains tetragonally distorted hexacoordinated Cu{sup 2+} as a result of the Jahn–Teller effect, with a significantly longer coordination distance for one of the three water molecules in the coordination sphere. That the copper centre is of type II was further confirmed by electron paramagnetic resonance (EPR). The coordination distances were refined from EXAFS with standard deviations within 0.01 Å. The insulin derivative containing Cu{sup 2+} is sensitive towards photoreduction when exposed to SR. During the reduction of Cu{sup 2+} to Cu{sup +}, the coordination geometry of copper changes towards lower coordination numbers. Primary damage, i.e. photoreduction, was followed directly by XANES as a function of radiation dose, while secondary damage in the form of structural changes around the Cu atoms after exposure to different radiation doses was studied by crystallography using a laboratory diffractometer. Protection against photoreduction and subsequent radiation damage was carried out by solid embedment of Cu insulin in a saccharose matrix. At 100 K the photoreduction was suppressed by ?15%, and it was suppressed by a further ?30% on cooling the samples to 20 K.

  9. The Shape of Protein Crowders is a Major Determinant of Protein Diffusion

    PubMed Central

    Balbo, Jessica; Mereghetti, Paolo; Herten, Dirk-Peter; Wade, Rebecca C.

    2013-01-01

    As a model for understanding how molecular crowding influences diffusion and transport of proteins in cellular environments, we combined experimental and theoretical approaches to study the diffusion of proteins in highly concentrated protein solutions. Bovine serum albumin and ?-Globulin were chosen as molecular crowders and as tracers. These two proteins are representatives of the main types of plasma protein and have different shapes and sizes. Solutions consisting of one or both proteins were studied. The self-diffusion coefficients of the fluorescently labeled tracer proteins were measured by means of fluorescence correlation spectroscopy at a total protein concentration of up to 400 g/L. ?-Globulin is found to have a stronger influence as a crowder on the tracer self-diffusion coefficient than Bovine serum albumin. Brownian dynamics simulations show that the excluded volume and the shape of the crowding protein have a significantly stronger influence on translational and rotational diffusion coefficients, as well as transient oligomerization, than hydrodynamic or direct interactions. Anomalous subdiffusion, which is not observed at the experimental fluorescence correlation spectroscopy timescales (>100 ?s), appears only at very short timescales (<1 ?s) in the simulations due to steric effects of the proteins. We envision that the combined experimental and computational approach employed here can be developed to unravel the different biophysical contributions to protein motion and interaction in cellular environments by systematically varying protein properties such as molecular weight, size, shape, and electrostatic interactions. PMID:23561534

  10. A continuum hard-sphere model of protein adsorption

    NASA Astrophysics Data System (ADS)

    Finch, Craig; Clarke, Thomas; Hickman, James J.

    2013-07-01

    Protein adsorption plays a significant role in biological phenomena such as cell-surface interactions and the coagulation of blood. Two-dimensional random sequential adsorption (RSA) models are widely used to model the adsorption of proteins on solid surfaces. Continuum equations have been developed so that the results of RSA simulations can be used to predict the kinetics of adsorption. Recently, Brownian dynamics simulations have become popular for modeling protein adsorption. In this work a continuum model was developed to allow the results from a Brownian dynamics simulation to be used as the boundary condition in a computational fluid dynamics (CFD) simulation. Brownian dynamics simulations were used to model the diffusive transport of hard-sphere particles in a liquid and the adsorption of the particles onto a solid surface. The configuration of the adsorbed particles was analyzed to quantify the chemical potential near the surface, which was found to be a function of the distance from the surface and the fractional surface coverage. The near-surface chemical potential was used to derive a continuum model of adsorption that incorporates the results from the Brownian dynamics simulations. The equations of the continuum model were discretized and coupled to a CFD simulation of diffusive transport to the surface. The kinetics of adsorption predicted by the continuum model closely matched the results from the Brownian dynamics simulation. This new model allows the results from mesoscale simulations to be incorporated into micro- or macro-scale CFD transport simulations of protein adsorption in practical devices.

  11. A CONTINUUM HARD-SPHERE MODEL OF PROTEIN ADSORPTION

    PubMed Central

    Finch, Craig; Clarke, Thomas; Hickman, James J.

    2012-01-01

    Protein adsorption plays a significant role in biological phenomena such as cell-surface interactions and the coagulation of blood. Two-dimensional random sequential adsorption (RSA) models are widely used to model the adsorption of proteins on solid surfaces. Continuum equations have been developed so that the results of RSA simulations can be used to predict the kinetics of adsorption. Recently, Brownian dynamics simulations have become popular for modeling protein adsorption. In this work a continuum model was developed to allow the results from a Brownian dynamics simulation to be used as the boundary condition in a computational fluid dynamics (CFD) simulation. Brownian dynamics simulations were used to model the diffusive transport of hard-sphere particles in a liquid and the adsorption of the particles onto a solid surface. The configuration of the adsorbed particles was analyzed to quantify the chemical potential near the surface, which was found to be a function of the distance from the surface and the fractional surface coverage. The near-surface chemical potential was used to derive a continuum model of adsorption that incorporates the results from the Brownian dynamics simulations. The equations of the continuum model were discretized and coupled to a CFD simulation of diffusive transport to the surface. The kinetics of adsorption predicted by the continuum model closely matched the results from the Brownian dynamics simulation. This new model allows the results from mesoscale simulations to be incorporated into micro- or macro-scale CFD transport simulations of protein adsorption in practical devices. PMID:23729843

  12. A CONTINUUM HARD-SPHERE MODEL OF PROTEIN ADSORPTION.

    PubMed

    Finch, Craig; Clarke, Thomas; Hickman, James J

    2013-07-01

    Protein adsorption plays a significant role in biological phenomena such as cell-surface interactions and the coagulation of blood. Two-dimensional random sequential adsorption (RSA) models are widely used to model the adsorption of proteins on solid surfaces. Continuum equations have been developed so that the results of RSA simulations can be used to predict the kinetics of adsorption. Recently, Brownian dynamics simulations have become popular for modeling protein adsorption. In this work a continuum model was developed to allow the results from a Brownian dynamics simulation to be used as the boundary condition in a computational fluid dynamics (CFD) simulation. Brownian dynamics simulations were used to model the diffusive transport of hard-sphere particles in a liquid and the adsorption of the particles onto a solid surface. The configuration of the adsorbed particles was analyzed to quantify the chemical potential near the surface, which was found to be a function of the distance from the surface and the fractional surface coverage. The near-surface chemical potential was used to derive a continuum model of adsorption that incorporates the results from the Brownian dynamics simulations. The equations of the continuum model were discretized and coupled to a CFD simulation of diffusive transport to the surface. The kinetics of adsorption predicted by the continuum model closely matched the results from the Brownian dynamics simulation. This new model allows the results from mesoscale simulations to be incorporated into micro- or macro-scale CFD transport simulations of protein adsorption in practical devices. PMID:23729843

  13. Modeling of chemical inhibition from amyloid protein aggregation kinetics

    PubMed Central

    2014-01-01

    Backgrounds The process of amyloid proteins aggregation causes several human neuropathologies. In some cases, e.g. fibrillar deposits of insulin, the problems are generated in the processes of production and purification of protein and in the pump devices or injectable preparations for diabetics. Experimental kinetics and adequate modelling of chemical inhibition from amyloid aggregation are of practical importance in order to study the viable processing, formulation and storage as well as to predict and optimize the best conditions to reduce the effect of protein nucleation. Results In this manuscript, experimental data of insulin, A?42 amyloid protein and apomyoglobin fibrillation from recent bibliography were selected to evaluate the capability of a bivariate sigmoid equation to model them. The mathematical functions (logistic combined with Weibull equation) were used in reparameterized form and the effect of inhibitor concentrations on kinetic parameters from logistic equation were perfectly defined and explained. The surfaces of data were accurately described by proposed model and the presented analysis characterized the inhibitory influence on the protein aggregation by several chemicals. Discrimination between true and apparent inhibitors was also confirmed by the bivariate equation. EGCG for insulin (working at pH?=?7.4/T?=?37°C) and taiwaniaflavone for A?42 were the compounds studied that shown the greatest inhibition capacity. Conclusions An accurate, simple and effective model to investigate the inhibition of chemicals on amyloid protein aggregation has been developed. The equation could be useful for the clear quantification of inhibitor potential of chemicals and rigorous comparison among them. PMID:24572069

  14. Bovine mastitis: frontiers in immunogenetics.

    PubMed

    Thompson-Crispi, Kathleen; Atalla, Heba; Miglior, Filippo; Mallard, Bonnie A

    2014-01-01

    Mastitis is one of the most prevalent and costly diseases in the dairy industry with losses attributable to reduced milk production, discarded milk, early culling, veterinary services, and labor costs. Typically, mastitis is an inflammation of the mammary gland most often, but not limited to, bacterial infection, and is characterized by the movement of leukocytes and serum proteins from the blood to the site of infection. It contributes to compromised milk quality and the potential spread of antimicrobial resistance if antibiotic treatment is not astutely applied. Despite the implementation of management practises and genetic selection approaches, bovine mastitis control continues to be inadequate. However, some novel genetic strategies have recently been demonstrated to reduce mastitis incidence by taking advantage of a cow's natural ability to make appropriate immune responses against invading pathogens. Specifically, dairy cattle with enhanced and balanced immune responses have a lower occurrence of disease, including mastitis, and they can be identified and selected for using the high immune response (HIR) technology. Enhanced immune responsiveness is also associated with improved response to vaccination, increased milk, and colostrum quality. Since immunity is an important fitness trait, beneficial associations with longevity and reproduction are also often noted. This review highlights the genetic regulation of the bovine immune system and its vital contributions to disease resistance. Genetic selection approaches currently used in the dairy industry to reduce the incidence of disease are reviewed, including the HIR technology, genomics to improve disease resistance or immune response, as well as the Immunity(+)™ sire line. Improving the overall immune responsiveness of cattle is expected to provide superior disease resistance, increasing animal welfare and food quality while maintaining favorable production levels to feed a growing population. PMID:25339959

  15. Bovine Mastitis: Frontiers in Immunogenetics

    PubMed Central

    Thompson-Crispi, Kathleen; Atalla, Heba; Miglior, Filippo; Mallard, Bonnie A.

    2014-01-01

    Mastitis is one of the most prevalent and costly diseases in the dairy industry with losses attributable to reduced milk production, discarded milk, early culling, veterinary services, and labor costs. Typically, mastitis is an inflammation of the mammary gland most often, but not limited to, bacterial infection, and is characterized by the movement of leukocytes and serum proteins from the blood to the site of infection. It contributes to compromised milk quality and the potential spread of antimicrobial resistance if antibiotic treatment is not astutely applied. Despite the implementation of management practises and genetic selection approaches, bovine mastitis control continues to be inadequate. However, some novel genetic strategies have recently been demonstrated to reduce mastitis incidence by taking advantage of a cow’s natural ability to make appropriate immune responses against invading pathogens. Specifically, dairy cattle with enhanced and balanced immune responses have a lower occurrence of disease, including mastitis, and they can be identified and selected for using the high immune response (HIR) technology. Enhanced immune responsiveness is also associated with improved response to vaccination, increased milk, and colostrum quality. Since immunity is an important fitness trait, beneficial associations with longevity and reproduction are also often noted. This review highlights the genetic regulation of the bovine immune system and its vital contributions to disease resistance. Genetic selection approaches currently used in the dairy industry to reduce the incidence of disease are reviewed, including the HIR technology, genomics to improve disease resistance or immune response, as well as the Immunity+™ sire line. Improving the overall immune responsiveness of cattle is expected to provide superior disease resistance, increasing animal welfare and food quality while maintaining favorable production levels to feed a growing population. PMID:25339959

  16. Templated Ultrathin Polyelectrolyte Microreservoir for Delivery of Bovine Serum Albumin: Fabrication and Performance Evaluation

    Microsoft Academic Search

    Girish K. Gupta; Vikas Jain; Prabhat Ranjan Mishra

    2011-01-01

    The aim of the study was to develop ultrathin polyelectrolyte microreservoir (UPM) using two combinations of synthetic\\/synthetic\\u000a (S\\/s; poly(allylamine hydrochloride) (PAH)\\/sodium poly(styrenesulfonate)) and synthetic\\/natural (S\\/n; PAH\\/sodium alginate)\\u000a polyelectrolytes over spherical porous CaCO3 core particles (CP) followed by core removal and to evaluate its biocompatibility and integrity of loaded model protein bovine\\u000a serum albumin (BSA). A novel process for synthesis of

  17. [Preparation of monoclonal antibody against bovine phosphoenolpyruvate carboxykinase].

    PubMed

    Sheng, Meilei; Zhang, Xiao; Zhang, Wei; Zhang, Yanfei; Wang, Ying; Dai, Yifan

    2015-06-01

    Objective To prepare the monoclonal antibody against bovine phosphoenolpyruvate carboxykinase (PEPCK) and characterize its biological functions. Methods The recombinant plasmid containing PEPCK gene was constructed and used to transfect Escherichia coli. After expression induction in E.coli, the recombinant protein PEPCK was purified and used to immunize BALB/c mice. After the spleen B cells in the immunized mice were fused with murine myeloma cells, the positive clones were identified and selected by indirect ELISA for titer determination. PEPCK mAb produced by the positive hybridoma cells was enriched and its biological functions were examined by Western blotting, immunohistochemistry and immunoprecipitation. Results One hybridoma cell line steadily secreting PEPCK mAb was successfully generated, namely 3D36H. Western blotting, immunohistochemistry and immunoprecipitation showed that the PEPCK mAb was able to specifically bind to bovine PEPCK protein. Conclusion The bovine PEPCK mAb was prepared successfully and had a good ability and specificity. PMID:26062429

  18. QSAR modeling of ?-lactam binding to human serum proteins

    Microsoft Academic Search

    L. Mark Hall; Lowell H. Hall; Lemont B. Kier

    2003-01-01

    The binding of beta-lactams to human serum proteins was modeled with topological descriptors of molecular structure. Experimental data was the concentration of protein-bound drug expressed as a percent of the total plasma concentration (percent fraction bound, PFB) for 87 penicillins and for 115 ß-lactams. The electrotopological state indices (E-State) and the molecular connectivity chi indices were found to be the

  19. Intermolecular protein interactions in solutions of bovine lens beta L-crystallin. Results from 1/T1 nuclear magnetic relaxation dispersion profiles.

    PubMed Central

    Koenig, S H; Brown, R D; Kenworthy, A K; Magid, A D; Ugolini, R

    1993-01-01

    We report the magnetic field dependence of 1/T1 of solvent water protons and deuterons (nuclear magnetic relaxation dispersion, or NMRD, profiles) for solutions of steer lens beta L-crystallin. Such data allow the study of intermolecular protein interactions over a wide concentration range, here 1-34% vol/vol, by providing a measure of the rotational relaxation time of solute macromolecules. We conclude that, for approximately less than 5% protein, the solute particles are noncompact, with a rotationally averaged volume approximately three times that of a compact 60-kD sphere. (Earlier results for alpha-crystallin, approximately 1,000 kD, from optical and osmotic measurements (Vérétout and Tardieu, 1989. J. Mol. Biol. 205:713-728), show a similar, approximately twofold, effect). At intermediate concentrations, to approximately 20% protein, there is evidence for limited association or oligomerization, as found for the structurally related gamma II-crystallin (Koenig et al. 1990. Biophys. J. 57:461-469), to a limiting size about two-thirds that of alpha-crystallin. The difference in NMRD behavior of the three classes of crystallins is consonant with their differing osmotic properties (Vérétout and Tardieu. J. Mol. Biol. 1989, 205:713-728; Kenworthy, McIntosh, and Magid. Biophys. J. 1992. 61:A477; Tardieu et al. 1992. Eur. Biophys. J. 21:1-12). We indicate how the unusual structures and interactions of these three classes of proteins can be combined to optimize transparency and minimize colloid osmotic difficulties in eye lens. PMID:8388267

  20. Activation of signal transducer and activator of transcription 5 (STAT5) is linked to ?1-integrin protein abundance in unilaterally milked bovine mammary glands.

