Sample records for seo proteins results

  1. deSEO: Combating Search-Result Poisoning John P. John,

    E-print Network

    Abadi, Martín

    deSEO: Combating Search-Result Poisoning John P. John, , Fang Yu§ , Yinglian Xie§ , Arvind malware by poisoning search results for popular queries. Such attacks, although recent, appear to be both through search engine optimiza- tion (SEO) techniques, attackers are able to poison the search results

  2. Calcium powered phloem protein of SEO gene family “Forisome” functions in wound sealing and act as biomimetic smart materials

    PubMed Central

    Srivastava, Vineet Kumar; Tuteja, Narendra

    2014-01-01

    Forisomes protein belongs to SEO gene family and is unique to Fabaceae family. These proteins are located in sieve tubes of phloem and function to prevent loss of nutrient-rich photoassimilates, upon mechanical injury/wounding. Forisome protein is also known as ATP independent, mechanically active proteins. Despite the wealth of information role of forisome in plants are not yet fully understood. Recent reports suggest that forisomes protein can act as ideal model to study self assembly mechanism for development of nanotechnological devices like microfluidic system application in space exploration mission. Improvement in micro instrument is highly demanding and has been a key technology by NASA in future space exploration missions. Based on its physical parameters, forisome are found to be ideal biomimetic materials for micro fluidic system because the conformational shifts can be replicated in vitro and are fully reversible over large number of cycles. By the use of protein engineering forisome recombinant protein can be tailored. Due to its unique ability to convert chemical energy into mechanical energy forisome has received much attention. For nanotechnological application and handling biomolecules such as DNA, RNA, protein and cell as a whole microfluidic system will be the most powerful technology. The discovery of new biomimetic smart materials has been a key factor in development of space science and its requirements in such a challenging environment. The field of microfludic, particularly in terms of development of its components along with identification of new biomimetic smart materials, deserves more attention. More biophysical investigation is required to characterize it to make it more suitable under parameters of performance. PMID:25763691

  3. Calcium powered phloem protein of SEO gene family "Forisome" functions in wound sealing and act as biomimetic smart materials.

    PubMed

    Srivastava, Vineet Kumar; Tuteja, Narendra

    2014-06-01

    Forisomes protein belongs to SEO gene family and is unique to Fabaceae family. These proteins are located in sieve tubes of phloem and function to prevent loss of nutrient-rich photoassimilates, upon mechanical injury/wounding. Forisome protein is also known as ATP independent, mechanically active proteins. Despite the wealth of information role of forisome in plants are not yet fully understood. Recent reports suggest that forisomes protein can act as ideal model to study self assembly mechanism for development of nanotechnological devices like microfluidic system application in space exploration mission. Improvement in micro instrument is highly demanding and has been a key technology by NASA in future space exploration missions. Based on its physical parameters, forisome are found to be ideal biomimetic materials for micro fluidic system because the conformational shifts can be replicated in vitro and are fully reversible over large number of cycles. By the use of protein engineering forisome recombinant protein can be tailored. Due to its unique ability to convert chemical energy into mechanical energy forisome has received much attention. For nanotechnological application and handling biomolecules such as DNA, RNA, protein and cell as a whole microfluidic system will be the most powerful technology. The discovery of new biomimetic smart materials has been a key factor in development of space science and its requirements in such a challenging environment. The field of microfludic, particularly in terms of development of its components along with identification of new biomimetic smart materials, deserves more attention. More biophysical investigation is required to characterize it to make it more suitable under parameters of performance. PMID:24905822

  4. SEOS frame camera applications study

    NASA Technical Reports Server (NTRS)

    1974-01-01

    A research and development satellite is discussed which will provide opportunities for observation of transient phenomena that fall within the fixed viewing circle of the spacecraft. The evaluation of possible applications for frame cameras, for SEOS, are studied. The computed lens characteristics for each camera are listed.

  5. Sodium selenite (Na2SeO3) induces apoptosis through the mitochondrial pathway in CNE-2 nasopharyngeal carcinoma cells.

    PubMed

    Cui, Zhongyi; Li, Caihong; Li, Xiangyong; Zhang, Qunzhou; Zhang, Yuefei; Shao, Jingjing; Zhou, Keyuan

    2015-06-01

    This study investigated the effect of sodium selenite (Na2SeO3) on proliferation, cell cycle, apoptosis as well as the underlying mechanism in CNE?2 nasopharyngeal carcinoma (NPC) cells. The CNE?2 cell line was treated with different concentrations of Na2SeO3, and the effects of Na2SeO3 on cell viability and proliferation were evaluated using Cell Counting kit?8 (CCK?8) assay. Cellular apoptosis and cell cycle were evaluated by flow cytomerty following Annexin V?FITC/PI double staining and PI single staining respectively; nuclei morphology stained with DAPI and Hoechst 333258 was observed under a fluorescence microscope, while DNA fragmentation was detected by agarose gel electrophoresis. The mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) were analyzed using fluorescent staining assays. Expression of Bcl?XL, Bax, Bak, and caspase?3 activation were examined by western blotting. The results showed that Na2SeO3 inhibited proliferation and induced apoptosis of CNE?2 cells in a dose? and time?dependent manner. Na2SeO3 at low concentrations induced cell cycle arrest at S phase, while high concentrations of Na2SeO3 induced cell cycle arrest at G0/G1 phase. Furthermore, Na2SeO3 increased ROS level and decreased MMP, upregulated caspase?3 activity and the expression of Bak and Bax but simultaneously downregulated Bcl?XL. In conclusion, our studies demonstrated that Na2SeO3 had significant anti?proliferative and apoptosis?inducing effects via arresting cell cycle and regulating mitochondria?mediated intrinsic caspase pathway in CNE?2 NPC cells, suggesting that Na2SeO3 might have therapeutic potentials in the treatment of NPC. PMID:25891011

  6. Synthesis, structure, and characterization of two new polar sodium tungsten selenites: Na2(WO3)3(SeO3)·2H2O and Na6(W6O19)(SeO3)2.

    PubMed

    Nguyen, Sau Doan; Halasyamani, P Shiv

    2013-03-01

    Two new quaternary sodium tungsten selenites, Na2(WO3)3(SeO3)·2H2O (P31c) and Na6(W6O19)(SeO3)2 (C2), have been synthesized and characterized. The former exhibits a hexagonal tungsten oxide layered structure, whereas the latter has a one-dimensional "ribbon" structure. The layers and "ribbons" consist of distorted WO6 and asymmetric SeO3 polyhedra. The layers in Na2(WO3)3(SeO3)·2H2O and the "ribbons" in Na6(W6O19)(SeO3)2 are separated by Na(+) cations. Powder second-harmonic-generation (SHG) measurements on Na2(WO3)3(SeO3)·2H2O and Na6(W6O19)(SeO3)2 using 1064 nm radiation reveal SHG efficiencies of approximately 450× and 20× ?-SiO2, respectively. Particle size versus SHG efficiency measurements indicate that the materials are type 1 non-phase-matchable. Converse piezoelectric measurements result in d33 values of approximately 23 and 12 pm/V, whereas pyroelectric measurements reveal coefficients of -0.41 and -1.10 ?C/m(2)·K at 60 °C for Na2(WO3)3(SeO3)·2H2O and Na6(W6O19)(SeO3)2, respectively. Frequency-dependent polarization measurements confirm that the materials are nonferroelectric; i.e., the macroscopic polarization is not reversible, or "switchable". IR and UV-vis spectroscopy, thermogravimetric and differential thermal analysis measurements, and electron localization function calculations were also done for the materials. Crystal data: Na2(WO3)3(SeO3)·2H2O, trigonal, space group P31c (No. 159), a = 7.2595(6) Ĺ, b = 7.2595(6) Ĺ, c = 12.4867(13) Ĺ, V = 569.89(9) Ĺ(3), Z = 2; Na6(W6O19)(SeO3)2, monoclinic, space group C2 (No. 5), a = 42.169(8) Ĺ, b = 7.2690(15) Ĺ, c = 6.7494(13) Ĺ, ? = 98.48(3)°, V = 2046.2(7) Ĺ(3), Z = 4. PMID:23425251

  7. J protein mutations and resulting proteostasis collapse

    PubMed Central

    Koutras, Carolina; Braun, Janice E. A.

    2014-01-01

    Despite a century of intensive investigation the effective treatment of protein aggregation diseases remains elusive. Ordinarily, molecular chaperones ensure that proteins maintain their functional conformation. The appearance of misfolded proteins that aggregate implies the collapse of the cellular chaperone quality control network. That said, the cellular chaperone network is extensive and functional information regarding the detailed action of specific chaperones is not yet available. J proteins (DnaJ/Hsp40) are a family of chaperone cofactors that harness Hsc70 (heat shock cognate protein of 70 kDa) for diverse conformational cellular tasks and, as such, represent novel clinically relevant targets for diseases resulting from the disruption of proteostasis. Here we review incisive reports identifying mutations in individual J protein chaperones and the proteostasis collapse that ensues. PMID:25071450

  8. Capital structures in an emerging market: a duration analysis of the time interval between IPO and SEO in China

    Microsoft Academic Search

    Yang Ni; Shasha Guo; David E. Giles

    2010-01-01

    We model the durations between firms’ ‘Initial Public Offerings’ (IPOs) and their subsequent ‘Seasoned Equity Offerings’ (SEOs) in China between 2001 and 2006. Our results have important implications for the capital structure in emerging markets. Our evidence on financing decisions in China contradicts the predictions of both the trade-off and pecking order theories. Firms do not issue equity after debt

  9. Capital Structures in an Emerging Market: A Duration Analysis of the Time Interval Between IPO and SEO in China

    Microsoft Academic Search

    Yang Ni; Shasha Guo; David E. Giles

    2009-01-01

    We model the durations between firms’ “Initial Public Offerings” (IPOs) and their subsequent “Seasoned Equity Offerings” (SEOs) in China during the period from 1 January 2001 to 1 July 2006. Duration analysis is applied by using the nonparametric Kaplan-Meier estimator of the hazard function, and parametric accelerated failure time models with time-varying covariates. The results of this analysis have important

  10. On the dielectric susceptibility calculation in the incommensurate phase of K2SeO4

    NASA Astrophysics Data System (ADS)

    Aslanyan, T. A.