    PubMed

    Murney, R; Stelwagen, K; Wheeler, T T; Margerison, J K; Singh, K

    2015-05-01

    Prolactin (PRL) is important in the regulation of milk synthesis in mammary epithelial cells (MEC). In cattle, circulating levels of PRL are not limiting, suggesting the possible involvement of other factors that may control the response to PRL at the cellular level. The effects of milking frequency (MF) on milk synthesis are controlled locally within mammary glands and involve PRL signaling. To further investigate this relationship between MF and PRL signaling, udder halves of 17 dairy cows were milked either 4 times a day (4×) or once a day (1×) for 14 d in early lactation. Mammary biopsies were obtained 3 to 5h following milking from both udder halves of 10 cows, and changes in PRL and associated pathways were measured. The abundance of STAT5A mRNA was higher after 4× milking, whereas that of the PRL receptor (PRLR) and STAT3 were lower relative to that after 1× milking. In 4× mammary tissues, the protein levels of STAT5, activated STAT5, and ?1-integrin were higher, whereas the those of the long isoform of PRL receptor and activated STAT3 were lower than 1× tissues. The activation of STAT5 correlated strongly with major milk protein mRNA abundance (r=0.86 to 0.94) and ?1-integrin protein levels (r=0.91). These results confirm that major milk protein gene expression is associated with STAT5 activation and suggests that the STAT5 and ?1-integrin signaling pathways are linked. Modulation of ?1-integrin abundance in response to changes in MF may be a mechanism that controls the MEC ability to respond to PRL and therefore its secretory activity. PMID:25771047

  1. Application of bovine pituitary extract and polyglycolide/poly(lactide-co-glycolide) scaffold to the cultivation of bovine knee chondrocytes.

    PubMed

    Kuo, Yung-Chih; Chung, Chiu-Yen

    2005-01-01

    In vitro cultivation of primary bovine knee chondrocytes (BKCs), using bovine pituitary extract (BPE) and porous scaffolds composed of polyglycolide (PGA) and 85/15 poly(lactide-co-glycolide) (PLGA), was investigated. Here, BPE was prepared from fresh bovine pituitaries, and cylindrical PGA/PLGA scaffolds with various chemical compositions were fabricated by solvent merging/particulate leaching method. Experimental results showed that in microcarrier systems, the rate of BKC growth on PGA surfaces is faster than that on PLGA surfaces, and the decrease in the medium pH value of BKCs-adsorbed PGA particles is faster than that of BKCs-adsorbed PLGA particles. After 28-day construct cultivation, the BKC amount and the content of glycosaminoglycans and collagen per construct increased with BPE protein concentration. For a constant BPE protein concentration, a higher PGA percentage in scaffold leads to a better biological environment for the growth of BKCs and the synthesis of extracellar matrices. PMID:16321055

  2. Bayesian Proteoform Modeling Improves Protein Quantification of Global Proteomic Measurements

    SciTech Connect

    Webb-Robertson, Bobbie-Jo M.; Matzke, Melissa M.; Datta, Susmita; Payne, Samuel H.; Kang, Jiyun; Bramer, Lisa M.; Nicora, Carrie D.; Shukla, Anil K.; Metz, Thomas O.; Rodland, Karin D.; Smith, Richard D.; Tardiff, Mark F.; McDermott, Jason E.; Pounds, Joel G.; Waters, Katrina M.

    2014-12-01

    As the capability of mass spectrometry-based proteomics has matured, tens of thousands of peptides can be measured simultaneously, which has the benefit of offering a systems view of protein expression. However, a major challenge is that with an increase in throughput, protein quantification estimation from the native measured peptides has become a computational task. A limitation to existing computationally-driven protein quantification methods is that most ignore protein variation, such as alternate splicing of the RNA transcript and post-translational modifications or other possible proteoforms, which will affect a significant fraction of the proteome. The consequence of this assumption is that statistical inference at the protein level, and consequently downstream analyses, such as network and pathway modeling, have only limited power for biomarker discovery. Here, we describe a Bayesian model (BP-Quant) that uses statistically derived peptides signatures to identify peptides that are outside the dominant pattern, or the existence of multiple over-expressed patterns to improve relative protein abundance estimates. It is a research-driven approach that utilizes the objectives of the experiment, defined in the context of a standard statistical hypothesis, to identify a set of peptides exhibiting similar statistical behavior relating to a protein. This approach infers that changes in relative protein abundance can be used as a surrogate for changes in function, without necessarily taking into account the effect of differential post-translational modifications, processing, or splicing in altering protein function. We verify the approach using a dilution study from mouse plasma samples and demonstrate that BP-Quant achieves similar accuracy as the current state-of-the-art methods at proteoform identification with significantly better specificity. BP-Quant is available as a MatLab ® and R packages at https://github.com/PNNL-Comp-Mass-Spec/BP-Quant.

  3. Bovine sperm plasma membrane proteomics through biotinylation and subcellular enrichment.

    PubMed

    Kasvandik, Sergo; Sillaste, Gerly; Velthut-Meikas, Agne; Mikelsaar, Aavo-Valdur; Hallap, Triin; Padrik, Peeter; Tenson, Tanel; Jaakma, Ülle; Kõks, Sulev; Salumets, Andres

    2015-06-01

    A significant proportion of mammalian fertilization is mediated through the proteomic composition of the sperm surface. These protein constituents can present as biomarkers to control and regulate breeding of agricultural animals. Previous studies have addressed the bovine sperm cell apical plasma membrane (PM) proteome with nitrogen cavitation enrichment. Alternative workflows would enable to expand the compositional data more globally around the entire sperm's surface. We used a cell surface biotin-labeling in combination with differential centrifugation to enrich sperm surface proteins. Using nano-LC MS/MS, 338 proteins were confidently identified in the PM-enriched proteome. Functional categories of sperm-egg interaction, protein turnover, metabolism as well as molecular transport, spermatogenesis, and signal transduction were represented by proteins with high quantitative signal in our study. A highly significant degree of enrichment was found for transmembrane and PM-targeted proteins. Among them, we also report proteins previously not described on bovine sperm (CPQ, CD58, CKLF, CPVL, GLB1L3, and LPCAT2B) of which CPQ and CPVL cell surface localization was further validated. A descriptive overview of the bovine sperm PM integral and peripheral proteins is provided to complement future studies on animal reproduction and its relation to sperm cell surface. All MS data have been deposited in the ProteomeXchange with identifier PXD001096 (http://proteomecentral.proteomexchange.org/dataset/PXD001096). PMID:25603787

  4. Modeling the flexibility of alpha helices in protein interfaces : structure based design and prediction of helix-mediated protein-protein interactions

    E-print Network

    Apgar, James R. (James Reasoner)

    2008-01-01

    Protein-protein interactions play an essential role in many biological functions. Prediction and design of these interactions using computational methods requires models that can be used to efficiently sample structural ...

  5. Molecular cloning and characterization of a novel bovine IFN-?.

    PubMed

    Guo, Yongli; Gao, Mingchun; Bao, Jun; Luo, Xiuxin; Liu, Ying; An, Dong; Zhang, Haili; Ma, Bo; Wang, Junwei

    2015-03-01

    A bovine IFN-? (BoIFN-?) gene was amplified from bovine liver genomic DNA consisting of a 463bp partial 5'UTR, 582bp complete ORF and 171bp partial 3'UTR, which encodes a protein of 193 amino acids with a 21-amino acid signal peptide and shares 61 to 87% identity with other species IFN-?. Then BoIFN-? gene was characterized, and it can be transcribed in EBK cells at a high level after being infected by VSV. Recombinant proteins were expressed in Escherichia coli and the antiviral activity was determined in vitro, which revealed that bovine IFN-? has less antiviral activity than bovine IFN-?. In addition, an immunofluorescence assay indicated that BoIFN-? expressed in MDBK cells could be detected by polyclonal antibody against BoIFN-?. Furthermore, the BoIFN-? gene can be constitutively expressed in the liver, thymus, kidney, small intestine and testis, but not in the heart. This study revealed that BoIFN-? has the typical characteristics of type I interferon and can be expressed constitutively in certain tissue, which not only can be a likely candidate for a novel, effective therapeutic agent, but also facilitate further research on the role of bovine IFN system. PMID:25523095

  6. Constitutive and Trophoblast-Specific Expression of a Class of Bovine Interferon Genes

    Microsoft Academic Search

    James C. Cross; R. Michael Roberts

    1991-01-01

    The early conceptus in sheep and cattle secretes a low molecular weight protein called ovine and bovine trophoblast protein 1 (TP-1) that is critical for establishment of pregnancy. TP-1 is a type I interferon (IFN) and is most related to IFN-omega. Here we have determined if TP-1 genes are regulated similarly to other type I IFNs. Single day 18 bovine

  7. Information-driven modeling of protein-peptide complexes.

    PubMed

    Trellet, Mikael; Melquiond, Adrien S J; Bonvin, Alexandre M J J

    2015-01-01

    Despite their biological importance in many regulatory processes, protein-peptide recognition mechanisms are difficult to study experimentally at the structural level because of the inherent flexibility of peptides and the often transient interactions on which they rely. Complementary methods like biomolecular docking are therefore required. The prediction of the three-dimensional structure of protein-peptide complexes raises unique challenges for computational algorithms, as exemplified by the recent introduction of protein-peptide targets in the blind international experiment CAPRI (Critical Assessment of PRedicted Interactions). Conventional protein-protein docking approaches are often struggling with the high flexibility of peptides whose short sizes impede protocols and scoring functions developed for larger interfaces. On the other side, protein-small ligand docking methods are unable to cope with the larger number of degrees of freedom in peptides compared to small molecules and the typically reduced available information to define the binding site. In this chapter, we describe a protocol to model protein-peptide complexes using the HADDOCK web server, working through a test case to illustrate every steps. The flexibility challenge that peptides represent is dealt with by combining elements of conformational selection and induced fit molecular recognition theories. PMID:25555727

  8. Evaluation of the antimicrobial effect of super-oxidized water (Sterilox®) and sodium hypochlorite against Enterococcus faecalis in a bovine root canal model

    PubMed Central

    ROSSI-FEDELE, Giampiero; de FIGUEIREDO, José Antonio Poli; STEIER, Liviu; CANULLO, Luigi; STEIER, Gabriela; ROBERTS, Adam P.

    2010-01-01

    Ideally root canal irrigants should have, amongst other properties, antimicrobial action associated with a lack of toxicity against periapical tissues. Sodium hypochlorite (NaOCl) is a widely used root canal irrigant, however it has been shown to have a cytotoxic effect on vital tissue and therefore it is prudent to investigate alternative irrigants. Sterilox's Aquatine Alpha Electrolyte® belongs to the group of the super-oxidized waters; it consists of a mixture of oxidizing substances, and has been suggested to be used as root canal irrigant. Super-oxidized waters have been shown to provide efficient cleaning of root canal walls, and have been proposed to be used for the disinfection of medical equipment. Objective To compare the antimicrobial action against Enterococcus faecalis of NaOCl, Optident Sterilox Electrolyte Solution® and Sterilox's Aquatine Alpha Electrolyte® when used as irrigating solutions in a bovine root canal model. Methodology Root sections were prepared and inoculated with E. faecalis JH2-2. After 10 days of incubation the root canals were irrigated using one of three solutions (NaOCl, Optident Sterilox Electrolyte Solution® and Sterilox's Aquatine Alpha Electrolyte®) and subsequently sampled by grinding dentin using drills. The debris was placed in BHI broth and dilutions were plated onto fresh agar plates to quantify growth. Results Sodium hypochlorite was the only irrigant to eliminate all bacteria. When the dilutions were made, although NaOCl was still statistically superior, Sterilox's Aquatine Alpha Electrolyte® solution was superior to Optident Sterilox Electrolyte Solution®. Conclusion Under the conditions of this study Sterilox's Aquatine Alpha Electrolyte® appeared to have significantly more antimicrobial action compared to the Optident Sterilox Electrolyte Solution® alone, however NaOCl was the only solution able to consistently eradicate E. faecalis in the model. PMID:21085808

  9. Solvated dissipative electro-elastic network model of hydrated proteins

    NASA Astrophysics Data System (ADS)

    Martin, Daniel R.; Matyushov, Dmitry V.

    2012-10-01

    Elastic network models coarse grain proteins into a network of residue beads connected by springs. We add dissipative dynamics to this mechanical system by applying overdamped Langevin equations of motion to normal-mode vibrations of the network. In addition, the network is made heterogeneous and softened at the protein surface by accounting for hydration of the ionized residues. Solvation changes the network Hessian in two ways. Diagonal solvation terms soften the spring constants and off-diagonal dipole-dipole terms correlate displacements of the ionized residues. The model is used to formulate the response functions of the electrostatic potential and electric field appearing in theories of redox reactions and spectroscopy. We also formulate the dielectric response of the protein and find that solvation of the surface ionized residues leads to a slow relaxation peak in the dielectric loss spectrum, about two orders of magnitude slower than the main peak of protein relaxation. Finally, the solvated network is used to formulate the allosteric response of the protein to ion binding. The global thermodynamics of ion binding is not strongly affected by the network solvation, but it dramatically enhances conformational changes in response to placing a charge at the active site of the protein.

  10. Learning sequence determinants of protein:protein interaction specificity with sparse graphical models.

    PubMed

    Kamisetty, Hetunandan; Ghosh, Bornika; Langmead, Christopher James; Bailey-Kellogg, Chris

    2015-06-01

    In studying the strength and specificity of interaction between members of two protein families, key questions center on which pairs of possible partners actually interact, how well they interact, and why they interact while others do not. The advent of large-scale experimental studies of interactions between members of a target family and a diverse set of possible interaction partners offers the opportunity to address these questions. We develop here a method, DgSpi (data-driven graphical models of specificity in protein:protein interactions), for learning and using graphical models that explicitly represent the amino acid basis for interaction specificity (why) and extend earlier classification-oriented approaches (which) to predict the ?G of binding (how well). We demonstrate the effectiveness of our approach in analyzing and predicting interactions between a set of 82 PDZ recognition modules against a panel of 217 possible peptide partners, based on data from MacBeath and colleagues. Our predicted ?G values are highly predictive of the experimentally measured ones, reaching correlation coefficients of 0.69 in 10-fold cross-validation and 0.63 in leave-one-PDZ-out cross-validation. Furthermore, the model serves as a compact representation of amino acid constraints underlying the interactions, enabling protein-level ?G predictions to be naturally understood in terms of residue-level constraints. Finally, the model DgSpi readily enables the design of new interacting partners, and we demonstrate that designed ligands are novel and diverse. PMID:25973864

  11. Bovine spongiform encephalopathy in Sweden: an H-type variant

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine spongiform encephalopathy (BSE) had never been detected in Sweden until 2006, when the active surveillance identified a case in a 12-year-old cow. The case was an unusual form since several molecular features of the protease-resistant prion protein (PrP**res) were different from classical BSE...

  12. Onapristone (ZK299) and mifepristone (RU486) regulate the messenger RNA and protein expression levels of the progesterone receptor isoforms A and B in the bovine endometrium.

    PubMed

    Rekawiecki, Robert; Kowalik, Magdalena K; Kotwica, Jan

    2015-08-01

    The aim of this study was to examine whether progesterone (P4) and its antagonists, onapristone (ZK299) and mifepristone (RU486), affect the levels of PGRA and PGRB messenger RNA (mRNA) and protein in the cow uterus which may be important in understanding whether the final physiological effect evoked by an antagonist depends on PGR isoform bound to the antagonist. Endometrial slices on Days 6 to 10 and 17 to 20 of the estrous cycle were treated for 6 or 24 hours for mRNA and protein expression analysis, respectively, with P4, ZK299, or RU486 at a dose of 10(-4), 10(-5), or 10(-6) M. In the samples on Days 6 to 10 of the estrous cycle, PGRAB mRNA was stimulated by P4 (10(-4) M; P < 0.01) and RU486 (10(-6); P < 0.001) and was decreased by ZK299 (10(-5); P < 0.05). In contrast, PGRB mRNA was decreased by the all P4 (P < 0.01) and ZK299 (P < 0.001) doses and by two of the RU486 doses (10(-4) M; P < 0.01 and 10(-5) M; P < 0.01). In samples on Days 17 to 20 of the estrous cycle, PGRAB mRNA was stimulated by RU486 (10(-5) M; P < 0.001). PGRB mRNA was decreased by P4 (10(-4) and 10(-5) M; P < 0.001), ZK299 (10(-4) and 10(-5) M; P < 0.001), and RU486 (10(-4) M; P < 0.01 and 10(-6) M; P < 0.001) and was increased by ZK299 (10(-6) M; P < 0.001) and RU486 (10(-5) M; P < 0.001). In samples on Days 6 to 10 of the estrous cycle, PGRB protein levels were decreased (P < 0.05) by all three ZK299 doses and by two of the RU486 doses (10(-4) M; P < 0.05 and 10(-5) M; P < 0.01). In contrast, in samples on Days 17 to 20, both PGRA and PGRB protein levels were decreased by ZK299 stimulation (10(-5) M; P < 0.05 and 10(-5) M; P < 0.01, respectively), whereas only PGRA protein levels were increased by RU486 (10(-5) M; P < 0.01). Both ZK299 and RU486 may exhibit both agonist and antagonist properties depending on which receptor isoform they affect. As a result, an increase or decrease in the expression of a particular PGR isoform will be observed. PMID:25976976

  13. Protein oxidation affects proteolysis in a meat model system.

    PubMed

    Berardo, Alberto; Claeys, Erik; Vossen, Els; Leroy, Frédéric; De Smet, Stefaan

    2015-08-01

    The effect of hydrogen peroxide-induced protein oxidation and pH (4.8 and 5.2) on meat proteolysis was investigated in a meat model system for dry fermented sausages. In oxidised samples, increased protein carbonyl contents and decreased thiol concentrations were found. The initial concentration of protein carbonyls was significantly lower in oxidised samples at pH4.8 than in ones at pH5.2, but after ten days comparable levels were reached. The inhibition of proteolysis by the addition of a protease inhibitor cocktail did not influence protein oxidation. Yet, proteolysis was negatively affected by low pH values as well as by oxidation, resulting in a reduced release of amino acids during ripening. PMID:25909819

  14. Evaluation of a Rapid Western Immunoblotting Procedure for the Diagnosis of Bovine Spongiform Encephalopathy (BSE) in the UK

    Microsoft Academic Search

    W. A. Cooley; J. K. Clark; S. J. Ryder; L. A. Davis; S. S. J. Farrelly; M. J. Stack

    2001-01-01

    Bovine brain tissue samples from 625 UK cattle, clinically suspected as bovine spongiform encephalopathy (BSE) cases, were used in a blind analysis to assess a rapid Western immunoblotting technique (Prionics Check; Prionics AG, Zurich), which detects bovine disease-specific protease-resistant prion protein (PrPSc). By means of statutory histopathological examination, 599 of the 625 cattle were confirmed as BSE cases by the

  15. Insights into protein evolution landscapes from folding models

    NASA Astrophysics Data System (ADS)

    Koonin, Eugene

    2011-03-01

    Off-lattice models of protein folding were employed to investigate the origins of the evolutionary rate distributions and fitness landscapes. For each robust folder, the network of sequences that share its native structure is identified. The fitness of a sequence is a simple function of the number of misfolded molecules produced to reach a characteristic protein abundance. Fixation probabilities of mutants are computed under a simple population dynamics model, and the fold-averaged evolution rate is computed a using a Markov chain on the fold network. The distribution of the logarithm of the evolution rates exhibits a peak with a long tail on the low rate side and resembles the universal empirical distribution of the evolutionary rates more closely than either distribution resembles the log-normal distribution. We next addressed the question of the extent of determinism in protein evolution. Limited empirical studies suggest that the fitness landscapes of protein evolution are significantly smoother, or more additive, than random landscapes. However, widespread sign epistasis seems to restrict evolution to a small fraction of available trajectories, thus making the evolutionary process substantially deterministic. Access to complete fitness landscapes within the model framework enables exhaustive analysis of evolutionary trajectories. The model landscapes were compared to a continuum of artificial landscapes of varying smoothness. In maximally smooth, fully additive landscapes, evolution cannot be predicted because all paths are accessible. However, a small amount of noise can make most paths inaccessible while preserving the overall structure of the landscape. Although the model landscapes are almost additive, most paths are non-monotonic with respect to fitness, so evolutionary trajectories can be approximately predicted. Thus, protein folding physics seems to dictate the universal distribution of the evolutionary rates of protein-coding genes and the quasi-deterministic character of evolution.

  16. Assignment of disulfide bridges in bovine CD36.

    PubMed

    Rasmussen, J T; Berglund, L; Rasmussen, M S; Petersen, T E

    1998-10-15

    The multifunctional membrane protein CD36 is expressed on platelets, mature monocytes and macrophages, microvascular endothelial cells and mammary epithelial cells. The exact physiological function of this glycoprotein is unclear. In order to determine the number and pattern of disulfide bridges, CD36 was purified from bovine milk fat globule membranes. The purification procedure involved Triton X-114 extraction, DEAE-Sepharose ion-exchange chromatography and reverse-phase chromatography on a Resource RPC column. The CD36 preparation was used for characterization of the disulfide bridge pattern, which was determined by peptide mapping, amino acid sequence analysis, and matrix-assisted laser-desorption ionization/time of flight mass spectrometry. We have found that there are no free cysteines in CD36 and that the six centrally clustered cysteines are linked by disulfide bonds, Cys242-Cys310, Cys271-Cys332 and Cys312-Cys321, resulting in a 1-3, 2-6 and 4-5 arrangement of the disulfide bridges. These data are in agreement with a model where the protein is oriented so that it has two short intracellular segments (residues 1-6 and 461-471) and two transmembrane domains (residues 7-28 and 439-460), and with four cysteines expected to be acylated placed near the intracellular side of the membrane. The remaining part of CD36 is extracellular, comprising eight glycosylations and three disulfide bridges. In the CD36 family of membrane proteins, it is likely that a similar pattern of disulfide bridges can be found in the sensory neuron membrane protein-1 from the silk moth Antheraea polyphemus and the mammalian scavenger receptor class B type I, whereas lysosome membrane protein II, and epithelial membrane protein from Drosophila melanogaster are both lacking one cysteine in the area of interest. PMID:9826197

  17. A backbone based protein model with explicit solvent

    NASA Astrophysics Data System (ADS)

    Sharma, Sumit; Buldyrev, Sergey; Rossky, Peter J.; Stanley, H. Eugene; Debenedetti, Pablo G.; Angell, C. Austen; Kumar, Sanat K.

    2011-03-01

    The computational expense of folding atomistically detailed protein models is prohibitive. Hence minimalist models of proteins are a popular choice. The minimalist models developed so far have excluded water, and treated the hydrophobic effect as an effective attraction between hydrophobic monomers. This simplified treatment does not capture the temperature-dependent variations in entropy and enthalpy of water molecules. Proteins have a predominantly water-screened hydrophobic core and water-exposed polar groups. This structural feature should alter the dynamics of proteins and surrounding water from that of a hydrophobic homopolymer in water. To include these features in a minimalist model, we designed heteropolymers of polar and hydrophobic monomers in explicit water-like medium. The polar monomers and water molecules were modeled with the Jagla potential, which has been shown to reproduce many water-like thermodynamic properties, and the hydrophobic monomers as hard spheres. We discuss a methodology for optimizing the sequence of these heteropolymers and how the hydrophobic collapse of these heteropolymers differs from that of a random heteropolymer.

  18. Sketching protein aggregation with a physics-based toy model

    NASA Astrophysics Data System (ADS)

    Enciso, Marta; Rey, Antonio

    2013-09-01

    We explore the applicability of a single-bead coarse-grained molecular model to describe the competition between protein folding and aggregation. We have designed very simple and regular sequences, based on our previous studies on peptide aggregation, that successfully fold into the three main protein structural families (all-?, all-?, and ? + ?). Thanks to equilibrium computer simulations, we evaluate how temperature and concentration promote aggregation. Aggregates have been obtained for all the amino acid sequences considered, showing that this process is common to all proteins, as previously stated. However, each structural family presents particular characteristics that can be related to its specific balance between hydrogen bond and hydrophobic interactions. The model is very simple and has limitations, yet it is able to reproduce both the cooperative folding of isolated polypeptide chains with regular sequences and the formation of different types of aggregates at high concentrations.

  19. Interaction of Zinc, Ascorbic Acid, and Folic Acid in Glycation with Albumin as Protein Model

    Microsoft Academic Search

    Rashmi Tupe; Vaishali Agte

    2010-01-01

    Using albumin as model, we conducted series of in vitro glycation experiments to examine role of zinc in glycation using glucose\\u000a at 4–100 mg\\/ml, incubations at 37°C or 60°C, duration of 2 or 4 weeks and in presence of zinc or ascorbic acid (AA) or folic\\u000a acid (FA). Modifications of bovine serum albumin (BSA) were examined by using fluorescence of advanced glycation

  20. Minimal models for proteins and RNA: From folding to function

    E-print Network

    D. L. Pincus; S. S. Cho; C. Hyeon; D. Thirumalai

    2008-08-22

    We present a panoramic view of the utility of coarse-grained (CG) models to study folding and functions of proteins and RNA. Drawing largely on the methods developed in our group over the last twenty years, we describe a number of key applications ranging from folding of proteins with disulfide bonds to functions of molecular machines. After presenting the theoretical basis that justifies the use of CG models, we explore the biophysical basis for the emergence of a finite number of folds from lattice models. The lattice model simulations of approach to the folded state show that non-native interactions are relevant only early in the folding process - a finding that rationalizes the success of structure-based models that emphasize native interactions. Applications of off-lattice $C_{\\alpha}$ and models that explicitly consider side chains ($C_{\\alpha}$-SCM) to folding of $\\beta$-hairpin and effects of macromolecular crowding are briefly discussed. Successful application of a new class of off-lattice model, referred to as the Self-Organized Polymer (SOP), is shown by describing the response of Green Fluorescent Protein (GFP) to mechanical force. The utility of the SOP model is further illustrated by applications that clarify the functions of the chaperonin GroEL and motion of the molecular motor kinesin. We also present two distinct models for RNA, namely, the Three Site Interaction (TIS) model and the SOP model, that probe forced unfolding and force quench refolding of a simple hairpin and {\\it Azoarcus} ribozyme. The predictions based on the SOP model show that force-induced unfolding pathways of the ribozyme can be dramatically changed by varying the loading rate. We conclude with a discussion of future prospects for the use of coarse-grained models in addressing problems of outstanding interest in biology.

  1. Comparative study of sperm washing and selection methods after cryopreservation and its influence on sperm subpopulational structure in a bovine model.

    PubMed

    García-Herreros, Manuel; Leal, Claudia L V

    2014-12-01

    The effect of different sperm washing-selection methods on sperm morphometric characteristics as a study to detect differences in the subpopulational structure has been carried out in detail in a bovine model. Cryopreserved sperm samples from 5 bulls were thawed, pooled, and processed by TALP-washing centrifugation method (TWCM), selective Percoll discontinuous density-gradient centrifugation method (PDGM), and self-migration swim-up separation method (SUMM). Live-dead assay (SYBR-14/ethidium homodimer-1), chlortetracycline assay (CTC), and sperm motility were assessed, and aliquots of sperm were processed for automated sperm morphometry analysis (ASMA) simultaneously before (raw thawed sperm used as control, RTS) and after different sperm washing-selection techniques. Deleterious effects of different methods were evident, particularly on sperm membrane integrity (p < 0.05) and capacitation status (p < 0.05). Moreover, each cell was measured for four primary dimensional parameters, and three shape parameters. All sperm morphometric parameters evaluated were analyzed by principal component analysis (PCA) and multivariate clustering analyses. PCA revealed two principal components for each sperm washing or separation method explaining more than the 91% of the variance. The number of subpopulations found was the same for all methods (four) except for PDGM (three). However, irrespective of the number of subpopulations defined by PCA and clustering analyses, the sperm subpopulational structure was found to be different and strongly influenced by the sperm selection procedure due to statistical differences found regarding the sperm biophysical changes induced by each method used (p < 0.001). It is concluded that different sperm washing-selection methods commonly used during IVF process, may lead to alterations in sperm morphometric characteristics, which might explain the different results seen after IVF, since an important influence of these methods on sperm subpopulational structure has been demonstrated. PMID:25007138

  2. Clathrin assembly proteins: affinity purification and a model for coat assembly

    PubMed Central

    1987-01-01

    Assembly protein (AP) preparations from bovine brain coated vesicles have been fractionated by clathrin-Sepharose affinity chromatography. Two distinct fractions that possess coat assembly activity were obtained and are termed AP-1 and AP-2. The AP-1, not retained on the resin, has principal components with molecular weights of 108,000, 100,000, 47,000, and 19,000. The AP-2, bound to the resin and eluted by Tris-HCl at a concentration that parallels the latter's effect on coat disassembly, corresponds to the active complex described previously (Zaremba, S., and J. H. Keen, 1983, J. Cell Biol., 97:1339-1347). Its composition is similar to that of the AP-1 in that it contains 100,000- , 50,000-, and 16,000-mol-wt polypeptides in equimolar amounts; minor amounts of 112,000- and 115,000-mol-wt polypeptides are also present. Both are distinct from a recently described assembly protein of larger subunit molecular weight that we term AP-3. These results indicate the existence of a family of assembly proteins within cells. On incubation with clathrin both AP-1 and AP-2 induce the formation of coat structures, those containing AP-1 slightly smaller (mean diameter = 72 nm) than those formed in the presence of AP-2 (mean diameter = 79 nm); both structures have been detected previously in coated vesicle preparations from brain. Coats formed in the presence of AP-2 consistently contain approximately one molecule each of the 100,000-, 50,000-, and 16,000-mol-wt polypeptides per clathrin trimer. By low angle laser light scattering the molecular weight of native AP-2 was determined to be approximately 343,000, indicating that it is a dimer of each of the three subunits, and implying that it is functionally bivalent in clathrin binding. A model for AP-mediated coat assembly is proposed in which a bivalent AP-2 molecule bridges the distal legs or terminal domains of two clathrin trimers that are destined to occupy adjacent vertices in the assembled coat. Binding of a second AP-2 molecule locks these two trimers in register for assembly and further addition of AP-2 to free trimer legs promotes completion of the clathrin lattice. Effects of AP binding on the angle and flexibility of the legs at the hub of the trimer (the "pucker") are suggested to account for the characteristic size distributions of coats formed under varied conditions and, more speculatively, to contribute to the transformation of flat clathrin lattices to curved coated vesicles that are thought to occur during endocytosis. PMID:2890644

  3. MetaMEME: Motifbased Hidden Markov Models of Protein Families

    E-print Network

    Bailey, Timothy L.