    2010-10-01

    It is shown that the thermodynamic potential of the domain-like incommensurate (IC) phase of the K2SeO4crystal (viewed as a model for the IC-C transition) should be supplemented with a term, taking into account the local, Lorentz electric field. The latter qualitatively changes the result of calculation of the dielectric susceptibility for this IC structure by Nattermann and Trimper, J. Phys. C: Solid State Phys. 14, 1603, (1981), and gives phase transition to the ferroelectric IC phase obtained by Aslanyan, Phys. Rev. B 70, 024102, (2004).

  11. Fecal Calprotectin is an Accurate Tool and Correlated to Seo Index in Prediction of Relapse in Iranian Patients With Ulcerative Colitis

    PubMed Central

    Hosseini, Seyed Vahid; Jafari, Peyman; Taghavi, Seyed Alireza; Safarpour, Ali Reza; Rezaianzadeh, Abbas; Moini, Maryam; Mehrabi, Manoosh

    2015-01-01

    Background: The natural clinical course of Ulcerative Colitis (UC) is characterized by episodes of relapse and remission. Fecal Calprotectin (FC) is a relatively new marker of intestinal inflammation and is an available, non-expensive tool for predicting relapse of quiescent UC. The Seo colitis activity index is a clinical index for assessment of the severity of UC. Objectives: The present study aimed to evaluate the accuracy of FC and the Seo colitis activity index and their correlation in prediction of UC exacerbation. Patients and Methods: In this prospective cohort study, 157 patients with clinical and endoscopic diagnosis of UC selected randomly from 1273 registered patients in Fars province’s IBD registry center in Shiraz, Iran, were followed from October 2012 to October 2013 for 12 months or shorter, if they had a relapse. Two patients left the study before completion and one patient had relapse because of discontinuation of drugs. The participants' clinical and serum factors were evaluated every three months. Furthermore, stool samples were collected at the beginning of study and every three months and FC concentration (commercially available enzyme linked immunoassay) and the Seo Index were assessed. Then univariate analysis, multiple variable logistic regression, Receiver Operating Characteristics (ROC) curve analysis, and Pearson’s correlation test (r) were used for statistical analysis of data. Results: According to the results, 74 patients (48.1%) relapsed during the follow-up (33 men and 41 women). Mean ± SD of FC was 862.82 ± 655.97 ?g/g and 163.19 ± 215.85 ?g/g in relapsing and non-relapsing patients, respectively (P < 0.001). Multiple logistic regression analysis revealed that age, number of previous relapses, FC and the Seo index were significant predictors of relapse. ROC curve analysis of FC level and Seo activity index for prediction of relapse demonstrated area under the curve of 0.882 (P < 0.001) and 0.92 1(P < 0.001), respectively. Besides, FC level of 341 ?g/g was identified as the cut-off point with 11.2% and 79.7% relapse rate below and above this point, respectively. Additionally, Pearson correlation coefficient (r) between FC and the Seo index was significant in prediction of relapse (r = 0.63, P < 0.001). Conclusions: As a simple and noninvasive marker, FC is highly accurate and significantly correlated to the Seo activity index in prediction of relapse in the course of quiescent UC in Iranian patients. PMID:25793117

  12. Syntheses and Crystal Structures of Er2(SeO3)3 and Dy3(SeO3)4F

    Microsoft Academic Search

    Ina Krügermann; Mathias S. Wickleder

    2002-01-01

    The decomposition of Er2(SeO4)3 in the presence of LiF in a sealed gold ampoule at 870°C yielded single crystals of Er2(SeO3)3. The triclinic crystal structure (P1, Z=2, a=698.2(1), b=800.6(1), c=895.0(1) pm, alpha=71.38(1), beta=70.13(1), gamma=65.87(1)°, Rall=0.0364) contains two crystallographically different Er3+ ions in seven- and eightfold coordination of oxygen atoms, respectively. The distances Er-O range from 220 to 256 pm. Each

  13. Syntheses and Crystal Structures of Er 2(SeO 3) 3 and Dy 3(SeO 3) 4F

    Microsoft Academic Search

    Ina Krügermann; Mathias S. Wickleder

    2002-01-01

    The decomposition of Er2(SeO4)3 in the presence of LiF in a sealed gold ampoule at 870°C yielded single crystals of Er2(SeO3)3. The triclinic crystal structure (P1, Z=2, a=698.2(1), b=800.6(1), c=895.0(1) pm, ?=71.38(1), ?=70.13(1), ?=65.87(1)°, Rall=0.0364) contains two crystallographically different Er3+ ions in seven- and eightfold coordination of oxygen atoms, respectively. The distances Er–O range from 220 to 256 pm. Each

  14. A physical approach to protein structure prediction: CASP4 results

    SciTech Connect

    Crivelli, Silvia; Eskow, Elizabeth; Bader, Brett; Lamberti, Vincent; Byrd, Richard; Schnabel, Robert; Head-Gordon, Teresa

    2001-02-27

    We describe our global optimization method called Stochastic Perturbation with Soft Constraints (SPSC), which uses information from known proteins to predict secondary structure, but not in the tertiary structure predictions or in generating the terms of the physics-based energy function. Our approach is also characterized by the use of an all atom energy function that includes a novel hydrophobic solvation function derived from experiments that shows promising ability for energy discrimination against misfolded structures. We present the results obtained using our SPSC method and energy function for blind prediction in the 4th Critical Assessment of Techniques for Protein Structure Prediction (CASP4) competition, and show that our approach is more effective on targets for which less information from known proteins is available. In fact our SPSC method produced the best prediction for one of the most difficult targets of the competition, a new fold protein of 240 amino acids.

  15. LAPLACE-BELTRAMI EIGENFUNCTION EXPANSION OF CORTICAL MANIFOLDS Seongho Seo1

    E-print Network

    Chung, Moo K.

    LAPLACE-BELTRAMI EIGENFUNCTION EXPANSION OF CORTICAL MANIFOLDS Seongho Seo1 Moo K. Chung1,2,3, 1 of Laplace-Beltrami operator and compare the performance with the conventional spherical harmonic (SPHARM) representation. Cortical manifolds are repre- sented as a linear combination of the Laplace-Beltrami eigen

  16. The RCS of Wire-type Scattering Structures Dong-wook Seo1

    E-print Network

    Myung, Noh-Hoon

    The RCS of Wire-type Scattering Structures Dong-wook Seo1 and Noh-Hoon Myung1 1 School works mainly utilized the method of moment (MoM) for predicting the radar cross section (RCS) of a large to minimize the calculation time. In this case, the total RCS of many wires is simply the product

  17. Protein-protein interactions: analysis of a false positive GST pulldown result.

    PubMed

    Wissmueller, Sandra; Font, Josep; Liew, Chu Wai; Cram, Edward; Schroeder, Thilo; Turner, Jeremy; Crossley, Merlin; Mackay, Joel P; Matthews, Jacqueline M

    2011-08-01

    One of the most common ways to demonstrate a direct protein-protein interaction in vitro is the glutathione-S-transferse (GST)-pulldown. Here we report the detailed characterization of a putative interaction between two transcription factor proteins, GATA-1 and Krüppel-like factor 3 (KLF3/BKLF) that show robust interactions in GST-pulldown experiments. Attempts to map the interaction interface of GATA-1 on KLF3 using a mutagenic screening approach did not yield a contiguous binding face on KLF3, suggesting that the interaction might be non-specific. NMR experiments showed that the proteins do not interact at protein concentrations of 50-100 ?M. Rather, the GST tag can cause part of KLF3 to misfold. In addition to misfolding, the fact that both proteins are DNA-binding domains appears to introduce binding artifacts (possibly nucleic acid bridging) that cannot be resolved by the addition of nucleases or ethidium bromide (EtBr). This study emphasizes the need for caution in relying on GST-pulldown results and related methods, without convincing confirmation from different approaches. PMID:21638332

  18. New vanadium selenites: centrosymmetric Ca2(VO2)2(SeO3)3(H2O)2, Sr2(VO2)2(SeO3)3, and Ba(V2O5)(SeO3), and noncentrosymmetric and polar A4(VO2)2(SeO3)4(Se2O5) (A = Sr2+ or Pb2+).

    PubMed

    Yeon, Jeongho; Kim, Sang-Hwan; Nguyen, Sau Doan; Lee, Hana; Halasyamani, P Shiv

    2012-01-01

    Five new vanadium selenites, Ca(2)(VO(2))(2)(SeO(3))(3)(H(2)O)(2), Sr(2)(VO(2))(2)(SeO(3))(3), Ba(V(2)O(5))(SeO(3)), Sr(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)), and Pb(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)), have been synthesized and characterized. Their crystal structures were determined by single crystal X-ray diffraction. The compounds exhibit one- or two-dimensional structures consisting of corner- and edge-shared VO(4), VO(5), VO(6), and SeO(3) polyhedra. Of the reported materials, A(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)) (A = Sr(2+) or Pb(2+)) are noncentrosymmetric (NCS) and polar. Powder second-harmonic generation (SHG) measurements revealed SHG efficiencies of approximately 130 and 150 × ?-SiO(2) for Sr(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)) and Pb(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)), respectively. Piezoelectric charge constants of 43 and 53 pm/V, and pyroelectric coefficients of -27 and -42 ?C/m(2)·K at 70 °C were obtained for Sr(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)) and Pb(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)), respectively. Frequency dependent polarization measurements confirmed that the materials are not ferroelectric, that is, the observed polarization cannot be reversed. In addition, the lone-pair on the Se(4+) cation may be considered as stereo-active consistent with calculations. For all of the reported materials, infrared, UV-vis, thermogravimetric, and differential thermal analysis measurements were performed. Crystal data: Ca(2)(VO(2))(2)(SeO(3))(3)(H(2)O)(2), orthorhombic, space group Pnma (No. 62), a = 7.827(4) Ĺ, b = 16.764(5) Ĺ, c = 9.679(5) Ĺ, V = 1270.1(9) Ĺ(3), and Z = 4; Sr(2)(VO(2))(2)(SeO(3))(3), monoclinic, space group P2(1)/c (No. 12), a = 14.739(13) Ĺ, b = 9.788(8) Ĺ, c = 8.440(7) Ĺ, ? = 96.881(11)°, V = 1208.8(18) Ĺ(3), and Z = 4; Ba(V(2)O(5))(SeO(3)), orthorhombic, space group Pnma (No. 62), a = 13.9287(7) Ĺ, b = 5.3787(3) Ĺ, c = 8.9853(5) Ĺ, V = 673.16(6) Ĺ(3), and Z = 4; Sr(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)), orthorhombic, space group Fdd2 (No. 43), a = 25.161(3) Ĺ, b = 12.1579(15) Ĺ, c = 12.8592(16) Ĺ, V = 3933.7(8) Ĺ(3), and Z = 8; Pb(4)(VO(2))(2)(SeO(3))(4)(Se(2)O(5)), orthorhombic, space group Fdd2 (No. 43), a = 25.029(2) Ĺ, b = 12.2147(10) Ĺ, c = 13.0154(10) Ĺ, V = 3979.1(6) Ĺ(3), and Z = 8. PMID:22145697

  19. Success in mathematics within a challenged minority: the case of students of Ethiopian origin in Israel (SEO)

    Microsoft Academic Search

    Tiruwork Mulat; Abraham Arcavi

    2009-01-01

    Many studies have reported on the economical, social, and educational difficulties encountered by Ethiopian Jews since their\\u000a immigration to Israel. Furthermore, the overall academic underachievement and poor representation of students of Ethiopian\\u000a origin (SEO) in the advanced mathematics and science classes were highlighted and described. Yet, studies focusing on differential\\u000a achievements within SEO and on students who succeed against all

  20. Ribosomal Protein Mutations Result in Constitutive p53 Protein Degradation through Impairment of the AKT Pathway

    PubMed Central

    Hermkens, Dorien; Wlodarski, Marcin W.; Da Costa, Lydie; MacInnes, Alyson W.

    2015-01-01

    Mutations in ribosomal protein (RP) genes can result in the loss of erythrocyte progenitor cells and cause severe anemia. This is seen in patients with Diamond-Blackfan anemia (DBA), a pure red cell aplasia and bone marrow failure syndrome that is almost exclusively linked to RP gene haploinsufficiency. While the mechanisms underlying the cytopenia phenotype of patients with these mutations are not completely understood, it is believed that stabilization of the p53 tumor suppressor protein may induce apoptosis in the progenitor cells. In stark contrast, tumor cells from zebrafish with RP gene haploinsufficiency are unable to stabilize p53 even when exposed to acute DNA damage despite transcribing wild type p53 normally. In this work we demonstrate that p53 has a limited role in eliciting the anemia phenotype of zebrafish models of DBA. In fact, we find that RP-deficient embryos exhibit the same normal p53 transcription, absence of p53 protein, and impaired p53 response to DNA damage as RP haploinsufficient tumor cells. Recently we reported that RP mutations suppress activity of the AKT pathway, and we show here that this suppression results in proteasomal degradation of p53. By re-activating the AKT pathway or by inhibiting GSK-3, a downstream modifier that normally represses AKT signaling, we are able to restore the stabilization of p53. Our work indicates that the anemia phenotype of zebrafish models of DBA is dependent on factors other than p53, and may hold clinical significance for both DBA and the increasing number of cancers revealing spontaneous mutations in RP genes. PMID:26132763

  1. Ribosomal Protein Mutations Result in Constitutive p53 Protein Degradation through Impairment of the AKT Pathway.

    PubMed

    Antunes, Ana T; Goos, Yvonne J; Pereboom, Tamara C; Hermkens, Dorien; Wlodarski, Marcin W; Da Costa, Lydie; MacInnes, Alyson W

    2015-07-01

    Mutations in ribosomal protein (RP) genes can result in the loss of erythrocyte progenitor cells and cause severe anemia. This is seen in patients with Diamond-Blackfan anemia (DBA), a pure red cell aplasia and bone marrow failure syndrome that is almost exclusively linked to RP gene haploinsufficiency. While the mechanisms underlying the cytopenia phenotype of patients with these mutations are not completely understood, it is believed that stabilization of the p53 tumor suppressor protein may induce apoptosis in the progenitor cells. In stark contrast, tumor cells from zebrafish with RP gene haploinsufficiency are unable to stabilize p53 even when exposed to acute DNA damage despite transcribing wild type p53 normally. In this work we demonstrate that p53 has a limited role in eliciting the anemia phenotype of zebrafish models of DBA. In fact, we find that RP-deficient embryos exhibit the same normal p53 transcription, absence of p53 protein, and impaired p53 response to DNA damage as RP haploinsufficient tumor cells. Recently we reported that RP mutations suppress activity of the AKT pathway, and we show here that this suppression results in proteasomal degradation of p53. By re-activating the AKT pathway or by inhibiting GSK-3, a downstream modifier that normally represses AKT signaling, we are able to restore the stabilization of p53. Our work indicates that the anemia phenotype of zebrafish models of DBA is dependent on factors other than p53, and may hold clinical significance for both DBA and the increasing number of cancers revealing spontaneous mutations in RP genes. PMID:26132763

  2. Applications of synchrotron radiation to protein crystallography: preliminary results.

    PubMed

    Phillips, J C; Wlodawer, A; Yevitz, M M; Hodgson, K O

    1976-01-01

    X-ray diffraction photographs of protein single crystals have been obtained using synchrotron radiation produced by an electron-positron storage ring. The diffracted intensities observed with this unconventional source are a factor of at least 60 greater than those obtained with a sealed x-ray tube using the same crystal and instrumental parameters. Diffraction data have been collected by the precession method to higher resolution and using smaller protein crystals than would have been possible with a conventional source. The crystal decay rate in the synchrotron beam for several proteins appears to be substantially less than that observed with Ni-filtered Cu radiation. The tunable nature of the source (which allows selective optimization of anomalous contributions to the scattering factors) and the low angular divergence of the beam make the source very useful for single crystal protein diffraction studies. PMID:1061106

  3. Diminished Self-Chaperoning Activity of the DF508 Mutant of CFTR Results in Protein Misfolding

    E-print Network

    Dokholyan, Nikolay V.

    Diminished Self-Chaperoning Activity of the DF508 Mutant of CFTR Results in Protein Misfolding) Diminished Self-Chaperoning Activity of the DF508 Mutant of CFTR Results in Protein Misfolding. PLoS Comput The absence of a functional ATP Binding Cassette (ABC) protein called the Cystic Fibrosis Transmembrane

  4. Synthesis, structure, and characterization of new Li+-d0-lone-pair-oxides: noncentrosymmetric polar Li6(Mo2O5)3(SeO3)6 and centrosymmetric Li2(MO3)(TeO3) (M = Mo6+ or W6+).

    PubMed

    Nguyen, Sau Doan; Halasyamani, P Shiv

    2012-09-01

    New quaternary lithium-d(0) cation-lone-pair oxides, Li(6)(Mo(2)O(5))(3)(SeO(3))(6) (Pmn2(1)) and Li(2)(MO(3))(TeO(3)) (P2(1)/n) (M = Mo(6+) or W(6+)), have been synthesized and characterized. The former is noncentrosymmetric and polar, whereas the latter is centrosymmetric. Their crystal structures exhibit zigzag anionic layers composed of distorted MO(6) and asymmetric AO(3) (A = Se(4+) or Te(4+)) polyhedra. The anionic layers stack along a 2-fold screw axis and are separated by Li(+) cations. Powder SHG measurements on Li(6)(Mo(2)O(5))(3)(SeO(3))(6) using 1064 nm radiation reveal a SHG efficiency of approximately 170 × ?-SiO(2). Particle size vs SHG efficiency measurements indicate Li(6)(Mo(2)O(5))(3)(SeO(3))(6) is type 1 nonphase-matchable. Converse piezoelectric measurements result in a d(33) value of ?28 pm/V and pyroelectric measurements reveal a pyroelectric coefficient of -0.43 ?C/m(2)K at 50 °C for Li(6)(Mo(2)O(5))(3)(SeO(3))(6). Frequency-dependent polarization measurements confirm that Li(6)(Mo(2)O(5))(3)(SeO(3))(6) is nonferroelectric, i.e., the macroscopic polarization is not reversible, or 'switchable'. Infrared, UV-vis, thermogravimetric, and differential thermal analysis measurements and electron localization function calculations were also done for all materials. PMID:22916963

  5. Combinatorial Algorithms for Protein Folding in Lattice Models: A Survey of Mathematical Results

    E-print Network

    Istrail, Sorin

    Combinatorial Algorithms for Protein Folding in Lattice Models: A Survey of Mathematical Results a comprehensive survey of combinatorial algorithms and theorems about lattice protein folding models obtained in the almost 15 years since the publication in 1995 of the first protein folding approximation algorithm

  6. Annotation of proteins of unknown function: initial enzyme results.

    PubMed

    McKay, Talia; Hart, Kaitlin; Horn, Alison; Kessler, Haeja; Dodge, Greg; Bardhi, Keti; Bardhi, Kostandina; Mills, Jeffrey L; Bernstein, Herbert J; Craig, Paul A

    2015-03-01

    Working with a combination of ProMOL (a plugin for PyMOL that searches a library of enzymatic motifs for local structural homologs), BLAST and Pfam (servers that identify global sequence homologs), and Dali (a server that identifies global structural homologs), we have begun the process of assigning functional annotations to the approximately 3,500 structures in the Protein Data Bank that are currently classified as having "unknown function". Using a limited template library of 388 motifs, over 500 promising in silico matches have been identified by ProMOL, among which 65 exceptionally good matches have been identified. The characteristics of the exceptionally good matches are discussed. PMID:25630330

  7. Highmobility inverted selectively doped heterojunctions Hadas Shtrikman, M. Heiblum, K. Seo, D. E. Galbi, and L. Osterling

    E-print Network

    Heiblum, Mordehai "Moty"

    Highmobility inverted selectively doped heterojunctions Hadas Shtrikman, M. Heiblum, K. Seo, D. E in inverted GaAsAlGaAs heterojunctions Appl. Phys. Lett. 52, 1268 (1988); 10.1063/1.99176 High mobility: 132.76.50.6 On: Thu, 10 Apr 2014 16:05:40 #12;High-mobility inverted selectively doped heterojunctions

  8. 31 CFR 30.3 - Q-3: How are the SEOs and most highly compensated employees identified for purposes of compliance...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... Money and Finance: Treasury Office of the Secretary of the Treasury TARP STANDARDS FOR COMPENSATION AND CORPORATE GOVERNANCE § 30.3 Q-3: How are the SEOs and most highly compensated employees identified for purposes of...