    Meta­MEME: Motif­based Hidden Markov Models of Protein Families William N. Grundy \\Lambda bgrundy by the EM algorithm using the MEME software. Because motif­based HMMs have relatively few parameters searches, especially when training sets contain few sequences. Availability: http://www.sdsc.edu/MEME

  4. Tools for comparative protein structure modeling and analysis

    E-print Network

    Sali, Andrej

    magnetic resonance (NMR) spectroscopy, have enhanced the quality and speed of structural studies (1 known protein structure (the template); (ii) alignment of the target sequence and the template(s); (iii-template sequence identity. The errors encountered in comparative modeling include fold assignment and alignment

  5. Anomalous diffusion in neutral evolution of model proteins

    NASA Astrophysics Data System (ADS)

    Nelson, Erik D.; Grishin, Nick V.

    2015-06-01

    Protein evolution is frequently explored using minimalist polymer models, however, little attention has been given to the problem of structural drift, or diffusion. Here, we study neutral evolution of small protein motifs using an off-lattice heteropolymer model in which individual monomers interact as low-resolution amino acids. In contrast to most earlier models, both the length and folded structure of the polymers are permitted to change. To describe structural change, we compute the mean-square distance (MSD) between monomers in homologous folds separated by n neutral mutations. We find that structural change is episodic, and, averaged over lineages (for example, those extending from a single sequence), exhibits a power-law dependence on n . We show that this exponent depends on the alignment method used, and we analyze the distribution of waiting times between neutral mutations. The latter are more disperse than for models required to maintain a specific fold, but exhibit a similar power-law tail.

  6. Comparative protein structure modeling by iterative alignment, model building and model assessment.

    PubMed

    John, Bino; Sali, Andrej

    2003-07-15

    Comparative or homology protein structure modeling is severely limited by errors in the alignment of a modeled sequence with related proteins of known three-dimensional structure. To ameliorate this problem, we have developed an automated method that optimizes both the alignment and the model implied by it. This task is achieved by a genetic algorithm protocol that starts with a set of initial alignments and then iterates through re-alignment, model building and model assessment to optimize a model assessment score. During this iterative process: (i) new alignments are constructed by application of a number of operators, such as alignment mutations and cross-overs; (ii) comparative models corresponding to these alignments are built by satisfaction of spatial restraints, as implemented in our program MODELLER; (iii) the models are assessed by a variety of criteria, partly depending on an atomic statistical potential. When testing the procedure on a very difficult set of 19 modeling targets sharing only 4-27% sequence identity with their template structures, the average final alignment accuracy increased from 37 to 45% relative to the initial alignment (the alignment accuracy was measured as the percentage of positions in the tested alignment that were identical to the reference structure-based alignment). Correspondingly, the average model accuracy increased from 43 to 54% (the model accuracy was measured as the percentage of the C(alpha) atoms of the model that were within 5 A of the corresponding C(alpha) atoms in the superposed native structure). The present method also compares favorably with two of the most successful previously described methods, PSI-BLAST and SAM. The accuracy of the final models would be increased further if a better method for ranking of the models were available. PMID:12853614

  7. The proteins of exocytosis: lessons from the sperm model.

    PubMed

    Tomes, Claudia Nora

    2015-02-01

    Exocytosis is a highly regulated process that consists of multiple functionally, kinetically and/or morphologically definable stages such as recruitment, targeting, tethering and docking of secretory vesicles with the plasma membrane, priming of the fusion machinery and calcium-triggered membrane fusion. After fusion, the membrane around the secretory vesicle is incorporated into the plasma membrane and the granule releases its contents. The proteins involved in these processes belong to several highly conserved families: Rab GTPases, SNAREs (soluble NSF-attachment protein receptors), ?-SNAP (?-NSF attachment protein), NSF (N-ethylmaleimide-sensitive factor), Munc13 and -18, complexins and synaptotagmins. In the present article, the molecules of exocytosis are reviewed, using human sperm as a model system. Sperm exocytosis is driven by isoforms of the same proteinaceous fusion machinery mentioned above, with their functions orchestrated in a hierarchically organized and unidirectional signalling cascade. In addition to the universal exocytosis regulator calcium, this cascade includes other second messengers such as diacylglycerol, inositol 1,4,5-trisphosphate and cAMP, as well as the enzymes that synthesize them and their target proteins. Of special interest is the cAMP-binding protein Epac (exchange protein directly activated by cAMP) due in part to its enzymatic activity towards Rap. The activation of Epac and Rap leads to a highly localized calcium signal which, together with assembly of the SNARE complex, governs the final stages of exocytosis. The source of this releasable calcium is the secretory granule itself. PMID:25609177

  8. Differential effects of dietary flaxseed protein and soy protein on plasma triglyceride and uric acid levels in animal models

    Microsoft Academic Search

    Sam J Bhathena; Ali A Ali; Ali I Mohamed; Carl T Hansen; Manuel T Velasquez

    2002-01-01

    The effect of dietary soy protein and flaxseed meal on metabolic parameters was studied in two animal models, F344 rats with normal lipid levels and obese SHR\\/N-cp rats with elevated levels of cholesterol and triglyceride. The rats were fed AIN 93 diet differing only in the source of protein. The rats were fed either 20% casein, 20% soy protein or

  9. Effect of Electrostatic Interaction on the Adsorption of Globular Proteins on Octacalcium Phosphate Crystal Film

    Microsoft Academic Search

    Woo-Kul Lee; Jea-Seung Ko; Hyun-Man Kim

    2002-01-01

    The electrostatic effect on the adsorption of globular proteins, such as bovine serum albumin (BSA), hen egg white lysozyme (LZM), and ?-lactoglobulin (?-Lg), on octacalcium phosphate (OCP)-like crystal thin films was investigated. A poorly crystalline thin film was synthesized on a tissue culture polystyrene (TCP) surface and used as a model surface in this study. The solution pH clearly affected

  10. A novel microgel and associated post-fabrication encapsulation technique of proteins

    Microsoft Academic Search

    Ying Zhang; Wen Zhu; Biaobing Wang; Jiandong Ding

    2005-01-01

    A novel negatively thermo-sensitive and biodegradable microgel was prepared by combination of macromer synthesis and inverse suspension polymerization. A new post-fabrication encapsulation technique based upon this kind of intelligent microgel was developed. Model proteins (hemoglobin, bovine serum albumin and insulin) were encapsulated into the microgels at 4 °C and released in vitro at 37 °C. Relatively high loading levels and

  11. Improving protein structure prediction with model-based search

    Microsoft Academic Search

    T. J. Brunette; Oliver Brock

    2005-01-01

    Motivation: De novo protein structure prediction can be formu- lated as search in a high-dimensional space. One of the most frequently used computational tools to solve such search pro- blems is the Monte Carlo method. We present a novel search technique, called model-based search. This method samples the high-dimensional search space to build an approximate model of the underlying function.

  12. Evaluation of the combined use of the recombinant Brucella abortus Omp10, Omp19 and Omp28 proteins for the clinical diagnosis of bovine brucellosis.

    PubMed

    Simborio, Hannah Leah Tadeja; Lee, Jin Ju; Bernardo Reyes, Alisha Wehdnesday; Hop, Huynh Tan; Arayan, Lauren Togonon; Min, Wongi; Lee, Hu Jang; Yoo, Han Sang; Kim, Suk

    2015-01-01

    Currently, there are several serodiagnostic tools available for brucellosis, however, it is difficult to differentiate an active infection from vaccination. Hence, there is a great need to develop alternative means that can distinguish between these two conditions without utilizing lipopolysaccharide (LPS). This study was an attempt to determine the efficacy of combined recombinant Brucella (B.) abortus outer membrane proteins (rOmps) and individual rOmps in the serodiagnosis of brucellosis by enzyme linked immunosorbent assay (ELISA), utilizing both that standard tube agglutination test (TAT)-positive and -negative serum samples from Korean native cattle. The results are very interesting and promising because the combined rOmp antigens used in the study were highly reactive with the TAT-positive serum samples. The combined rOmps sensitivity, specificity and accuracy were 215/232 (92.67%), 294/298 (98.66%) and 509/530 (96.04%), respectively. While these results are preliminary, the tests performed have very high potential in the serodiagnosis of brucellosis and likewise, the combined rOmps can be used for future vaccine production. PMID:25988974

  13. Thermodynamics of Protein Folding from Coarse-Grained Models' Perspectives

    E-print Network

    Michael Bachmann; Wolfhard Janke

    2007-10-25

    Folding and aggregation of proteins, the interaction between proteins and membranes, as well as the adsorption of organic soft matter to inorganic solid substrates belong to the most interesting challenges in understanding structure and function of complex macromolecules. This is reasoned by the interdisciplinary character of the associated questions ranging from the molecular origin of the loss of biological functionality as, for example, in Alzheimer's disease to the development of organic circuits for biosensory applications. In this lecture, we focus on the analysis of mesoscopic models for protein folding, aggregation, and hybrid systems of soft and solid condensed matter. The simplicity of the coarse-grained models allows for a more universal description of the notoriously difficult problem of protein folding. In this approach, classifications of structure formation processes with respect to the conformational pseudophases are possible. This is similar in aggregation and adsorption processes, where the individual folding propensity is influenced by external forces. The main problem in studies of conformational transitions is that the sequences of amino acids proteins are built up of are necessarily of finite length and, therefore, a thermodynamic limit does not exist. Thus, structural transitions are not phase transitions in the strict thermodynamic sense and the analysis of pseudouniversal aspects is intricate, as apparently small-system effects accompany all conformational transitions and cannot be neglected.

  14. A Collaboration Network Model Of Cytokine-Protein Network

    NASA Astrophysics Data System (ADS)

    Zou, Sheng-Rong; Zhou, Ta; Peng, Yu-Jing; Guo, Zhong-Wei; Gu, Chang-Gui; He, Da-Ren

    2008-03-01

    Complex networks provide us a new view for investigation of immune systems. We collect data through STRING database and present a network description with cooperation network model. The cytokine-protein network model we consider is constituted by two kinds of nodes, one is immune cytokine types which can be regarded as collaboration acts, the other one is protein type which can be regarded as collaboration actors. From act degree distribution that can be well described by typical SPL (shifted power law) functions [1], we find that HRAS, TNFRSF13C, S100A8, S100A1, MAPK8, S100A7, LIF, CCL4, CXCL13 are highly collaborated with other proteins. It reveals that these mediators are important in cytokine-protein network to regulate immune activity. Dyad in the collaboration networks can be defined as two proteins and they appear in one cytokine collaboration relationship. The dyad act degree distribution can also be well described by typical SPL functions. [1] Assortativity and act degree distribution of some collaboration networks, Hui Chang, Bei-Bei Su, Yue-Ping Zhou, Daren He, Physica A, 383 (2007) 687-702

  15. The effect of the number of observations used for Fourier transform infrared model calibration for bovine milk fat composition on the estimated genetic parameters of the predicted data.

    PubMed

    Rutten, M J M; Bovenhuis, H; van Arendonk, J A M

    2010-10-01

    Fourier transform infrared spectroscopy is a suitable method to determine bovine milk fat composition. However, the determination of fat composition by gas chromatography, required for calibration of the infrared prediction model, is expensive and labor intensive. It has recently been shown that the number of calibration samples is strongly related to the model's validation r(2) (i.e., accuracy of prediction). However, the effect of the number of calibration samples used, and therefore validation r(2), on the estimated genetic parameters of data predicted using the model needs to be established. To this end, 235 calibration data subsets of different sizes were sampled: n=100, n=250, n=500, and n=1,000 calibration samples. Subsequently, these data subsets were used to calibrate fat composition prediction models for 2 specific fatty acids: C16:0 and C18u (where u=unsaturated). Next, genetic parameters were estimated on predicted fat composition data for these fatty acids. Strong relationships between the number of calibration samples and validation r(2), as well as strong genetic correlations were found. However, the use of n=100 calibration samples resulted in a broad range of validation r(2) values and genetic correlations. Subsequent increases of the number of calibration samples resulted in narrowing patterns for validation r(2) as well as genetic correlations. The use of n=1,000 calibration samples resulted in estimated genetic correlations varying within a range of 0.10 around the average, which seems acceptable. Genetic analyses for the human health-related fatty acids C14:0, C16:0, and C18u, and the ratio of saturated fatty acids to unsaturated fatty acids showed that replacing observations on fat composition determined by gas chromatography by predictions based on infrared spectra reduced the potential genetic gain to 98, 86, 96, and 99% for the 4 fatty acid traits, respectively, in dairy breeding schemes where progeny testing is practiced. We conclude that a relatively large number of calibration samples is required to be able to obtain genetic correlations that lie within a limited range. Considering that the routine recording of infrared spectra is relatively cheap and straightforward, we concluded that this methodology provides an excellent means for the dairy industry to genetically alter milk fat composition. PMID:20855022

  16. An Unusual Intrinsically Disordered Protein from the Model Legume Lotus japonicus Stabilizes Proteins in Vitro*

    PubMed Central

    Haaning, Svend; Radutoiu, Simona; Hoffmann, Søren V.; Dittmer, Jens; Giehm, Lise; Otzen, Daniel E.; Stougaard, Jens

    2008-01-01

    Intrinsic structural disorder is a prevalent feature of proteins with chaperone activity. Using a complementary set of techniques, we have structurally characterized LjIDP1 (intrinsically disordered protein 1) from the model legume Lotus japonicus, and our results provide the first structural characterization of a member of the Lea5 protein family (PF03242). Contrary to in silico predictions, we show that LjIDP1 is intrinsically disordered and probably exists as an ensemble of conformations with limited residual ?-sheet, turn/loop, and polyproline II secondary structure. Furthermore, we show that LjIDP1 has an inherent propensity to undergo a large conformational shift, adopting a largely ?-helical structure when it is dehydrated and in the presence of different detergents and alcohols. This is consistent with an overrepresentation of order-promoting residues in LjIDP1 compared with the average of intrinsically disordered proteins. In line with functioning as a chaperone, we show that LjIDP1 effectively prevents inactivation of two model enzymes under conditions that promote protein misfolding and aggregation. The LjIdp1 gene is expressed in all L. japonicus tissues tested. A higher expression level was found in the root tip proximal zone, in roots inoculated with compatible endosymbiotic M. loti, and in functional nitrogen-fixing root nodules. We suggest that the ability of LjIDP1 to prevent protein misfolding and aggregation may play a significant role in tissues, such as symbiotic root nodules, which are characterized by high metabolic activity. PMID:18779323

  17. Cytotoxic Activity of Coagulase-Negative Staphylococci in Bovine Mastitis

    Microsoft Academic Search

    SONGLIN ZHANG; CAROL W. MADDOX

    2000-01-01

    Secreted toxins play important roles in the pathogenesis of bacterial infections. In this study, we examined the presence of secreted cytotoxic factors of coagulase-negative staphylococci (CoNS) from bovine clinical and subclinical mastitis. A 34- to 36-kDa protein with cell-rounding cytotoxic activity was found in many CoNS strains, especially in Staphylococcus chromogenes strains. The protein caused cell detachment and cell rounding

  18. Homology modelling and analysis of structure predictions of the bovine rhinitis B virus RNA-dependent RNA polymerase (RdRp)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine Rhinitis B Viruses (BRBV) are picornaviruses responsible for mild respiratory infection of cattle and probably the least characterized member of the Aphthoviruses. BRBV is the closest relative known to Foot and Mouth Disease virus (FMDV) with around a 43 percent identical polyprotein sequenc...