  9. 31 CFR 30.3 - Q-3: How are the SEOs and most highly compensated employees identified for purposes of compliance...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... Money and Finance: Treasury Office of the Secretary of the Treasury TARP STANDARDS FOR COMPENSATION AND CORPORATE GOVERNANCE § 30.3 Q-3: How are the SEOs and most highly compensated employees identified for purposes of...

  10. 31 CFR 30.3 - Q-3: How are the SEOs and most highly compensated employees identified for purposes of compliance...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... Money and Finance: Treasury Office of the Secretary of the Treasury TARP STANDARDS FOR COMPENSATION AND CORPORATE GOVERNANCE § 30.3 Q-3: How are the SEOs and most highly compensated employees identified for purposes of...

  11. 31 CFR 30.3 - Q-3: How are the SEOs and most highly compensated employees identified for purposes of compliance...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... Money and Finance: Treasury Office of the Secretary of the Treasury TARP STANDARDS FOR COMPENSATION AND CORPORATE GOVERNANCE § 30.3 Q-3: How are the SEOs and most highly compensated employees identified for purposes of...

  12. Proteins

    NSDL National Science Digital Library

    Mowery, Jeanette

    Laboratory manual and supplemental resources that were developed for a college laboratory course in protein purification. The enzyme, Beta-galactosidase, is purified in two steps, with analysis and verification of results. Course materials are divided into four units: Why Proteins, Assays, The Purification Process, and Analysis and Verification. Powerpoint lectures and study guides are provided.

  13. Casein protein results in higher prandial and exercise induced whole body protein anabolism than whey protein in Chronic Obstructive Pulmonary Disease

    PubMed Central

    Engelen, Mariëlle PKJ; Rutten, Erica PA; De Castro, Carmen LN; Wouters, Emiel FM; Schols, Annemie MWJ; Deutz, Nicolaas EP

    2012-01-01

    Objective Exercise is known to improve physical functioning and health status in Chronic Obstructive Pulmonary Disease (COPD). Recently, disturbances in protein turnover and amino acid kinetics have been observed after exercise in COPD. The objective was to investigate which dairy protein is able to positively influence the protein metabolic response to exercise in COPD. Materials and Methods 8 COPD patients and 8 healthy subjects performed a cycle test on two days while ingesting casein or whey protein. Whole body protein breakdown (WbPB), synthesis (WbPS), splanchnic amino acid extraction (SPE), and NetWbPS (=WbPS–WbPB) were measured using stable isotope methodology during 20 minutes of exercise (at 50% peak work load of COPD group). The controls performed a second exercise test at the same relative workload. Exercise was followed by 1 hour of recovery. Results In the healthy group, WbPS, SPE, and NetPS were higher during casein than during whey feeding (p<0.01). WbPS and NetPS were higher during exercise, independent of exercise intensity (p<0.01). NetPS was higher during casein feeding in COPD due to lower WbPB (p<0.05). Higher SPE was found during exercise during casein and whey feeding in COPD (p<0.05). Lactate levels during exercise were higher in COPD (p<0.05) independent of the protein. Post-exercise, lower NetPS values were found independent of protein type in both groups. Conclusion Casein resulted in more protein anabolism than whey protein which was maintained during and following exercise in COPD. Optimizing protein intake might be of importance for muscle maintenance during daily physical activities in COPD. PMID:22512824

  14. Recent results and new hardware developments for protein crystal growth in microactivity

    NASA Technical Reports Server (NTRS)

    Delucas, L. J.; Long, M. M.; Moore, K. M.; Smith, C.; Carson, M.; Narayana, S. V. L.; Carter, D.; Clark, A. D., Jr.; Nanni, R. G.; Ding, J.

    1993-01-01

    Protein crystal growth experiments have been performed on 16 space shuttle missions since April, 1985. The initial experiments utilized vapor diffusion crystallization techniques similar to those used in laboratories for earth-based experiments. More recent experiments have utilized temperature induced crystallization as an alternative method for growing high quality protein crystals in microgravity. Results from both vapor diffusion and temperature induced crystallization experiments indicate that proteins grown in microgravity may be larger, display more uniform morphologies, and yield diffraction data to significantly higher resolutions than the best crystals of these proteins grown on earth.

  15. Synthesis, characterization, and structure-property relationships in two new polar oxides: Zn2(MoO4)(SeO3) and Zn2(MoO4)(TeO3).

    PubMed

    Nguyen, Sau Doan; Kim, Sang-Hwan; Halasyamani, P Shiv

    2011-06-01

    Two new noncentrosymmetric (NCS) polar oxide materials, Zn(2)(MoO(4))(AO(3)) (A = Se(4+) or Te(4+)), have been synthesized by hydrothermal and solid-state techniques. Their crystal structures have been determined, and characterization of their functional properties (second-harmonic generation, piezoelectricity, and polarization) has been performed. The isostructural materials exhibit a three-dimensional network consisting of ZnO(4), ZnO(6), MoO(4), and AO(3) polyhedra that share edges and corners. Powder second-harmonic generation (SHG) measurements using 1064 nm radiation indicate the materials exhibit moderate SHG efficiencies of 100 × and 80 × ?-SiO(2) for Zn(2)(MoO(4))(SeO(3)) and Zn(2)(MoO(4))(TeO(3)), respectively. Particle size vs SHG efficiency measurements indicate the materials are type 1 non-phase-matchable. Converse piezoelectric measurements resulted in d(33) values of ?14 and ?30 pm/V for Zn(2)(MoO(4))(SeO(3)) and Zn(2)(MoO(4))(TeO(3)), respectively, whereas pyroelectric measurements revealed coefficients of -0.31 and -0.64 ?C/m(2) K at 55 °C for Zn(2)(MoO(4))(SeO(3)) and Zn(2)(MoO(4))(TeO(3)), respectively. Frequency-dependent polarization measurements confirmed that all of the materials are nonferroelectric; that is, the macroscopic polarization is not reversible, or "switchable". Infrared, UV-vis, thermogravimetric, and differential thermal analysis measurements were also performed. First-principles density functional theory (DFT) electronic structure calculations were also done. Crystal data: Zn(2)(MoO(4))(SeO(3)), monoclinic, space group P2(1) (No. 4), a = 5.1809(4) Ĺ, b = 8.3238(7) Ĺ, c = 7.1541(6) Ĺ, ? = 99.413(1)°, V = 305.2(1) Ĺ(3), Z = 2; Zn(2)(MoO(4))(TeO(3)), monoclinic, space group P2(1) (No. 4), a = 5.178(4) Ĺ, b = 8.409(6) Ĺ, c = 7.241(5) Ĺ, ? = 99.351(8)°, V = 311.1(4) Ĺ(3), Z = 2. PMID:21557565

  16. Microhydration of the selenite dianion: a theoretical study of structures, hydration energies, and electronic stabilities of SeO(3)(2-)(H(2)O)(n) (n = 0-6, 9) clusters.

    PubMed

    Wicke, Henryk; Meleshyn, Artur

    2010-09-01

    In extension of the ongoing investigations of oxyanion-water clusters, we studied energetically low-lying configurations of hydrated selenite dianion (and in select cases, SeO(3)(-)) clusters using density functional theory (B3LYP, M05-2X, PBE0) and second-order Mřller-Plesset perturbation theory (MP2). Water molecules doubly hydrogen bond to the selenite oxygens for n Se-O bond length of 1.69-1.71 A and selenite tetrahedron height of 0.64-0.73 A are in accordance with recent experimental results for selenite in aqueous solution or adsorbed on calcite. Structural perturbations due to the hydration are accompanied by a considerable charge transfer (up to 0.55|e|) from the selenite substructure to the water molecules. Furthermore, the calculated electron binding energies evidence that selenite-water clusters are electronically stable only for n >or= 4 (according to M05-2X) or n >or= 5 (according to B3LYP and PBE0). The hitherto unknown hydration free energy of selenite was calculated using a cluster/continuum approach to fall into the range from -224.6 to -245.5 kcal/mol depending on the applied continuum solvation model. PMID:20690659

  17. Enhanced sensitivity to conformation in various proteins. Vibrational circular dichroism results

    SciTech Connect

    Pancoska, P.; Yasui, S.C.; Keiderling, T.A. (Univ. of Illinois, Chicago (USA))

    1989-07-11

    Vibrational circular dichroism (VCD) spectra of several globular proteins dissolved in D2O are presented and compared to conventional UV-CD results. It can be seen that, for the alpha, beta, and alpha + beta categories of Levitt and Chothia, VCD evidences much larger band shape variations, including sign alteration, than does UV-CD. A direct parallel is seen between the VCD of the alpha-helix found in model polypeptides and the amide I' VCD of myoglobin. Since all structural aspects of the protein contribute to the VCD on a roughly equal footing, a similar correlation of the chymotrypsin amide I' VCD with that of beta-sheet models is not as clear. In addition, the VCD of random-coil-type proteins is found to be clearly related to VCD results from random-coil polypeptides. Finally, simulations are presented to postulate the expected VCD for protein structures having conformations that lie between the limiting cases discussed here.

  18. Ablation of retinal ciliopathy protein RPGR results in altered photoreceptor ciliary composition

    PubMed Central

    Rao, Kollu N.; Li, Linjing; Anand, Manisha; Khanna, Hemant

    2015-01-01

    Cilia regulate several developmental and homeostatic pathways that are critical to survival. Sensory cilia of photoreceptors regulate phototransduction cascade for visual processing. Mutations in the ciliary protein RPGR (retinitis pigmentosa GTPase regulator) are a prominent cause of severe blindness disorders due to degeneration of mature photoreceptors. However, precise function of RPGR is still unclear. Here we studied the involvement of RPGR in ciliary trafficking by analyzing the composition of photoreceptor sensory cilia (PSC) in Rpgrko retina. Using tandem mass spectrometry analysis followed by immunoblotting, we detected few alterations in levels of proteins involved in proteasomal function and vesicular trafficking in Rpgrko PSC, prior to onset of degeneration. We also found alterations in the levels of high molecular weight soluble proteins in Rpgrko PSC. Our data indicate RPGR regulates entry or retention of soluble proteins in photoreceptor cilia but spares the trafficking of key structural and phototransduction-associated proteins. Given a frequent occurrence of RPGR mutations in severe photoreceptor degeneration due to ciliary disorders, our results provide insights into pathways resulting in altered mature cilia function in ciliopathies. PMID:26068394

  19. Analyzing Proteomes and Protein Function Using Graphical Comparative Analysis of Tandem Mass Spectrometry Results

    Microsoft Academic Search

    K. Jill McAfee; Dexter T. Duncan; Michael Assink; Andrew J. Link

    2006-01-01

    Although generating large amounts of proteomic data us- ing tandem mass spectrometry has become routine, there is currently no single set of comprehensive tools for the rigorous analysis of tandem mass spectrometry results given the large variety of possible experimental aims. Cur- rently available applications are typically designed for dis- playing proteins and posttranslational modifications from the point of view

  20. Phylogenetic continuum indicates "galaxies" in the protein universe: preliminary results on the natural group structures of proteins.

    PubMed

    Ladunga, I

    1992-04-01

    The markedly nonuniform, even systematic distribution of sequences in the protein "universe" has been analyzed by methods of protein taxonomy. Mapping of the natural hierarchical system of proteins has revealed some dense cores, i.e., well-defined clusterings of proteins that seem to be natural structural groupings, possibly seeds for a future protein taxonomy. The aim was not to force proteins into more or less man-made categories by discriminant analysis, but to find structurally similar groups, possibly of common evolutionary origin. Single-valued distance measures between pairs of superfamilies from the Protein Identification Resource were defined by two chi 2-like methods on tripeptide frequencies and the variable-length subsequence identity method derived from dot-matrix comparisons. Distance matrices were processed by several methods of cluster analysis to detect phylogenetic continuum between highly divergent proteins. Only well-defined clusters characterized by relatively unique structural, intracellular environmental, organismal, and functional attribute states were selected as major protein groups, including subsets of viral and Escherichia coli proteins, hormones, inhibitors, plant, ribosomal, serum and structural proteins, amino acid synthases, and clusters dominated by certain oxidoreductases and apolar and DNA-associated enzymes. The limited repertoire of functional patterns due to small genome size, the high rate of recombination, specific features of the bacterial membranes, or of the virus cycle canalize certain proteins of viruses and Gram-negative bacteria, respectively, to organismal groups. PMID:1569589

  1. Developmental expression of MARCKS and protein kinase C in mice in relation to the exencephaly resulting from MARCKS deficiency

    Microsoft Academic Search

    Perry J. Blackshear; Wi S. Lai; Jane S. Tuttle; Deborah J. Stumpo; Elizabeth Kennington; Angus C. Nairn; Kathleen K. Sulik

    1996-01-01

    The roles of protein kinase C and its substrates in development are poorly understood. Recently, we disrupted the mouse gene for a major cellular substrate for protein kinase C, the MARCKS protein (Proc. Natl. Acad. Sci. USA, 92, 944–948, 1995). The resulting phenotype consisted of universal perinatal lethality, agenesis of the corpus callosum and other forebrain commissures, and neuronal ectopia

  2. Pokeweed antiviral protein alters splicing of HIV-1 RNAs, resulting in reduced virus production.

    PubMed

    Zhabokritsky, Alice; Mansouri, Sheila; Hudak, Katalin A

    2014-08-01

    Processing of HIV-1 transcripts results in three populations in the cytoplasm of infected cells: full-length RNA, singly spliced, and multiply spliced RNAs. Rev, regulator of virion expression, is an essential regulatory protein of HIV-1 required for transporting unspliced and singly spliced viral transcripts from the nucleus to the cytoplasm. Export allows these RNAs to be translated and the full-length RNA to be packaged into virus particles. In our study, we investigate the activity of pokeweed antiviral protein (PAP), a glycosidase isolated from the pokeweed plant Phytolacca americana, on the processing of viral RNAs. We show that coexpression of PAP with a proviral clone alters the splicing ratio of HIV-1 RNAs. Specifically, PAP causes the accumulation of multiply spliced 2-kb RNAs at the expense of full-length 9-kb and singly spliced 4-kb RNAs. The change in splicing ratio is due to a decrease in activity of Rev. We show that PAP depurinates the rev open reading frame and that this damage to the viral RNA inhibits its translation. By decreasing Rev expression, PAP indirectly reduces the availability of full-length 9-kb RNA for packaging and translation of the encoded structural proteins required for synthesis of viral particles. The decline we observe in virus protein expression is not due to cellular toxicity as PAP did not diminish translation rate. Our results describing the reduced activity of a regulatory protein of HIV-1, with resulting change in virus mRNA ratios, provides new insight into the antiviral mechanism of PAP. PMID:24951553

  3. Diurnal Rhythms Result in Significant Changes in the Cellular Protein Complement in the Cyanobacterium Cyanothece 51142

    SciTech Connect

    Stockel, Jana; Jacobs, Jon M.; Elvitigala, Thanura R.; Liberton, Michelle L.; Welsh, Eric A.; Polpitiya, Ashoka D.; Gritsenko, Marina A.; Nicora, Carrie D.; Koppenaal, David W.; Smith, Richard D.; Pakrasi, Himadri B.

    2011-02-22

    Cyanothece sp. ATCC 51142 is a diazotrophic cyanobacterium notable for its ability to perform oxygenic photosynthesis and dinitrogen fixation in the same single cell. Previous transcriptional analysis revealed that the existence of these incompatible cellular processes largely depends on tightly synchronized expression programs involving ,30% of genes in the genome. To expand upon current knowledge, we have utilized sensitive proteomic approaches to examine the impact of diurnal rhythms on the protein complement in Cyanothece 51142. We found that 250 proteins accounting for,5% of the predicted ORFs from the Cyanothece 51142 genome and 20% of proteins detected under alternating light/dark conditions exhibited periodic oscillations in their abundances. Our results suggest that altered enzyme activities at different phases during the diurnal cycle can be attributed to changes in the abundance of related proteins and key compounds. The integration of global proteomics and transcriptomic data further revealed that post-transcriptional events are important for temporal regulation of processes such as photosynthesis in Cyanothece 51142. This analysis is the first comprehensive report on global quantitative proteomics in a unicellular diazotrophic cyanobacterium and uncovers novel findings about diurnal rhythms.

  4. Aging results in an unusual expression of Drosophila heat shock proteins

    SciTech Connect

    Fleming, J.E.; Walton, J.K.; Dubitsky, R.; Bensch, K.G. (Linus Pauling Institute of Science and Medicine, Palo Alto, CA (USA))

    1988-06-01

    The authors used high-resolution two-dimensional polyacrylamide gel electrophoresis to evaluate the effect of aging on the heat shock response in Drosophila melanogaster. Although the aging process is not well understood at the molecular level, recent observations suggest that quantitative changes in gene expression occur as these fruit flies approach senescence. Such genetic alterations are in accord with our present data, which clearly show marked differences in the synthesis of heat shock proteins between young and old fruit flies. In 10-day-old flies, a heat shock of 20 min results in the expression of 14 new proteins as detectable by two-dimensional electrophoresis of ({sup 35}S)methionine-labeled polypeptides, whereas identical treatment of 45-day-old flies leads to the expression of at least 50 new or highly up-regulated proteins. In addition, there is also a concomitant increase in the rate of synthesis of a number of the normal proteins in the older animals. Microdensitometric determinations of the low molecular weight heat shock polypeptides on autoradiographs of five age groups revealed that their maximum expression occurs at 47 days for a population of flies with a mean life span of 33.7 days. Moreover, a heat shock effect similar to that observed in senescent flies occurs in young flies fed canavanine, an arginine analogue, before heat shock.

  5. Preparation and characterization of SeO2/TiO2 composite photocatalyst with excellent performance for sunset yellow azo dye degradation under natural sunlight illumination

    NASA Astrophysics Data System (ADS)

    Rajamanickam, D.; Dhatshanamurthi, P.; Shanthi, M.

    2015-03-01

    To improve the solar light induced photocatalytic application performances of TiO2, in this study, the SeO2 modified TiO2 composite photocatalysts with various ratios of SeO2 to TiO2 were prepared by sol-gel method. The catalyst was characterized by X-ray diffraction (XRD), high resolution scanning electron microscope (HR-SEM), energy dispersive spectra (EDS), diffuse reflectance spectra (DRS), photoluminescence spectra (PL), X-ray photoelectron spectroscopy (XPS) and Brunauer-Emmett-Teller (BET) surface area measurement methods. The photocatalytic activity of SeO2/TiO2 was investigated for the degradation of sunset yellow (SY) in aqueous solution using solar light. The SeO2/TiO2 is found to be more efficient than prepared TiO2 and TiO2-P25 at pH 7 for the mineralization of SY. The effects of operational parameters such as the amount of photocatalyst, dye concentration and initial pH on photo mineralization of SY have been analyzed. The degradation was strongly enhanced in the presence of electron acceptors such as oxone, KIO4 and KBrO3. The kinetics of SY photodegradation was found to follow the pseudo-first order rate law and could be described in terms of Langmuir-Hinshelwood model. The mineralization of SY has been confirmed by COD measurements. The catalyst is found to be reusable.

  6. Crystallization Optimum Solubility Screening: using crystallization results to identify the optimal buffer for protein crystal formation.