  19. Infectious bovine keratoconjunctivitis (pinkeye).

    PubMed

    Angelos, John A

    2015-03-01

    As is the case for controlling other infectious livestock diseases, the most successful efforts to control infectious bovine keratoconjunctivitis (IBK) will include consideration of the host, the environment, herd management, and ongoing surveillance even after the immediate crisis has passed. Research over many years has led to the discovery of a variety of antibiotic treatments and antibiotic regimens that can be effective against IBK. The discoveries of Mor bovoculi and reports of IBK associated with Mycoplasma spp without concurrent Mor bovis or Mor bovoculi have raised new questions into the roles that other organisms may play in IBK pathogenesis. PMID:25576389

  20. Large-scale protein-protein interactions detection by integrating big biosensing data with computational model.

    PubMed

    You, Zhu-Hong; Li, Shuai; Gao, Xin; Luo, Xin; Ji, Zhen

    2014-01-01

    Protein-protein interactions are the basis of biological functions, and studying these interactions on a molecular level is of crucial importance for understanding the functionality of a living cell. During the past decade, biosensors have emerged as an important tool for the high-throughput identification of proteins and their interactions. However, the high-throughput experimental methods for identifying PPIs are both time-consuming and expensive. On the other hand, high-throughput PPI data are often associated with high false-positive and high false-negative rates. Targeting at these problems, we propose a method for PPI detection by integrating biosensor-based PPI data with a novel computational model. This method was developed based on the algorithm of extreme learning machine combined with a novel representation of protein sequence descriptor. When performed on the large-scale human protein interaction dataset, the proposed method achieved 84.8% prediction accuracy with 84.08% sensitivity at the specificity of 85.53%. We conducted more extensive experiments to compare the proposed method with the state-of-the-art techniques, support vector machine. The achieved results demonstrate that our approach is very promising for detecting new PPIs, and it can be a helpful supplement for biosensor-based PPI data detection. PMID:25215285

  1. Polyether sulfone/hydroxyapatite mixed matrix membranes for protein purification

    NASA Astrophysics Data System (ADS)

    Sun, Junfen; Wu, Lishun

    2014-07-01

    This work proposes a novel approach for protein purification from solution using mixed matrix membranes (MMMs) comprising of hydroxyapatite (HAP) inside polyether sulfone (PES) matrix. The influence of HAP particle loading on membrane morphology is studied. The MMMs are further characterized concerning permeability and adsorption capacity. The MMMs show purification of protein via both diffusion as well as adsorption, and show the potential of using MMMs for improvements in protein purification techniques. The bovine serum albumin (BSA) was used as a model protein. The properties and structures of MMMs prepared by immersion phase separation process were characterized by pure water flux, BSA adsorption and scanning electron microscopy (SEM).

  2. Animal models of protein allergenicity: potential benefits, pitfalls and challenges.

    PubMed

    Dearman, R J; Kimber, I

    2009-04-01

    Food allergy is an important health issue. With an increasing interest in novel foods derived from transgenic crop plants, there is a growing need for the development of approaches suitable for the characterization of the allergenic potential of proteins. There are methods available currently (such as homology searches and serological testing) that are very effective at identifying proteins that are likely to cross-react with known allergens. However, animal models may play a role in the identification of truly novel proteins, such as bacterial or fungal proteins, that have not been experienced previously in the diet. We consider here the potential benefits, pitfalls and challenges of the selection of various animal models, including the mouse, the rat, the dog and the neonatal swine. The advantages and disadvantages of various experimental end-points are discussed, including the measurement of specific IgE by ELISA, Western blotting or functional tests such as the passive cutaneous anaphylaxis assay, and the assessment of challenge-induced clinical symptoms in previously sensitized animals. The experimental variables of route of exposure to test proteins and the incorporation of adjuvant to increase the sensitivity of the responses are considered also. It is important to emphasize that currently none of these approaches has been validated for the purposes of hazard identification in the context of a safety assessment. However, the available evidence suggests that the judicious use of an accurate and robust animal model could provide important additional data that would contribute significantly to the assessment of the potential allergenicity of novel proteins. PMID:19178540

  3. Protein encapsulation in polymeric microneedles by photolithography

    PubMed Central

    Kochhar, Jaspreet Singh; Zou, Shui; Chan, Sui Yung; Kang, Lifeng

    2012-01-01

    Background Recent interest in biocompatible polymeric microneedles for the delivery of biomolecules has propelled considerable interest in fabrication of microneedles. It is important that the fabrication process is feasible for drug encapsulation and compatible with the stability of the drug in question. Moreover, drug encapsulation may offer the advantage of higher drug loading compared with other technologies, such as drug coating. Methods and results In this study, we encapsulated a model protein drug, namely, bovine serum albumin, in polymeric microneedles by photolithography. Drug distribution within the microneedle array was found to be uniform. The encapsulated protein retained its primary, secondary, and tertiary structural characteristics. In vitro release of the encapsulated protein showed that almost all of the drug was released into phosphate buffered saline within 6 hours. The in vitro permeation profile of encapsulated bovine serum albumin through rat skin was also tested and shown to resemble the in vitro release profile, with an initial release burst followed by a slow release phase. The cytotoxicity of the microneedles without bovine serum albumin was tested in three different cell lines. High cell viabilities were observed, demonstrating the innocuous nature of the microneedles. Conclusion The microneedle array can potentially serve as a useful drug carrier for proteins, peptides, and vaccines. PMID:22787403

  4. Potential applications for antiviral therapy and prophylaxis in bovine medicine.

    PubMed

    Newcomer, Benjamin W; Walz, Paul H; Givens, M Daniel

    2014-06-01

    Viral disease is one of the major causes of financial loss and animal suffering in today's cattle industry. Increases in global commerce and average herd size, urbanization, vertical integration within the industry and alterations in global climate patterns have allowed the spread of pathogenic viruses, or the introduction of new viral species, into regions previously free of such pathogens, creating the potential for widespread morbidity and mortality in naïve cattle populations. Despite this, no antiviral products are currently commercially licensed for use in bovine medicine, although significant progress has been made in the development of antivirals for use against bovine viral diarrhea virus (BVDV), foot and mouth disease virus (FMDV) and bovine herpesvirus (BHV). BVDV is extensively studied as a model virus for human antiviral studies. Consequently, many compounds with efficacy have been identified and a few have been successfully used to prevent infection in vivo although commercial development is still lacking. FMDV is also the subject of extensive antiviral testing due to the importance of outbreak containment for maintenance of export markets. Thirdly, BHV presents an attractive target for antiviral development due to its worldwide presence. Antiviral studies for other bovine viral pathogens are largely limited to preliminary studies. This review summarizes the current state of knowledge of antiviral compounds against several key bovine pathogens and the potential for commercial antiviral applications in the prevention and control of several selected bovine diseases. PMID:24810855

  5. Identification, purification, and characterization of subunits of cAMP-dependent protein kinase in human testis. Reverse mobilities of human RII alpha and RII beta on sodium dodecyl sulfate-polyacrylamide gel electrophoresis compared with rat and bovine RIIs.

    PubMed

    Skålhegg, B S; Landmark, B; Foss, K B; Lohmann, S M; Hansson, V; Lea, T; Jahnsen, T

    1992-03-15

    We have previously identified and characterized regulatory (R) subunits of cyclic AMP-dependent protein kinase, particularly the RII subunits in rat tissues (Jahnsen, T., Lohmann, S. M., Walter, U., Hedin, L., and Richards, J. S. (1985) J. Biol. Chem. 260, 15980-15987; Jahnsen, T., Hedin, L., Lohmann, S. M., Walter, U., and Richards, J. S. (1986) J. Biol. Chem. 261, 6637-6639; Jahnsen, T., Hedin, L., Kidd, V. J., Beattie, W. G., Lohmann, S. M., Walter, U., Durica, J., Schulz, T. Z., Schiltz, E., Browner, M., Lawrence, C. B., Goldman, D., Ratoosh, S. L., and Richards, J. S. (1986) J. Biol. Chem. 261, 12352-12361). These studies showed that rat RII alpha and RII beta had apparent molecular masses of 54 and 52 kDa, respectively. The aim of the present study was to purify and characterize cAMP-dependent protein kinase R subunits in human testis and to examine which of the subunits (mRNAs and proteins) are present in this tissue. Our results show that human testis contains mRNAs for five out of the seven known subunits of cAMP-dependent protein kinase. We observed strong expression of mRNAs for RI alpha (1.5 and 3.2 kilobases (kb)), RII alpha (2.2, 2.4, and 7.0 kb), and RII beta (3.3 kb). We also demonstrated mRNAs for two of the three catalytic subunits, C alpha (2.7 kb) and C gamma (1.7 kb). Purification of R subunits by DEAE-cellulose and cAMP affinity chromatography revealed three distinct forms with apparent molecular masses of 49, 51, and 53 kDa, respectively. Characterization of these R subunits by their 8-azido-cAMP photoaffinity labeling and immunoreactivity, as well as by a phosphorylation-dependent mobility shift on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), indicated subunit sizes of RII beta (53 kDa) greater than RII alpha dephosphoform (51 kDa) greater than RI alpha (49 kDa). This conclusion was verified by the analysis of RII subunits produced by in vitro transcription/translation of full-length cDNAs for both human RII alpha and RII beta in wheat germ lysates. The in vitro translated products were the same size as the purified human testis subunits, and only the smallest RII subunit (RII alpha) revealed a distinct mobility shift on SDS-PAGE after phosphorylation/dephosphorylation. This study supports the conclusion that the mobilities of human RII subunits (RII alpha, RII beta) on SDS-PAGE are reversed in contrast with those of other species such as rat and bovine.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:1544918

  6. A Simple Protein Synthesis Model for the PURE System Operation.

    PubMed

    Mavelli, Fabio; Marangoni, Roberto; Stano, Pasquale

    2015-06-01

    The encapsulation of transcription-translation (TX-TL) cell-free machinery inside lipid vesicles (liposomes) is a key element in synthetic cell technology. The PURE system is a TX-TL kit composed of well-characterized parts, whose concentrations are fine tunable, which works according to a modular architecture. For these reasons, the PURE system perfectly fulfils the requirements of synthetic biology and is widely used for constructing synthetic cells. In this work, we present a simplified mathematical model to simulate the PURE system operations. Based on Michaelis-Menten kinetics and differential equations, the model describes protein synthesis dynamics by using 9 chemical species, 6 reactions and 16 kinetic parameters. The model correctly predicts the time course for messenger RNA and protein production and allows quantitative predictions. By means of this model, it is possible to foresee how the PURE system species affect the mechanism of proteins synthesis and therefore help in understanding scenarios where the concentration of the PURE system components has been modified purposely or as a result of stochastic fluctuations (for example after random encapsulation inside vesicles). The model also makes the determination of response coefficients for all species involved in the TX-TL mechanism possible and allows for scrutiny on how chemical energy is consumed by the three PURE system modules (transcription, translation and aminoacylation). PMID:25911591

  7. Bioanalytical methods for the metalloproteomics study of bovine longissimus thoracis muscle tissue with different grades of meat tenderness in the Nellore breed (Bos indicus).

    PubMed

    Baldassini, Welder Angelo; Braga, Camila Pereira; Chardulo, Luis Artur Loyola; Vasconcelos Silva, Josineudson Augusto; Malheiros, Jessica Moraes; de Albuquerque, Lúcia Galvão; Fernandes, Talita Tanaka; Padilha, Pedro de Magalhães

    2015-02-15

    The work describes a metalloproteomics study of bovine muscle tissue with different grades of meat tenderness from animals of the Nellore breed (Bos indicus) based on protein separation by two-dimensional gel electrophoresis, the identification of calcium ions in protein spots by X-ray fluorescence (SR-XRF) and the characterisation of proteins by electrospray ionisation mass spectrometry. Forty (40) specimens were selected and divided into two experimental groups: animals with tough meat (TO) and animals with tender meat (TE). A third group (P) of Piedmontese breed animals (Bos taurus) was included to serve as a comparative model for the level of meat tenderness. The procedures were efficient and preserved the metal-protein structure, enabling calcium detection in protein spots by SR-XRF at a given molecular weight range of 14-97kDa. Two proteins (pyruvate kinase and albumin) were inferred to be related to the phenotypical differences in animals from the different groups. PMID:25236199

  8. Designability, thermodynamic stability, and dynamics in protein folding: A lattice model study

    E-print Network

    Levine, Alex J.

    Designability, thermodynamic stability, and dynamics in protein folding: A lattice model study Re October 1998 In the framework of a lattice-model study of protein folding, we investigate the interplay model. Lattice models have been widely used in the study of protein folding dynamics.2­8 The main

  9. A minimum-reaction-flux solution to master-equation models of protein folding

    E-print Network

    Weston, Ken

    A minimum-reaction-flux solution to master-equation models of protein folding Huan-Xiang Zhoua; published online 20 May 2008 Master equations are widely used for modeling protein folding. Here- ceptual and quantitative models for protein folding.1­15 In such models, the conformational space

  10. HMMSTR: a Hidden Markov Model for Local Sequence-Structure Correlations in Proteins

    E-print Network

    Bystroff, Chris

    HMMSTR: a Hidden Markov Model for Local Sequence-Structure Correlations in Proteins Christopher describe a hidden Markov model, HMMSTR, for general protein sequence based on the I-sites library of sequence-structure motifs. Unlike the linear hidden Markov models used to model individual protein families

  11. Dynamics of Proteins in Crystals: Comparison of Experiment with Simple Models

    E-print Network

    Phillips, George N. Jr.

    -factor, Gaussian Network Model, normal modes, protein dynamics ABSTRACT The dynamic behavior of proteins to understand a protein's function, one must know about both its structure and dynamics. X-ray crystallographyDynamics of Proteins in Crystals: Comparison of Experiment with Simple Models Sibsankar Kundu1

  12. Genetic Algorithm for Predicting Protein Folding in the 2D HP Model

    E-print Network

    Emmerich, Michael

    Genetic Algorithm for Predicting Protein Folding in the 2D HP Model A Parameter Tuning Case Study of a protein, predicting its tertiary structure is known as the protein folding problem. This problem has been. The protein folding problem in the HP model is to find a conformation (a folded sequence) with the lowest

  13. Automatic protein structure prediction system enabling rapid and accurate model building for enzyme screening

    Microsoft Academic Search

    Joo-Hyun Seo; Gang-Seong Lee; Juhan Kim; Byung-Kwan Cho; Keehyoung Joo; Jooyoung Lee; Byung-Gee Kim

    2009-01-01

    Protein structure prediction has great potential of understanding the function of proteins at the molecular level and designing novel protein functions. Here, we report rapid and accurate structure prediction system running in an automated manner. Since fold recognition of the target protein to be modeled is the starting point of the template-guided model building process, various approaches – such as

  14. Assembly of Protein Structure From Sparse Experimental Data: An Efficient Monte Carlo Model

    E-print Network

    Skolnick, Jeff

    Kolinski*1,2 and Jeffrey Skolnick1 1Department of Molecular Biology, The Scripps Research Institute, La. Due to the simplic- ity of the protein representation and definition of the model force field:475­494, 1998. 1998 Wiley-Liss, Inc. Key words: protein assembly; protein struc- ture; protein reduced models

  15. Modeling the contribution of individual proteins to mixed skeletal muscle protein synthetic rates over increasing periods of label incorporation.

    PubMed

    Miller, Benjamin F; Wolff, Christopher A; Peelor, Fredrick F; Shipman, Patrick D; Hamilton, Karyn L

    2015-03-15

    Advances in stable isotope approaches, primarily the use of deuterium oxide ((2)H2O), allow for long-term measurements of protein synthesis, as well as the contribution of individual proteins to tissue measured protein synthesis rates. Here, we determined the influence of individual protein synthetic rates, individual protein content, and time of isotopic labeling on the measured synthesis rate of skeletal muscle proteins. To this end, we developed a mathematical model, applied the model to an established data set collected in vivo, and, to experimentally test the impact of different isotopic labeling periods, used (2)H2O to measure protein synthesis in cultured myotubes over periods of 2, 4, and 7 days. We first demonstrated the influence of both relative protein content and individual protein synthesis rates on measured synthesis rates over time. When expanded to include 286 individual proteins, the model closely approximated protein synthetic rates measured in vivo. The model revealed a 29% difference in measured synthesis rates from the slowest period of measurement (20 min) to the longest period of measurement (6 wk). In support of these findings, culturing of C2C12 myotubes with isotopic labeling periods of 2, 4, or 7 days revealed up to a doubling of the measured synthesis rate in the shorter labeling period compared with the longer period of labeling. From our model, we conclude that a 4-wk period of labeling is ideal for considering all proteins in a mixed-tissue fraction, while minimizing the slowing effect of fully turned-over proteins. In addition, we advocate that careful consideration must be paid to the period of isotopic labeling when comparing mixed protein synthetic rates between studies. PMID:25593288

  16. Modeling protein misfolding in charcot-marie-tooth disease.

    PubMed

    Theocharopoulou, Georgia; Vlamos, Panayiotis

    2015-01-01

    Charcot-Marie-Tooth (CMT) disease is the most common inherited neuromuscular disorder. Recent advancements in molecular biology have elucidated the molecular bases of this genetically heterogeneous neuropathy. Still, the major challenge lies in determining the individual contributions by malfunctions of proteins to the disease's pathology. This paper reviews the identified molecular mechanisms underlying major forms of CMT disease. A growing body of evidence has highlighted the role of protein misfolding in demyelinating peripheral neuropathies and neurodegenerative diseases. Several hypotheses have been proposed to explain how misfolded aggregates induce neuronal damage. Current research focuses on developing novel therapeutic targets which aim to prevent, or even reverse the formation of protein aggregation. Interestingly, the role of the cellular defence mechanisms against accumulation of misfolded proteins may play a key role leading to novel strategies for treatment accelerating the clearance of their toxic early aggregates. Based on these findings we propose a model for describing in terms of a formal computer language, the biomolecular processes involving proteins associated with CMT disease. PMID:25417019

  17. Model discrimination and estimation of the intraparticle mass transfer parameters for the adsorption of bovine serum albumin onto porous adsorbent particles by the use of experimental frontal analysis data.

    PubMed

    Heeter, G A; Liapis, A I

    1997-07-25

    Experimental data from a chromatographic system involving the adsorption of bovine serum albumin (BSA) onto porous anion-exchange adsorbent particles packed in a column are presented. The parameters that characterize the mass transfer mechanisms of intraparticle diffusion and convection are estimated by fitting the predictions of dynamic mathematical models describing adsorption in column systems having spherical perfusive and purely diffusive adsorbent particles to the experimental breakthrough data obtained from the column adsorption system. Both linear and nonlinear expressions for the equilibrium isotherm are considered. The values of the transport parameters are estimated in the time domain for the nonlinear adsorption models and in the Laplace transform domain for the linear adsorption models. The capabilities of the different models to describe satisfactorily the dynamic behavior of the adsorption system are compared. The dynamic nonlinear adsorption model for purely diffusive particles is found to describe most appropriately the dynamic behavior of the experimental chromatographic system studied in this work. PMID:9286073

  18. [Modeling of protein spatial structure using tritium planigraphy].

    PubMed

    Bogacheva, E N; Bogachev, A N; Dmitriev, I B; Dolgov, A A; Chulichkov, A L; Shishkov, A V; Baratova, L A

    2011-01-01

    The results of proteins spatial structure modeling using the tritium planigraphy technique are presented. The knowledge of three-dimensional structure of macromolecules is extremely necessary to understand the basic mechanisms of interaction in biological systems and complex technological processes. Known limitations of the X-ray analysis (crystal state) and NMR (molecular weight) make it necessary to seek new approaches to modeling the spatial structure of proteins. Semiempirical tritium planigraphy technique is one of these approaches. The method is based on the bombardment of the object by beam of hot tritium atoms (E(at) > or = 0.3 eV) and a computer simulation experiment. On the example of proteins of the different structural classes we set that by using this integrated approach can be obtained by three-dimensional model of the structure, well consistent with the data of X-ray analysis. An important factor is a sequence search of contacts between secondary structure elements: the best fit model with the native structure is achieved by assembling the elements of a vector in the sequence from the N- to C-terminus of the polypeptide chain. PMID:22279745

  19. Affinity purification of aprotinin from bovine lung.

    PubMed

    Xin, Yu; Liu, Lanhua; Chen, Beizhan; Zhang, Ling; Tong, Yanjun

    2015-05-01

    An affinity protocol for the purification of aprotinin from bovine lung was developed. To simulate the structure of sucrose octasulfate, a natural specific probe for aprotinin, the affinity ligand was composed of an acidic head and a hydrophobic stick, and was then linked with Sepharose. The sorbent was then subjected to adsorption analysis with pure aprotinin. The purification process consisted of one step of affinity chromatography and another step of ultrafiltration. Then purified aprotinin was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis, trypsin inhibitor activity, gel-filtration, and thin-layer chromatography analysis. As calculated, the theoretical maximum adsorption (Qmax ) of the affinity sorbent was 25,476.0 ± 184.8 kallikrein inactivator unit/g wet gel; the dissociation constant of the complex "immobilized ligand-aprotinin" (Kd ) was 4.6 ± 0.1 kallikrein inactivator unit/mL. After the affinity separation of bovine lung aprotinin, reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis and gel-filtration chromatography revealed that the protein was a single polypeptide, and the purities were ? 97 and 100%, respectively; the purified peptide was also confirmed with aprotinin standard by gel-filtration chromatography and thin-layer chromatography. After the whole purification process, protein, and bioactivity recoveries were 2.2 and 92.6%, respectively; and the specific activity was up to 15,907.1 ± 10.2 kallikrein inactivator unit/mg. PMID:25677462

  20. Purification and electrophoretic characterization of bovine conglutinin.

    PubMed

    Dec, Marta; Wernicki, Andrzej; Puchalski, Andrzej; Urban-Chmiel, Renata; Gr?z, Marcin

    2012-06-01

    In this study a two-stage procedure for purification of conglutinin using affinity and ion-exchange chromatography was developed. To isolate conglutinin from bovine serum, its unique ability to bind to complement component iC3b was exploited. Incubation of bovine serum with chromatographic beads (TSK, Toyopearl HW-75 F) at 37 °C allows for iC3b deposition and subsequent binding of conglutinin. A single protein fraction eluted with ethylenediaminetetraacetic acid (EDTA) was then separated on an ion-exchange column in an NaCl gradient. The purification was evaluated by SDS-PAGE and western blotting. Conglutinin analyzed by SDS-PAGE under reducing conditions showed two main bands at 41 and 47 kDa and eight weaker bands. Nonreduced conglutinin appeared as a ladder pattern composed of many fractions ranging from 34 to 630 kDa. The bands at 34, 153, 174, 247, 338 and 387 kDa displayed the highest optical density. In the native conglutinin profile four fractions were observed, and the pI of this protein was below 8.5. The presence of sugar residues in the conglutinin molecule was detected using Schiff's reagent. PMID:21932412

  1. Modeling G Protein-Coupled Receptors: a Concrete Possibility

    PubMed Central

    Costanzi, Stefano

    2010-01-01

    G protein-coupled receptors (GPCRs) are a large superfamily of membrane bound signaling proteins that are involved in the regulation of a wide range of physiological functions and constitute the most common target for therapeutic intervention. Due to the paucity of crystal structures, homology modeling has become a widespread technique for the construction of GPCR models, which have been applied to the study of their structure-function relationships and to the identification of lead ligands through virtual screening. Rhodopsin has been for years the only available template. However, recent breakthroughs in GPCR crystallography have led to the solution of the structures of a few additional receptors. In light of these newly elucidated crystal structures, we have been able to produce a substantial amount of data to demonstrate that accurate models of GPCRs in complex with their ligands can be constructed through homology modeling followed by fully flexible molecular docking. These results have been confirmed by our success in the first blind assessment of GPCR modeling and docking, organized in coordination with the solution of the X-ray structure of the adenosine A2A receptor. Taken together, these data indicate that: a) the transmembrane helical bundle can be modeled with considerable accuracy; b) predicting the binding mode of a ligand, although doable, is challenging; c) modeling of the extracellular and intracellular loops is still problematic. PMID:21253444

  2. Analysis and Ranking of Protein-Protein Docking Models Using Inter-Residue Contacts and Inter-Molecular Contact Maps.

    PubMed

    Oliva, Romina; Chermak, Edrisse; Cavallo, Luigi

    2015-01-01

    In view of the increasing interest both in inhibitors of protein-protein interactions and in protein drugs themselves, analysis of the three-dimensional structure of protein-protein complexes is assuming greater relevance in drug design. In the many cases where an experimental structure is not available, protein-protein docking becomes the method of choice for predicting the arrangement of the complex. However, reliably scoring protein-protein docking poses is still an unsolved problem. As a consequence, the screening of many docking models is usually required in the analysis step, to possibly single out the correct ones. Here, making use of exemplary cases, we review our recently introduced methods for the analysis of protein complex structures and for the scoring of protein docking poses, based on the use of inter-residue contacts and their visualization in inter-molecular contact maps. We also show that the ensemble of tools we developed can be used in the context of rational drug design targeting protein-protein interactions. PMID:26140438

  3. Collective aspects of protein folding illustrated by a toy model

    SciTech Connect

    Stillinger, F.H. [AT& T Bell Laboratories, Murray Hill, New Jersey 07974 (United States)] [AT& T Bell Laboratories, Murray Hill, New Jersey 07974 (United States); Head-Gordon, T. [Life Sciences Division, Lawrence Berkeley Laboratory, University of California, Berkeley, California 94720 (United States)] [Life Sciences Division, Lawrence Berkeley Laboratory, University of California, Berkeley, California 94720 (United States)

    1995-09-01

    A simple toy model for polypeptides serves as a testbed to illuminate some nonlocal, or collective, aspects of protein folding phenomena. The model is two dimensional and has only two amino acids, but involves a continuous range of backbone bend angles. Global potential energy minima and their folding structures have been determined for leading members of two special and contrasting polypeptide sequences, center doped and Fibonacci, named descriptively for their primary structures. The results display the presence of spontaneous symmetry breaking, elastic strain, and substantial conformational variation for specific embedded amino acid strings. We conclude that collective variables generated by the primary amino acid structure may be required for fully effective protein folding predictors, including those based on neural networks.

  4. Protein self-diffusion in crowded solutions

    PubMed Central

    Roosen-Runge, Felix; Hennig, Marcus; Zhang, Fajun; Jacobs, Robert M. J.; Sztucki, Michael; Schober, Helmut; Seydel, Tilo; Schreiber, Frank

    2011-01-01

    Macromolecular crowding in biological media is an essential factor for cellular function. The interplay of intermolecular interactions at multiple time and length scales governs a fine-tuned system of reaction and transport processes, including particularly protein diffusion as a limiting or driving factor. Using quasielastic neutron backscattering, we probe the protein self-diffusion in crowded aqueous solutions of bovine serum albumin on nanosecond time and nanometer length scales employing the same protein as crowding agent. The measured diffusion coefficient D(?) strongly decreases with increasing protein volume fraction ? explored within 7% ? ? ? 30%. With an ellipsoidal protein model and an analytical framework involving colloid diffusion theory, we separate the rotational Dr(?) and translational Dt(?) contributions to D(?). The resulting Dt(?) is described by short-time self-diffusion of effective spheres. Protein self-diffusion at biological volume fractions is found to be slowed down to 20% of the dilute limit solely due to hydrodynamic interactions. PMID:21730176

  5. Redesigning Protein Cavities as a Strategy for Increasing Affinity in Protein-Protein Interaction: Interferon-? Receptor 1 as a Model

    PubMed Central

    Biedermannová, Lada; Mikulecký, Pavel; Zahradník, Ji?í; Charnavets, Tatsiana; Šebo, Peter

    2015-01-01

    Combining computational and experimental tools, we present a new strategy for designing high affinity variants of a binding protein. The affinity is increased by mutating residues not at the interface, but at positions lining internal cavities of one of the interacting molecules. Filling the cavities lowers flexibility of the binding protein, possibly reducing entropic penalty of binding. The approach was tested using the interferon-? receptor 1 (IFN?R1) complex with IFN? as a model. Mutations were selected from 52 amino acid positions lining the IFN?R1 internal cavities by using a protocol based on FoldX prediction of free energy changes. The final four mutations filling the IFN?R1 cavities and potentially improving the affinity to IFN? were expressed, purified, and refolded, and their affinity towards IFN? was measured by SPR. While individual cavity mutations yielded receptor constructs exhibiting only slight increase of affinity compared to WT, combinations of these mutations with previously characterized variant N96W led to a significant sevenfold increase. The affinity increase in the high affinity receptor variant N96W+V35L is linked to the restriction of its molecular fluctuations in the unbound state. The results demonstrate that mutating cavity residues is a viable strategy for designing protein variants with increased affinity.