    PubMed

    Collins, Bernard; Stevens, Raymond C; Page, Rebecca

    2005-12-01

    An optimal solubility screen is described that uses the results of crystallization trials to identify buffers that improve protein solubility and, in turn, crystallization success. This screen is useful not only for standard crystallization experiments, but also can easily be implemented into any high-throughput structure-determination pipeline. As a proof of principle, the predicted novel-fold protein AF2059 from Archaeoglobus fulgidus, which was known to precipitate in most buffers and particularly during concentration experiments, was selected. Using the crystallization results of 192 independent crystallization trials, it was possible to identify a buffer containing 100 mM CHES pH 9.25 that significantly improves its solubility. After transferring AF2059 into this ;optimum-solubility' buffer, the protein was rescreened for crystal formation against these same 192 conditions. Instead of extensive precipitation, as observed initially, it was found that 24 separate conditions produced crystals and the exchange of AF2059 into CHES buffer significantly improved crystallization success. Fine-screen optimization of these conditions led to the production of a crystal suitable for high-resolution (2.2 A) structure determination. PMID:16511228

  7. Protein crystal growth results from the United States Microgravity Laboratory-1 mission

    NASA Technical Reports Server (NTRS)

    Delucas, Lawrence J.; Moore, K. M.; Vanderwoerd, M.; Bray, T. L.; Smith, C.; Carson, M.; Narayana, S. V. L.; Rosenblum, W. M.; Carter, D.; Clark, A. D, Jr.

    1994-01-01

    Protein crystal growth experiments have been performed by this laboratory on 18 Space Shuttle missions since April, 1985. In addition, a number of microgravity experiments also have been performed and reported by other investigators. These Space Shuttle missions have been used to grow crystals of a variety of proteins using vapor diffusion, liquid diffusion, and temperature-induced crystallization techniques. The United States Microgravity Laboratory - 1 mission (USML-1, June 25 - July 9, 1992) was a Spacelab mission dedicated to experiments involved in materials processing. New protein crystal growth hardware was developed to allow in orbit examination of initial crystal growth results, the knowledge from which was used on subsequent days to prepare new crystal growth experiments. In addition, new seeding hardware and techniques were tested as well as techniques that would prepare crystals for analysis by x-ray diffraction, a capability projected for the planned Space Station. Hardware that was specifically developed for the USML-1 mission will be discussed along with the experimental results from this mission.

  8. Divergent Evolution of CHD3 Proteins Resulted in MOM1 Refining Epigenetic Control in Vascular Plants

    PubMed Central

    ?aikovski, Marian; Yokthongwattana, Chotika; Habu, Yoshiki; Nishimura, Taisuke; Mathieu, Olivier; Paszkowski, Jerzy

    2008-01-01

    Arabidopsis MOM1 is required for the heritable maintenance of transcriptional gene silencing (TGS). Unlike many other silencing factors, depletion of MOM1 evokes transcription at selected loci without major changes in DNA methylation or histone modification. These loci retain unusual, bivalent chromatin properties, intermediate to both euchromatin and heterochromatin. The structure of MOM1 previously suggested an integral nuclear membrane protein with chromatin-remodeling and actin-binding activities. Unexpected results presented here challenge these presumed MOM1 activities and demonstrate that less than 13% of MOM1 sequence is necessary and sufficient for TGS maintenance. This active sequence encompasses a novel Conserved MOM1 Motif 2 (CMM2). The high conservation suggests that CMM2 has been the subject of strong evolutionary pressure. The replacement of Arabidopsis CMM2 by a poplar motif reveals its functional conservation. Interspecies comparison suggests that MOM1 proteins emerged at the origin of vascular plants through neo-functionalization of the ubiquitous eukaryotic CHD3 chromatin remodeling factors. Interestingly, despite the divergent evolution of CHD3 and MOM1, we observed functional cooperation in epigenetic control involving unrelated protein motifs and thus probably diverse mechanisms. PMID:18725928

  9. Correlating labeling chemistry and in-vitro test results with the biological behavior of radiolabeled proteins

    SciTech Connect

    Srivastava, S.C.; Meinken, G.E.

    1985-01-01

    Monoclonal antibodies possess enormous potential for delivery of therapeutic amounts of radionuclides to target antigens in vivo, in particular for tumor imaging and therapy. Translation of this concept into practice has encountered numerous problems. Specifically whereas general protein radiolabeling methods are applicable to antibodies, immunological properties of the antibodies are often compromised resulting in reduced in-vivo specificity for the target antigens. The bifunctional chelating agent approach shows the most promise, however, development of other agents will be necessary for widespread usefulness of this technique. The effects of labeling chemistry on the in-vivo behavior of several monoclonal antibodies are described. 30 refs., 4 figs., 10 tabs.

  10. The relationship between dietary protein intake and blood pressure: results from the PREMIER study

    Microsoft Academic Search

    Y F Wang; WS Yancy Jr; D Yu; C Champagne; L J Appel; P-H Lin

    2008-01-01

    Observational and clinical studies suggest that high protein intake, particularly protein from plant sources, might reduce blood pressure (BP). To examine the association of dietary protein with BP, we analysed data from PREMIER, an 18-month clinical trial (n=810) that examined the effects of two multi-component lifestyle modifications on BP. We examined the association of protein intake with BP, and in

  11. The sieve element occlusion gene family in dicotyledonous plants

    PubMed Central

    Jekat, Stephan B; Nordzieke, Steffen; Reineke, Anna R; Müller, Boje; Bornberg-Bauer, Erich; Noll, Gundula A

    2011-01-01

    Sieve element occlusion (SEO) genes encoding forisome subunits have been identified in Medicago truncatula and other legumes. Forisomes are structural phloem proteins uniquely found in Fabaceae sieve elements. They undergo a reversible conformational change after wounding, from a condensed to a dispersed state, thereby blocking sieve tube translocation and preventing the loss of photoassimilates. Recently, we identified SEO genes in several non-Fabaceae plants (lacking forisomes) and concluded that they most probably encode conventional non-forisome P-proteins. Molecular and phylogenetic analysis of the SEO gene family has identified domains that are characteristic for SEO proteins. Here, we extended our phylogenetic analysis by including additional SEO genes from several diverse species based on recently published genomic data. Our results strengthen the original assumption that SEO genes seem to be widespread in dicotyledonous angiosperms, and further underline the divergent evolution of SEO genes within the Fabaceae. PMID:21422825

  12. Biochemical and biophysical characterization of the selenium-binding and reducing site in Arabidopsis thaliana homologue to mammals selenium-binding protein 1.

    PubMed

    Schild, Florie; Kieffer-Jaquinod, Sylvie; Palencia, Andrés; Cobessi, David; Sarret, Géraldine; Zubieta, Chloé; Jourdain, Agnčs; Dumas, Renaud; Forge, Vincent; Testemale, Denis; Bourguignon, Jacques; Hugouvieux, Véronique

    2014-11-14

    The function of selenium-binding protein 1 (SBP1), present in almost all organisms, has not yet been established. In mammals, SBP1 is known to bind the essential element selenium but the binding site has not been identified. In addition, the SBP family has numerous potential metal-binding sites that may play a role in detoxification pathways in plants. In Arabidopsis thaliana, AtSBP1 over-expression increases tolerance to two toxic compounds for plants, selenium and cadmium, often found as soil pollutants. For a better understanding of AtSBP1 function in detoxification mechanisms, we investigated the chelating properties of the protein toward different ligands with a focus on selenium using biochemical and biophysical techniques. Thermal shift assays together with inductively coupled plasma mass spectrometry revealed that AtSBP1 binds selenium after incubation with selenite (SeO3(2-)) with a ligand to protein molar ratio of 1:1. Isothermal titration calorimetry confirmed the 1:1 stoichiometry and revealed an unexpectedly large value of binding enthalpy suggesting a covalent bond between selenium and AtSBP1. Titration of reduced Cys residues and comparative mass spectrometry on AtSBP1 and the purified selenium-AtSBP1 complex identified Cys(21) and Cys(22) as being responsible for the binding of one selenium. These results were validated by site-directed mutagenesis. Selenium K-edge x-ray absorption near edge spectroscopy performed on the selenium-AtSBP1 complex demonstrated that AtSBP1 reduced SeO3(2-) to form a R-S-Se(II)-S-R-type complex. The capacity of AtSBP1 to bind different metals and selenium is discussed with respect to the potential function of AtSBP1 in detoxification mechanisms and selenium metabolism. PMID:25274629

  13. Next-generation protein-rich potato expressing the seed protein gene AmA1 is a result of proteome rebalancing in transgenic tuber

    PubMed Central

    Chakraborty, Subhra; Chakraborty, Niranjan; Agrawal, Lalit; Ghosh, Sudip; Narula, Kanika; Shekhar, Shubhendu; Naik, Prakash S.; Pande, P. C.; Chakrborti, Swarup Kumar; Datta, Asis

    2010-01-01

    Protein deficiency is the most crucial factor that affects physical growth and development and that increases morbidity and mortality especially in developing countries. Efforts have been made to improve protein quality and quantity in crop plants but with limited success. Here, we report the development of transgenic potatoes with enhanced nutritive value by tuber-specific expression of a seed protein, AmA1 (Amaranth Albumin 1), in seven genotypic backgrounds suitable for cultivation in different agro-climatic regions. Analyses of the transgenic tubers revealed up to 60% increase in total protein content. In addition, the concentrations of several essential amino acids were increased significantly in transgenic tubers, which are otherwise limited in potato. Moreover, the transgenics also exhibited enhanced photosynthetic activity with a concomitant increase in total biomass. These results are striking because this genetic manipulation also resulted in a moderate increase in tuber yield. The comparative protein profiling suggests that the proteome rebalancing might cause increased protein content in transgenic tubers. Furthermore, the data on field performance and safety evaluation indicate that the transgenic potatoes are suitable for commercial cultivation. In vitro and in vivo studies on experimental animals demonstrate that the transgenic tubers are also safe for human consumption. Altogether, these results emphasize that the expression of AmA1 is a potential strategy for the nutritional improvement of food crops. PMID:20855595

  14. Probity: A Protein Identification Algorithm with Accurate Assignment of the Statistical Significance of the Results

    E-print Network

    Chait, Brian T.