  6. Viral infections and bovine mastitis: a review.

    PubMed

    Wellenberg, G J; van der Poel, W H M; Van Oirschot, J T

    2002-08-01

    This review deals with the role of viruses in the aetiology of bovine mastitis. Bovine herpesvirus 1, bovine herpesvirus 4, foot-and-mouth disease virus, and parainfluenza 3 virus have been isolated from milk from cows with clinical mastitis. Intramammary inoculations of bovine herpesvirus 1 or parainfluenza 3 virus-induced clinical mastitis, while an intramammary inoculation of foot-and-mouth disease virus resulted in necrosis of the mammary gland. Subclinical mastitis has been induced after a simultaneous intramammary and intranasal inoculation of lactating cows with bovine herpesvirus 4. Bovine leukaemia virus has been detected in mammary tissue of cows with subclinical mastitis, but whether this virus was able to induce bovine mastitis has not been reported. Bovine herpesvirus 2, vaccinia, cowpox, pseudocowpox, vesicular stomatitis, foot-and-mouth disease viruses, and bovine papillomaviruses can play an indirect role in the aetiology of bovine mastitis. These viruses can induce teat lesions, for instance in the ductus papillaris, which result in a reduction of the natural defence mechanisms of the udder and indirectly in bovine mastitis due to bacterial pathogens. Bovine herpesvirus 1, bovine viral diarrhoea virus, bovine immunodeficiency virus, and bovine leukaemia virus infections may play an indirect role in bovine mastitis, due to their immunosuppressive properties. But, more research is warranted to underline their indirect role in bovine mastitis. We conclude that viral infections can play a direct or indirect role in the aetiology of bovine mastitis; therefore, their importance in the aetiology of bovine mastitis and their economical impact needs further attention. PMID:12119136

  7. MODBASE, a database of annotated comparative protein structure models, and associated resources

    Microsoft Academic Search

    Ursula Pieper; Narayanan Eswar; Hannes Braberg; Mallur S. Madhusudhan; Fred P. Davis; Ashley C. Stuart; Nebojsa Mirkovic; Andrea Rossi; Marc A. Martí-renom; András Fiser; Ben M. Webb; Daniel M. Greenblatt; Conrad C. Huang; Thomas E. Ferrin; Andrej Sali

    2004-01-01

    MODBASE (http:\\/\\/salilab.org\\/modbase) is a rela- tional database of annotated comparative protein structure models for all available protein sequences matched to at least one known protein structure. The models are calculated by MODPIPE, an auto- mated modeling pipeline that relies on the MODELLER package for fold assignment, sequence-structure alignment, model building and model assessment (http:\\/salilab.org\\/modeller). MODBASE uses the MySQL relational database

  8. Crucial stages of protein folding through a solvable model: Predicting target sites

    E-print Network

    Cecconi, Fabio

    Crucial stages of protein folding through a solvable model: Predicting target sites for enzyme. Keywords: Protein-folding modeling; prediction of key folding sites; HIV-1 protease; drug resistance One

  9. Characterizing the Freeze–Drying Behavior of Model Protein Formulations

    Microsoft Academic Search

    Lavinia M. Lewis; Robert E. Johnson; Megan E. Oldroyd; Saleem S. Ahmed; Liji Joseph; Ilie Saracovan; Sandipan Sinha

    2010-01-01

    The freeze–drying behavior of three model proteins, namely, lysozyme, BSA, and IgG, has been studied using a variety of techniques\\u000a under two different primary drying conditions (shelf temperatures of ?25°C and +25°C, respectively) in an amorphous formulation.\\u000a Manometric temperature measurements were used to characterize product temperature (T\\u000a pr), sublimation rates, and product resistance (R\\u000a p) during primary drying. Biophysical techniques

  10. Native Fluorescence and Mag-Indo-1Protein Interaction as Tools for Probing Unfolding and Refolding Sequences of the Bovine Serum Albumin Subdomain in the Presence of Guanidine Hydrochloride

    Microsoft Academic Search

    Pierre M. Viallet; Tuan Vo-Dinh; Anne-Cécile Ribou; Jean Vigo; Jean-Marie Salmon

    2000-01-01

    Changes in the fluorescence spectrum of tryptophans Trp 134 and Trp 212 in bovine serum albumin (BSA) and of Trp 214 of human serum albumin in the presence of the chaotropic agent guanidine hydrochloride (Gnd) were studied. A detailed analysis of the fluorescence spectrum of native BSA yielded the fluorescence spectrum for each tryptophan of BSA. Modifications in the binding

  11. Large offspring syndrome: a bovine model for the human loss-of-imprinting overgrowth syndrome Beckwith-Wiedemann.

    PubMed

    Chen, Zhiyuan; Robbins, Katherine Marie; Wells, Kevin Dale; Rivera, Rocío Melissa

    2013-06-01

    Beckwith-Wiedemann syndrome (BWS) is a human loss-of-imprinting syndrome primarily characterized by macrosomia, macroglossia, and abdominal wall defects. BWS has been associated with misregulation of two clusters of imprinted genes. Children conceived with the use of assisted reproductive technologies (ART) appear to have an increased incidence of BWS. As in humans, ART can also induce a similar overgrowth syndrome in ruminants which is referred to as large offspring syndrome (LOS). The main goal of our study is to determine if LOS shows similar loss-of-imprinting at loci known to be misregulated in BWS. To test this, Bos taurus indicus × Bos taurus taurus F1 hybrids were generated by artificial insemination (AI; control) or by ART. Seven of the 27 conceptuses in the ART group were in the > 97th percentile body weight when compared with controls. Further, other characteristics reported in BWS were observed in the ART group, such as large tongue, umbilical hernia, and ear malformations. KCNQ1OT1 (the most-often misregulated imprinted gene in BWS) was biallelically-expressed in various organs in two out of seven overgrown conceptuses from the ART group, but shows monoallelic expression in all tissues of the AI conceptuses. Furthermore, biallelic expression of KCNQ1OT1 is associated with loss of methylation at the KvDMR1 on the maternal allele and with downregulation of the maternally-expressed gene CDKN1C. In conclusion, our results show phenotypic and epigenetic similarities between LOS and BWS, and we propose the use of LOS as an animal model to investigate the etiology of BWS. PMID:23751783

  12. Isolation and multilineage differentiation of bovine bone marrow mesenchymal stem cells

    Microsoft Academic Search

    Darko Bosnakovski; Morimichi Mizuno; Gonhyung Kim; Satoshi Takagi; Masahiro Okumura; Toru Fujinaga

    2005-01-01

    The bone marrow harbors a population of mesenchymal stem cells (MSCs) that possess the potential to differentiate into bone, cartilage, and fat, and along other tissue pathways. To date, MSCs from various species have been studied. Despite the bovine experimental model being widely used in experiments in vivo and in vitro, only a limited amount of information regarding bovine MSCs

  13. Rapid detection of bovine viral diarrhea virus as surrogate of bioterrorism agents

    Microsoft Academic Search

    Zarini Muhammad-Tahir; Evangelyn C. Alocilja; Daniel L. Grooms

    2005-01-01

    Bovine viral diarrhea virus (BVDV) is a major pathogen of cattle that is chosen as a model for select agents associated with agricultural bioterrorism, such as foot and mouth disease virus. Bovine viral diarrhea virus causes early embryonic death, abortion, respiratory problems, and immune system dysfunction in cattle throughout the world. Due to the insidious nature and economic loses from

  14. A novel affinity disks for bovine serum albumin purification.

    PubMed

    Tuzmen, Nalan; Kalburcu, Tülden; Uygun, Deniz Akta?; Akgol, Sinan; Denizli, Adil

    2015-01-01

    The adsorption characteristics of bovine serum albumin (BSA) onto the supermacroporous poly(hydroxyethylmethacrylate)-Reactive Green 19 [p(HEMA)-RG] cryogel disks have been investigated in this paper. p(HEMA) cryogel disks were prepared by radical polymerization initiated by N,N,N',N'-tetramethylene diamine (TEMED) and ammonium persulfate (APS) pair in an ice bath. Reactive Green (RG) 19 was covalently attached to the p(HEMA) cryogel disks. These disks were used in BSA adsorption studies to interrogate the effects of pH, initial protein concentration, ionic strength, and temperature. BSA adsorption capacity of the p(HEMA)-RG cryogel disk was significantly improved after the incorporation of RG. Adsorption capacity reached a plateau value at about 0.8 mg/mL at pH 4.0. The amount of adsorbed BSA decreased from 37.7 to 13.9 mg/g with increasing NaCl concentration. The enthalpy of BSA adsorption onto the p(HEMA)-RG cryogel disk was calculated as -58.4 kJ/mol. The adsorption equilibrium isotherm was fitted well by the Freundlich model. BSA was desorbed from cryogel disks (over 90 %) using 0.5 M NaSCN, and the purity of desorbed BSA was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The experimental results showed that the p(HEMA)-RG cryogel disks have potential for the quick protein separation and purification process. PMID:25308615

  15. Generic Coarse-Grained Model for Protein Folding and Aggregation

    NASA Astrophysics Data System (ADS)

    Bereau, Tristan; Deserno, Markus

    2009-03-01

    The complexity involved in protein structure is not only due to the rich variety of amino acids, but also the inherent weak interactions, comparable to thermal energy, and important cooperative phenomena. This presents a challenge in atomistic simulations, as it is associated with high-dimensionality and ruggedness of the energy landscape as well as long equilibration times. We have recently developed a coarse-grained (CG) implicit solvent peptide model which has been designed to reproduce key consequences of the abovementioned weak interactions. Its intermediate level of resolution, four beads per amino acid, allows for accurate sampling of local conformations by designing a force field that relies on simple interactions. A realistic ratio of ?-helix to ?-sheet content is achieved by mimicking a nearest-neighbor dipole interaction. We tune the model in order to fold helical proteins while systematically comparing the structure with NMR data. Very good agreement is achieved for proteins that have simple tertiary structures. We further probe the effects of cooperativity between amino acids by looking at peptide aggregation, where hydrophobic peptide fragments cooperatively form large-scale ?-sheet structures. The model is able to reproduce features from atomistic simulations on a qualitative basis.

  16. Molecular size of bovine lipoprotein lipase as determined by radiation inactivation

    SciTech Connect

    Olivecrona, T.; Bengtsson-Olivecrona, G.; Osborne, J.C. Jr.; Kempner, E.S.

    1985-06-10

    The authors have determined the size of the functional unit of bovine lipoprotein lipase by radiation inactivation. This was done in five different situations: 1) in a buffer with high salt concentration. In this situation the enzyme is relatively soluble and stable. 2) For an enzyme-heparin complex. This may reflect the physiological state of the enzyme at the vascular endothelium, where it is believed to be bound to a heparin-like molecule. 3) In the presence of lipid substrate and 4) with lipid substrate and activator protein. Here most of the enzyme is adsorbed to the substrate droplets. 5) For an enzyme-detergent complex; another model for enzyme-lipid interaction. In all five situations the enzyme activity decayed as an exponential function of radiation dose, and the target sizes were similar. The target size did not vary with the concentration of lipase protein. The combined data for bovine lipoprotein lipase yield a functional size of 72 kDa which is close to that expected for a dimer, 77 kDa.

  17. Texas Bovine Trichomoniasis Control Program 

    E-print Network

    Machen, Richard V.; Gill, Ronald J.; Faries Jr., Floron C.; Hairgrove, Thomas B.

    2009-04-30

    This publication summarizes the disease and transmission characteristics of bovine trichomoniasis in bulls and cows. It also outlines new measures being implemented in Texas to control the spread of the disease....

  18. Cartilage (Bovine and Shark) (PDQ®)

    Cancer.gov

    Expert-reviewed information summary about the use of bovine and shark cartilage as a treatment for people with cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  19. Antithrombotic activity of protein S infused without activated protein C in a baboon thrombosis model

    PubMed Central

    Heeb, Mary J.; Marzec, Ulla; Gruber, Andras; Hanson, Stephen R.

    2012-01-01

    Summary Protein S (ProS) is an essential plasma protein that enhances the anticoagulant activity of activated protein C (APC). In vitro, purified native human Zn2+-containing ProS also exerts direct anticoagulant activity by inhibiting prothrombinase and extrinsic FXase activities independently of APC. We investigated antithrombotic effects of ProS infused without APC in a baboon shunt model of thrombogenesis that employs a device consisting of arterial and venous shear flow segments. In in vitro experiments, the Zn2+-containing human ProS used for the studies displayed >10-fold higher prothrombinase inhibitory activity and anticoagulant activity in tissue factor-stimulated plasma, and 4-fold higher inhibition of the intrinsic pathway than the Zn2+-deficient ProS used. In the thrombosis model, ProS (33 µg/min for 1 hr) or saline was infused locally; platelet and fibrin deposition in the shunt were measured over 2 hr. During experiments performed at 50 mL/min blood flow, Zn2+-containing ProS inhibited platelet deposition 73–96% in arterial-type flow segments and 90–99% in venous-type flow segments; Zn2+-deficient ProS inhibited platelet deposition 52% in arterial-type flow segments and 65–73% in venous-type flow segments. At 100 mL/min blood flow rate, Zn2+-containing ProS inhibited platelet deposition by 39% and 73% in the respective segments; Zn2+-deficient ProS inhibited platelet deposition by 5% and 0% in the respective segments. Zn2+-containing ProS suppressed fibrin deposition by 67–90%. Systemic APC-independent ProS activity was significantly increased and thrombin-antithrombin complex levels were significantly decreased after infusion of ProS. Thus, infused human Zn2+-containing ProS is antithrombotic in primates, and may have therapeutic potential even in protein C-deficient human patients. PMID:22370911

  20. Antithrombotic activity of protein S infused without activated protein C in a baboon thrombosis model.

    PubMed

    Heeb, M J; Marzec, U; Gruber, A; Hanson, S R

    2012-04-01

    Protein S (ProS) is an essential plasma protein that enhances the anticoagulant activity of activated protein C (APC). In vitro , purified native human Zn2+-containing ProS also exerts direct anticoagulant activity by inhibiting prothrombinase and extrinsic FXase activities independently of APC. We investigated antithrombotic effects of ProS infused without APC in a baboon shunt model of thrombogenesis that employs a device consisting of arterial and venous shear flow segments. In in vitro experiments, the Zn2+-containing human ProS used for the studies displayed >10-fold higher prothrombinase inhibitory activity and anticoagulant activity in tissue factor-stimulated plasma, and four-fold higher inhibition of the intrinsic pathway than the Zn2+-deficient ProS used. In the thrombosis model, ProS (33 ?g/minute for 1 hour) or saline was infused locally; platelet and fibrin deposition in the shunt were measured over 2 hours. During experiments performed at 50 ml/minute blood flow, Zn2+-containing ProS inhibited platelet deposition 73-96% in arterial-type flow segments and 90-99% in venous-type flow segments; Zn2+-deficient ProS inhibited platelet deposition 52% in arterial-type flow segments and 65-73% in venous-type flow segments. At 100 ml/min blood flow rate, Zn2+-containing ProS inhibited platelet deposition by 39% and 73% in the respective segments; Zn2+-deficient ProS inhibited platelet deposition by 5% and 0% in the respective segments. Zn2+-containing ProS suppressed fibrin deposition by 67-90%. Systemic APC-independent ProS activity was significantly increased and thrombin-antithrombin complex levels were significantly decreased after infusion of ProS. Thus, infused human Zn2+-containing ProS is antithrombotic in primates, and may have therapeutic potential even in protein C-deficient human patients. PMID:22370911

  1. Conditional Random Fields for Classification of Protein Families: An Alternative to Hidden Markov Models

    E-print Network

    on the basis of its amino acid sequence is frequently done via a probabilistic model, usually a hidden Markov model chosen to model a protein family has been the hidden Markov model, or HMM. 1 #12;2 THOMAS J reasons why hidden Markov models are not the most suitable class of probabilistic models for the protein

  2. Allostery without conformation change: modelling protein dynamics at multiple scales

    NASA Astrophysics Data System (ADS)

    McLeish, T. C. B.; Rodgers, T. L.; Wilson, M. R.

    2013-10-01

    The original ideas of Cooper and Dryden, that allosteric signalling can be induced between distant binding sites on proteins without any change in mean structural conformation, has proved to be a remarkably prescient insight into the rich structure of protein dynamics. It represents an alternative to the celebrated Monod-Wyman-Changeux mechanism and proposes that modulation of the amplitude of thermal fluctuations around a mean structure, rather than shifts in the structure itself, give rise to allostery in ligand binding. In a complementary approach to experiments on real proteins, here we take a theoretical route to identify the necessary structural components of this mechanism. By reviewing and extending an approach that moves from very coarse-grained to more detailed models, we show that, a fundamental requirement for a body supporting fluctuation-induced allostery is a strongly inhomogeneous elastic modulus. This requirement is reflected in many real proteins, where a good approximation of the elastic structure maps strongly coherent domains onto rigid blocks connected by more flexible interface regions.

  3. A New Algorithm for Protein Folding in the HP Model Alantha Newman

    E-print Network

    Newman, Alantha

    A New Algorithm for Protein Folding in the HP Model Alantha Newman #3; Abstract We consider the problem of protein folding in the HP model on the two-dimensional square lattice. This problem but not in the string) are present. The protein folding problem in the hydrophobic-hydrophilic (HP) model on the 2D

  4. Protein Folding in the Hydrophobic-Hydrophilic (HP) Model is NP-Complete

    E-print Network

    Istrail, Sorin

    Protein Folding in the Hydrophobic-Hydrophilic (HP) Model is NP-Complete Bonnie Berger* Tom Leightont Abstract One of the simplest and most popular biophysical mod- els of protein folding is the hydrophobic-hydrophilic (HP) model. The HP model abstracts the hydrophobic in- teraction in protein folding

  5. Simple Physical Models Connect Theory and Experiment in Protein Folding Kinetics

    E-print Network

    Morozov, Alexandre V.

    Simple Physical Models Connect Theory and Experiment in Protein Folding Kinetics Eric Alm1 underlying the protein-folding problem can be tested by developing and characterizing simple models that make prefactor for protein folding. Finally, we discuss the limitations of simple native-state-based models

  6. Performance of Ecient Minimization Algorithms as Applied to Models of Peptides and Proteins

    E-print Network

    Aluffi, Paolo

    Performance of EÆcient Minimization Algorithms as Applied to Models of Peptides and Proteins CÆciency. Keywords: energy minimization; cyclic peptides and proteins; implicit solvation models; truncated and quasi, as applied to peptide and protein models. Such a study is necessary because the performance of minimization

  7. How Good Are Simplified Models for Protein Structure Prediction?

    PubMed Central

    Newton, M. A. Hakim; Rashid, Mahmood A.; Pham, Duc Nghia; Sattar, Abdul

    2014-01-01

    Protein structure prediction (PSP) has been one of the most challenging problems in computational biology for several decades. The challenge is largely due to the complexity of the all-atomic details and the unknown nature of the energy function. Researchers have therefore used simplified energy models that consider interaction potentials only between the amino acid monomers in contact on discrete lattices. The restricted nature of the lattices and the energy models poses a twofold concern regarding the assessment of the models. Can a native or a very close structure be obtained when structures are mapped to lattices? Can the contact based energy models on discrete lattices guide the search towards the native structures? In this paper, we use the protein chain lattice fitting (PCLF) problem to address the first concern; we developed a constraint-based local search algorithm for the PCLF problem for cubic and face-centered cubic lattices and found very close lattice fits for the native structures. For the second concern, we use a number of techniques to sample the conformation space and find correlations between energy functions and root mean square deviation (RMSD) distance of the lattice-based structures with the native structures. Our analysis reveals weakness of several contact based energy models used that are popular in PSP. PMID:24876837

  8. A Bovine Pancreatic Enzyme Catalyzing the Conversion of Proinsulin to Insulin

    PubMed Central

    Yip, C. C.

    1971-01-01

    An enzyme that catalyzes the conversion of bovine proinsulin to insulin has been purified from a bovine pancreatic extract. The product of conversion was identified as insulin by aminoacid analysis and determination of carboxyl terminal aminoacid residues. The purified enzyme preparation showed one major protein band on polyacrylamide gel disc electrophoresis; it had a molecular weight of about 70,000 and an isoelectric point (pI) at a pH of 4.82. Images PMID:5288380

  9. Culturing of BHK-21 cells in a medium containing adult bovine serum and pituitary extract

    Microsoft Academic Search

    H. P. M. Strouken; J.-W. Oberink; A. Bantjes

    1994-01-01

    Summary To achieve a comparable protein and cell production by culturing of BHK-21 cells in monolayer, in a medium containing adult bovine serum (ABS) and in a medium containing fetal bovine serum (FBS), 1.5 times (v\\/v) more ABS than FBS has to be added. At a volume fraction in the medium of less than 2% ABS, no proliferation is observed.

  10. Structures of Metal Sites of Oxidized Bovine Heart Cytochrome c Oxidase at 2.8 Å

    Microsoft Academic Search

    Tomitake Tsukihara; Hiroshi Aoyama; Eiki Yamashita; Takashi Tomizaki; Hiroshi Yamaguchi; Kyoko Shinzawa-Itoh; Ryosuke Nakashima; Reiko Yaono; Shinya Yoshikawa

    1995-01-01

    The high resolution three-dimensional x-ray structure of the metal sites of bovine heart cytochrome c oxidase is reported. Cytochrome c oxidase is the largest membrane protein yet crystallized and analyzed at atomic resolution. Electron density distribution of the oxidized bovine cytochrome c oxidase at 2.8 Å resolution indicates a dinuclear copper center with an unexpected structure similar to a [2Fe-2S]-type

  11. Polypeptide Transforming Growth Factors Isolated from Bovine Sources and Used for Wound Healing in vivo

    NASA Astrophysics Data System (ADS)

    Sporn, Michael B.; Roberts, Anita B.; Shull, James H.; Smith, Joseph M.; Ward, Jerrold M.; Sodek, Jaro

    1983-03-01

    Transforming growth factors, which are polypeptides that induce the transformed phenotype in nonneoplastic cells, have been isolated in bulk amounts from bovine salivary gland and kidney. In experiments in which wound healing chambers were implanted subcutaneously in the backs of rats, these bovine transforming growth factors accelerated the accumulation of total protein, collagen, and DNA in treated chambers. These studies thus show an effect of an isolated transforming growth factor in vivo.

  12. Cloning of bovine estrogen receptor beta (ER?): expression of novel deleted isoforms in reproductive tissues

    Microsoft Academic Search

    Norbert Walther; Chrisostomos Lioutas; Gina Tillmann; Richard Ivell

    1999-01-01

    cDNAs coding for bovine estrogen receptor beta (ER?) isoforms were cloned from bovine granulosa cells using a combination of several RT-PCR strategies. The cloned full-length receptor contains an open reading frame of 474 amino acids encoding a protein with high homology to the ER? sequences from other species. A second isoform nearly totally lacking the ligand binding domain was cloned

  13. Suppression of T-cell activation by Lactobacillus rhamnosus GG-degraded bovine casein

    Microsoft Academic Search

    Tanja Pessi; Erika Isolauri; Yelda Sütas; Hannu Kankaanranta; Eeva Moilanen; Mikko Hurme

    2001-01-01

    Earlier data indicate that Lactobacillus rhamnosus GG ATCC 53103 (L. GG), a commensal intestinal bacterial strain, promotes the degradation of proteins in the gut in vivo, and bovine casein hydrolysed with L. GG-derived proteases suppresses lymphocyte proliferation in vitro. The present study aimed to evaluate the effect of L. GG-degraded bovine casein on T-cell activation, i.e. IL-2 mRNA expression and

  14. Relaxation of backbone bond geometry improves protein energy landscape modeling.

    PubMed

    Conway, Patrick; Tyka, Michael D; DiMaio, Frank; Konerding, David E; Baker, David

    2014-01-01

    A key issue in macromolecular structure modeling is the granularity of the molecular representation. A fine-grained representation can approximate the actual structure more accurately, but may require many more degrees of freedom than a coarse-grained representation and hence make conformational search more challenging. We investigate this tradeoff between the accuracy and the size of protein conformational search space for two frequently used representations: one with fixed bond angles and lengths and one that has full flexibility. We performed large-scale explorations of the energy landscapes of 82 protein domains under each model, and find that the introduction of bond angle flexibility significantly increases the average energy gap between native and non-native structures. We also find that incorporating bonded geometry flexibility improves low resolution X-ray crystallographic refinement. These results suggest that backbone bond angle relaxation makes an important contribution to native structure energetics, that current energy functions are sufficiently accurate to capture the energetic gain associated with subtle deformations from chain ideality, and more speculatively, that backbone geometry distortions occur late in protein folding to optimize packing in the native state. PMID:24265211

  15. Engineering a model protein cavity to catalyze the Kemp elimination.

    PubMed

    Merski, Matthew; Shoichet, Brian K

    2012-10-01

    Synthetic cavitands and protein cavities have been widely studied as models for ligand recognition. Here we investigate the Met102 ? His substitution in the artificial L99A cavity in T4 lysozyme as a Kemp eliminase. The resulting enzyme had k(cat)/K(M) = 0.43 M(-1) s(-1) and a (k(cat)/K(M))/k(uncat) = 10(7) at pH 5.0. The crystal structure of this enzyme was determined at 1.30 ?, as were the structures of four complexes of substrate and product analogs. The absence of ordered waters or hydrogen bonding interactions, and the presence of a common catalytic base (His102) in an otherwise hydrophobic, buried cavity, facilitated detailed analysis of the reaction mechanism and its optimization. Subsequent substitutions increased eliminase activity by an additional four-fold. As activity-enhancing substitutions were engineered into the cavity, protein stability decreased, consistent with the stability-function trade-off hypothesis. This and related model cavities may provide templates for studying protein design principles in radically simplified environments. PMID:22988064

  16. Engineering a model protein cavity to catalyze the Kemp elimination

    PubMed Central

    Merski, Matthew; Shoichet, Brian K.

    2012-01-01

    Synthetic cavitands and protein cavities have been widely studied as models for ligand recognition. Here we investigate the Met102 ? His substitution in the artificial L99A cavity in T4 lysozyme as a Kemp eliminase. The resulting enzyme had kcat/KM = 0.43 M-1 s-1 and a (kcat/KM)/kuncat = 107 at pH 5.0. The crystal structure of this enzyme was determined at 1.30 ?, as were the structures of four complexes of substrate and product analogs. The absence of ordered waters or hydrogen bonding interactions, and the presence of a common catalytic base (His102) in an otherwise hydrophobic, buried cavity, facilitated detailed analysis of the reaction mechanism and its optimization. Subsequent substitutions increased eliminase activity by an additional four-fold. As activity-enhancing substitutions were engineered into the cavity, protein stability decreased, consistent with the stability-function trade-off hypothesis. This and related model cavities may provide templates for studying protein design principles in radically simplified environments. PMID:22988064

  17. Statistical mechanics of simple models of protein folding and design.

    PubMed Central

    Pande, V S; Grosberg, A Y; Tanaka, T

    1997-01-01

    It is now believed that the primary equilibrium aspects of simple models of protein folding are understood theoretically. However, current theories often resort to rather heavy mathematics to overcome some technical difficulties inherent in the problem or start from a phenomenological model. To this end, we take a new approach in this pedagogical review of the statistical mechanics of protein folding. The benefit of our approach is a drastic mathematical simplification of the theory, without resort to any new approximations or phenomenological prescriptions. Indeed, the results we obtain agree precisely with previous calculations. Because of this simplification, we are able to present here a thorough and self contained treatment of the problem. Topics discussed include the statistical mechanics of the random energy model (REM), tests of the validity of REM as a model for heteropolymer freezing, freezing transition of random sequences, phase diagram of designed ("minimally frustrated") sequences, and the degree to which errors in the interactions employed in simulations of either folding and design can still lead to correct folding behavior. Images FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 6 PMID:9414231

  18. Multi-Dimensional Scaling and MODELLER-Based Evolutionary Algorithms for Protein Model Refinement

    PubMed Central

    Chen, Yan; Shang, Yi; Xu, Dong

    2015-01-01

    Protein structure prediction, i.e., computationally predicting the three-dimensional structure of a protein from its primary sequence, is one of the most important and challenging problems in bioinformatics. Model refinement is a key step in the prediction process, where improved structures are constructed based on a pool of initially generated models. Since the refinement category was added to the biennial Critical Assessment of Structure Prediction (CASP) in 2008, CASP results show that it is a challenge for existing model refinement methods to improve model quality consistently. This paper presents three evolutionary algorithms for protein model refinement, in which multidimensional scaling(MDS), the MODELLER software, and a hybrid of both are used as crossover operators, respectively. The MDS-based method takes a purely geometrical approach and generates a child model by combining the contact maps of multiple parents. The MODELLER-based method takes a statistical and energy minimization approach, and uses the remodeling module in MODELLER program to generate new models from multiple parents. The hybrid method first generates models using the MDS-based method and then run them through the MODELLER-based method, aiming at combining the strength of both. Promising results have been obtained in experiments using CASP datasets. The MDS-based method improved the best of a pool of predicted models in terms of the global distance test score (GDT-TS) in 9 out of 16test targets. PMID:25844403

  19. Ionophoric properties of the proteolipid apoprotein from bovine brain myelin.

    PubMed

    de Cózar, M; Lucas, M; Monreal, J

    1987-05-01

    Ionophoric properties of the Proteolipid Apoprotein have been assayed. This is a highly purified and delipidated intrinsic myelin membrane protein, isolated from bovine brain white matter. The preparation of myelin membrane vesicles or the incorporation of purified protein into Dimiristoylphosphatidylcholine liposomes have been carried out. According to our results, the myelin Proteolipid protein may act as a Na+ and Rb+ (K+) unidirectional ionophoretic channel, which main physiological role could be related to the maintenance of ionic equilibrium of myelin sheath around the axons. PMID:2457371

  20. PRISM: a web server and repository for prediction of protein–protein interactions and modeling their 3D complexes

    PubMed Central

    Baspinar, Alper; Cukuroglu, Engin; Nussinov, Ruth; Keskin, Ozlem; Gursoy, Attila

    2014-01-01

    The PRISM web server enables fast and accurate prediction of protein–protein interactions (PPIs). The prediction algorithm is knowledge-based. It combines structural similarity and accounts for evolutionary conservation in the template interfaces. The predicted models are stored in its repository. Given two protein structures, PRISM will provide a structural model of their complex if a matching template interface is available. Users can download the complex structure, retrieve the interface residues and visualize the complex model. The PRISM web server is user friendly, free and open to all users at http://cosbi.ku.edu.tr/prism. PMID:24829450