    . State-of-the-art proteome analysis often involves protein separation by 2D-gel electrophoresis.7-10 The level of a protein is typically determined from the intensity of the spot on the gel. Each gel-spot of interest is excised and the protein is subjected to in-gel digestion using an enzyme that has high

  15. Altered protein prenylation in Sertoli cells is associated with adult infertility resulting from childhood mumps infection

    PubMed Central

    Wang, Xiu-Xing; Ying, Pu; Diao, Fan; Wang, Qiang; Ye, Dan; Jiang, Chen; Shen, Ning; Xu, Na; Chen, Wei-Bo; Lai, Shan-Shan; Jiang, Shan; Miao, Xiao-Li; Feng, Jin; Tao, Wei-Wei; Zhao, Ning-Wei; Yao, Bing; Xu, Zhi-Peng; Sun, Hai-Xiang; Sha, Jia-Hao; Huang, Xing-Xu; Shi, Qing-Hua; Tang, Hong

    2013-01-01

    Mumps commonly affects children 5–9 yr of age, and can lead to permanent adult sterility in certain cases. However, the etiology of this long-term effect remains unclear. Mumps infection results in progressive degeneration of the seminiferous epithelium and, occasionally, Sertoli cell–only syndrome. Thus, the remaining Sertoli cells may be critical to spermatogenesis recovery after orchitis healing. Here, we report that the protein farnesylation/geranylgeranylation balance is critical for patients’ fertility. The expression of geranylgeranyl diphosphate synthase 1 (GGPPS) was decreased due to elevated promoter methylation in the testes of infertile patients with mumps infection history. When we deleted GGPPS in mouse Sertoli cells, these cells remained intact, whereas the adjacent spermatogonia significantly decreased after the fifth postnatal day. The proinflammatory MAPK and NF-?B signaling pathways were constitutively activated in GGPPS?/? Sertoli cells due to the enhanced farnesylation of H-Ras. GGPPS?/? Sertoli cells secreted an array of cytokines to stimulate spermatogonia apoptosis, and chemokines to induce macrophage invasion into the seminiferous tubules. Invaded macrophages further blocked spermatogonia development, resulting in a long-term effect through to adulthood. Notably, this defect could be rescued by GGPP administration in EMCV-challenged mice. Our results suggest a novel mechanism by which mumps infection during childhood results in adult sterility. PMID:23825187

  16. SANS study of interaction of silica nanoparticles with BSA protein and their resultant structure

    SciTech Connect

    Yadav, Indresh, E-mail: vkaswal@barc.gov.in; Aswal, V. K., E-mail: vkaswal@barc.gov.in [Solid State Physics Division, Bhabha Atomic Research Centre, Mumbai-400085 (India); Kohlbrecher, J. [Laboratory for Neutron Scattering, Paul Scherrer Institute, CH-5232 PSI Villigen Switzerland (Switzerland)

    2014-04-24

    Small angle neutron scattering (SANS) has been carried out to study the interaction of anionic silica nanoparticles (88 Ĺ) with globular protein Bovine Serum Albumin (BSA) (M.W. 66.4 kD) in aqueous solution. The measurements have been carried out on fixed concentration (1 wt %) of Ludox silica nanoparticles with varying concentration of BSA (0–5 wt %) at pH7. Results show that silica nanoparticles and BSA coexist as individual entities at low concentration of BSA where electrostatic repulsive interactions between them prevent their aggregation. However, as the concentration of BSA increases (? 0.5 wt %), it induces the attractive depletion interaction among nanoparticles leading to finally their aggregation at higher BSA concentration (2 wt %). The aggregates are found to be governed by the diffusion limited aggregation (DLA) morphology of fractal nature having fractal dimension about 2.4.

  17. Experimental validation of FINDSITEcomb virtual ligand screening results for eight proteins yields novel nanomolar and micromolar binders

    PubMed Central

    2014-01-01

    Background Identification of ligand-protein binding interactions is a critical step in drug discovery. Experimental screening of large chemical libraries, in spite of their specific role and importance in drug discovery, suffer from the disadvantages of being random, time-consuming and expensive. To accelerate the process, traditional structure- or ligand-based VLS approaches are combined with experimental high-throughput screening, HTS. Often a single protein or, at most, a protein family is considered. Large scale VLS benchmarking across diverse protein families is rarely done, and the reported success rate is very low. Here, we demonstrate the experimental HTS validation of a novel VLS approach, FINDSITEcomb, across a diverse set of medically-relevant proteins. Results For eight different proteins belonging to different fold-classes and from diverse organisms, the top 1% of FINDSITEcomb’s VLS predictions were tested, and depending on the protein target, 4%-47% of the predicted ligands were shown to bind with ?M or better affinities. In total, 47 small molecule binders were identified. Low nanomolar (nM) binders for dihydrofolate reductase and protein tyrosine phosphatases (PTPs) and micromolar binders for the other proteins were identified. Six novel molecules had cytotoxic activity (<10 ?g/ml) against the HCT-116 colon carcinoma cell line and one novel molecule had potent antibacterial activity. Conclusions We show that FINDSITEcomb is a promising new VLS approach that can assist drug discovery. PMID:24936211

  18. Human amyloidogenic light chain proteins result in cardiac dysfunction, cell death, and early mortality in zebrafish

    PubMed Central

    Mishra, Shikha; Guan, Jian; Plovie, Eva; Seldin, David C.; Connors, Lawreen H.; Merlini, Giampaolo; Falk, Rodney H.; MacRae, Calum A.

    2013-01-01

    Systemic amyloid light-chain (AL) amyloidosis is associated with rapidly progressive and fatal cardiomyopathy resulting from the direct cardiotoxic effects of circulating AL light chain (AL-LC) proteins and the indirect effects of AL fibril tissue infiltration. Cardiac amyloidosis is resistant to standard heart failure therapies, and, to date, there are limited treatment options for these patients. The mechanisms underlying the development of cardiac amyloidosis and AL-LC cardiotoxicity are largely unknown, and their study has been limited by the lack of a suitable in vivo model system. Here, we establish an in vivo zebrafish model of human AL-LC-induced cardiotoxicity. AL-LC isolated from AL cardiomyopathy patients or control nonamyloidogenic LC protein isolated from multiple myeloma patients (Con-LC) was directly injected into the circulation of zebrafish at 48 h postfertilization. AL-LC injection resulted in impaired cardiac function, pericardial edema, and increased cell death relative to Con-LC, culminating in compromised survival with 100% mortality within 2 wk, independent of AL fibril deposition. Prior work has implicated noncanonical p38 MAPK activation in the pathogenesis of AL-LC-induced cardiotoxicity, and p38 MAPK inhibition via SB-203580 rescued AL-LC-induced cardiac dysfunction and cell death and attenuated mortality in zebrafish. This in vivo zebrafish model of AL-LC cardiotoxicity demonstrates that antagonism of p38 MAPK within the AL-LC cardiotoxic signaling response may serve to improve cardiac function and mortality in AL cardiomyopathy. Furthermore, this in vivo model system will allow for further study of the molecular underpinnings of AL cardiotoxicity and identification of novel therapeutic strategies. PMID:23624626

  19. 39K and 77Se NMR study of the paraelectric-to-incommensurate phase transition of K2SeO4

    NASA Astrophysics Data System (ADS)

    Topi?, B.; von Kienlin, A.; Gölzhäuser, A.; Haeberlen, U.; Blinc, R.

    1988-11-01

    The 39K quadrupole-coupling and chemical-shift tensors have been determined from the angular dependences of the 39K line shifts of the 39K+/-(1/2?-/+ 1) / 2 central NMR transitions in the paraelectric (P) and incommensurate (I) phases of K2SeO4. The main effect of the P-I phase transition on these tensors is the appearance of nonzero off-diagonal elements Vab and Vbc which reflects the destruction of mirror planes by frozen-in soft-mode displacements along the b axis. From the angular dependences of the 77Se line shifts the 77Se chemical-shift tensor has been determined in the paraelectric phase of K2SeO4. In contrast to the 39K quadrupole-coupling and chemical-shift tensors it remains unaffected on going through TI and changes only slightly at TC.

  20. Transiting Exoplanet Survey Satellite (TESS) Community Observer Program including the Science Enhancement Option Box (SEO Box) - 12 TB On-board Flash Memory for Serendipitous Science

    NASA Astrophysics Data System (ADS)

    Schingler, Robert; Villasenor, J. N.; Ricker, G. R.; Latham, D. W.; Vanderspek, R. K.; Ennico, K. A.; Lewis, B. S.; Bakos, G.; Brown, T. M.; Burgasser, A. J.; Charbonneau, D.; Clampin, M.; Deming, L. D.; Doty, J. P.; Dunham, E. W.; Elliot, J. L.; Holman, M. J.; Ida, S.; Jenkins, J. M.; Jernigan, J. G.; Kawai, N.; Laughlin, G. P.; Lissauer, J. J.; Martel, F.; Sasselov, D. D.; Seager, S.; Torres, G.; Udry, S.; Winn, J. N.; Worden, S. P.

    2010-01-01

    The Transiting Exoplanet Survey Satellite (TESS) will perform an all-sky survey in a low-inclination, low-Earth orbit. TESS's 144 GB of raw data collected each orbit will be stacked, cleaned, cut, compressed and downloaded. The Community Observer Program is a Science Enhancement Option (SEO) that takes advantage of the low-radiation environment, technology advances in flash memory, and the vast amount of astronomical data collected by TESS. The Community Observer Program requires the addition of a 12 TB "SEO Box” inside the TESS Bus. The hardware can be built using low-cost Commercial Off-The-Shelf (COTS) components and fits within TESS's margins while accommodating GSFC gold rules. The SEO Box collects and stores a duplicate of the TESS camera data at a "raw” stage ( 4.3 GB/orbit, after stacking and cleaning) and makes them available for on-board processing. The sheer amount of onboard storage provided by the SEO Box allows the stacking and storing of several months of data, allowing the investigator to probe deeper in time prior to a given event. Additionally, with computation power and data in standard formats, investigators can utilize data-mining techniques to investigate serendipitous phenomenon, including pulsating stars, eclipsing binaries, supernovae or other transient phenomena. The Community Observer Program enables ad-hoc teams of citizen scientists to propose, test, refine and rank algorithms for on-board analysis to support serendipitous science. Combining "best practices” of online collaboration, with careful moderation and community management, enables this `crowd sourced’ participatory exploration with a minimal risk and impact on the core TESS Team. This system provides a powerful and independent tool opening a wide range of opportunity for science enhancement and secondary science. Support for this work has been provided by NASA, the Kavli Foundation, Google, and the Smithsonian Institution.

  1. Transiting Exoplanet Survey Satellite (TESS) Community Observer Program including the Science Enhancement Option Box (SEO Box) - 12 TB On-board Flash Memory for Serendipitous Science

    Microsoft Academic Search

    Robert Schingler; J. N. Villasenor; G. R. Ricker; D. W. Latham; R. K. Vanderspek; K. A. Ennico; B. S. Lewis; G. Bakos; T. M. Brown; A. J. Burgasser; D. Charbonneau; M. Clampin; L. D. Deming; J. P. Doty; E. W. Dunham; J. L. Elliot; M. J. Holman; S. Ida; J. M. Jenkins; J. G. Jernigan; N. Kawai; G. P. Laughlin; J. J. Lissauer; F. Martel; D. D. Sasselov; S. Seager; G. Torres; S. Udry; J. N. Winn; S. P. Worden

    2010-01-01

    The Transiting Exoplanet Survey Satellite (TESS) will perform an all-sky survey in a low-inclination, low-Earth orbit. TESS's 144 GB of raw data collected each orbit will be stacked, cleaned, cut, compressed and downloaded. The Community Observer Program is a Science Enhancement Option (SEO) that takes advantage of the low-radiation environment, technology advances in flash memory, and the vast amount of

  2. Pseudomorphic InGaAs base ballistic hotelectron device K. Seo, M. Heiblum, C. M. Knoedler, WP. Hong, and P. Bhattacharya

    E-print Network

    Heiblum, Mordehai "Moty"

    Pseudomorphic InGaAs base ballistic hotelectron device K. Seo, M. Heiblum, C. M. Knoedler, WP. Hong. Phys. 69, 4454 (1991); 10.1063/1.348378 Enhanced ballistic transport in InGaAs/InAlAs hotelectron://scitation.aip.org/termsconditions. Downloaded to IP: 132.76.50.6 On: Thu, 10 Apr 2014 16:10:12 #12;Pseudomorphic InGaAs base ballistic hot

  3. Synthesis, Crystal Structure and Thermal Decomposition of the New Cadmium Selenite Chloride, Cd4(SeO3)2OCl2

    PubMed Central

    Rabbani, Faiz; Ajaz, Humayun; Zimmermann, Iwan; Johnsson, Mats

    2014-01-01

    A synthetic study in the Cd-Se-O-Cl system led to formation of the new oxochloride compound Cd4(SeO3)2OCl2 via solid state reactions. The compound crystallizes in the orthorhombic space group Fmmm with cell parameters a?=?7.3610(3) Ĺ, b?=?15.4936(2) Ĺ, c?=?17.5603(3) Ĺ, Z?=?8, S?=?0.969, F(000)?=?2800, R?=?0.0185, Rw?=?0.0384. Single crystal X-ray data were collected at 293 K. The crystal structure can be considered as layered and the building units are distorted [Cd(1)O6] octahedra, distorted [Cd(2)O8] cubes, irregular [Cd(3)O4Cl2] polyhedra and SeO3E trigonal pyramids. There are two crystallographically unique Cl atoms that both are half occupied. Thermogravimetric studies show that the compound starts to decompose at 500°C. The crystal structure of the new compound is closely related to the previously described compound Cd4(SeO3)2Cl4(H2O). PMID:24844633

  4. Electrophoretic analysis of sheep plasma protein labeled with Na2 75SeO3 in vivo

    SciTech Connect

    Davidson, W.B.; McMurray, C.H.

    1987-05-01

    Following an intravenous injection of /sup 75/Se, sodium selenite plasma samples were analyzed by two-dimensional electrophoresis. /sup 75/Se was detected by indirect autoradiography. From 0.5 to 53 hr postinjection of /sup 75/Se, 21 /sup 75/Se peptides were detected. Both the isoelectric points and molecular weights of these peptides are reported. The molecular weights of the peptides ranged from 20,000 to 70,000 daltons.

  5. Pooled Results From 5 Validation Studies of Dietary Self-Report Instruments Using Recovery Biomarkers for Energy and Protein Intake

    PubMed Central

    Freedman, Laurence S.; Commins, John M.; Moler, James E.; Arab, Lenore; Baer, David J.; Kipnis, Victor; Midthune, Douglas; Moshfegh, Alanna J.; Neuhouser, Marian L.; Prentice, Ross L.; Schatzkin, Arthur; Spiegelman, Donna; Subar, Amy F.; Tinker, Lesley F.; Willett, Walter

    2014-01-01

    We pooled data from 5 large validation studies of dietary self-report instruments that used recovery biomarkers as references to clarify the measurement properties of food frequency questionnaires (FFQs) and 24-hour recalls. The studies were conducted in widely differing US adult populations from 1999 to 2009. We report on total energy, protein, and protein density intakes. Results were similar across sexes, but there was heterogeneity across studies. Using a FFQ, the average correlation coefficients for reported versus true intakes for energy, protein, and protein density were 0.21, 0.29, and 0.41, respectively. Using a single 24-hour recall, the coefficients were 0.26, 0.40, and 0.36, respectively, for the same nutrients and rose to 0.31, 0.49, and 0.46 when three 24-hour recalls were averaged. The average rate of under-reporting of energy intake was 28% with a FFQ and 15% with a single 24-hour recall, but the percentages were lower for protein. Personal characteristics related to under-reporting were body mass index, educational level, and age. Calibration equations for true intake that included personal characteristics provided improved prediction. This project establishes that FFQs have stronger correlations with truth for protein density than for absolute protein intake, that the use of multiple 24-hour recalls substantially increases the correlations when compared with a single 24-hour recall, and that body mass index strongly predicts under-reporting of energy and protein intakes. PMID:24918187

  6. Protein

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteins are the major structural and functional components of all cells in the body. They are macromolecules that comprise 1 or more chains of amino acids that vary in their sequence and length and are folded into specific 3-dimensional structures. The sizes and conformations of proteins, therefor...

  7. Proteins.

    ERIC Educational Resources Information Center

    Doolittle, Russell F.

    1985-01-01

    Examines proteins which give rise to structure and, by virtue of selective binding to other molecules, make genes. Binding sites, amino acids, protein evolution, and molecular paleontology are discussed. Work with encoding segments of deoxyribonucleic acid (exons) and noncoding stretches (introns) provides new information for hypotheses. (DH)

  8. Heterologous Activation of Protein Kinase C Stimulates Phosphorylation of -Opioid Receptor at Serine 344, Resulting

    E-print Network

    Tian, Weidong

    Heterologous Activation of Protein Kinase C Stimulates Phosphorylation of -Opioid Receptor of the current study is to investigate the effect of opioid-independent, heterologous activation of protein kinase C (PKC) on the responsiveness of opioid receptor and the underlying molecular mechanisms. Our

  9. A loss in cellular protein partners promotes ?-synuclein aggregation in cells resulting from oxidative stress.

    PubMed

    Guo, Yuanjian; Scarlata, Suzanne

    2013-06-01

    There is a consensus that oxidative stress promotes neurodegeneration and may be linked to plaque formation. ?-Synuclein is the main component of neurodegenerative plaques. We have found that ?-synuclein binds strongly to the enzyme phospholipase C?1 (PLC?1) in vitro and in cells affecting both its G protein activation and its degradation. Because PLC?1 binds to ?-synuclein in cells, we tested whether decreasing its level would promote ?-synuclein aggregation and whether overproducing PLC?1 would inhibit aggregation. By imaging fluorescent ?-synuclein in living HEK293, PC12, and SK-H-SH cells, we find that ?-synuclein aggregation is directly related to the level of PLC?1. Importantly, we found that oxidative stress does not affect the cellular levels of ?-synuclein but results in the down-regulation of PLC?1 thereby promoting ?-synuclein aggregation. A peptide that mimics part of the ?-synuclein binding site to PLC? prevents aggregation. Our studies indicate that PLC?1 can reduce cell damage under oxidative stress and offers a potential site that might be exploited to prevent ?-synuclein aggregation. PMID:23659438

  10. Interference with the expression of a novel human polycomb protein, hPc2, results in cellular transformation and apoptosis.

    PubMed Central

    Satijn, D P; Olson, D J; van der Vlag, J; Hamer, K M; Lambrechts, C; Masselink, H; Gunster, M J; Sewalt, R G; van Driel, R; Otte, A P

    1997-01-01

    Polycomb (Pc) is involved in the stable and heritable repression of homeotic gene activity during Drosophila development. Here, we report the identification of a novel human Pc homolog, hPc2. This gene is more closely related to a Xenopus Pc homolog, XPc, than to a previously described human Pc homolog, CBX2 (hPc1). However, the hPc2 and CBX2/hPc1 proteins colocalize in interphase nuclei of human U-2 OS osteosarcoma cells, suggesting that the proteins are part of a common protein complex. To study the functions of the novel human Pc homolog, we generated a mutant protein, delta hPc2, which lacks an evolutionarily conserved C-terminal domain. This C-terminal domain is important for hPc2 function, since the delta hPc2 mutant protein which lacks the C-terminal domain is unable to repress gene activity. Expression of the delta hPc2 protein, but not of the wild-type hPc2 protein, results in cellular transformation of mammalian cell lines as judged by phenotypic changes, altered marker gene expression, and anchorage-independent growth. Specifically in delta hPc2-transformed cells, the expression of the c-myc proto-oncogene is strongly enhanced and serum deprivation results in apoptosis. In contrast, overexpression of the wild-type hPc2 protein results in decreased c-myc expression. Our data suggest that hPc2 is a repressor of proto-oncogene activity and that interference with hPc2 function can lead to derepression of proto-oncogene transcription and subsequently to cellular transformation. PMID:9315667

  11. Loss of Clcc1 results in ER stress, misfolded protein accumulation, and neurodegeneration.

    PubMed

    Jia, Yichang; Jucius, Thomas J; Cook, Susan A; Ackerman, Susan L

    2015-02-18

    Folding of transmembrane and secretory proteins occurs in the lumen of the endoplasmic reticulum (ER) before transportation to the cell surface and is monitored by the unfolded protein response (UPR) signaling pathway. The accumulation of unfolded proteins in the ER activates the UPR that restores ER homeostasis by regulating gene expression that leads to an increase in the protein-folding capacity of the ER and a decrease in the ER protein-folding load. However, prolonged UPR activity has been associated with cell death in multiple pathological conditions, including neurodegeneration. Here, we report a spontaneous recessive mouse mutation that causes progressive cerebellar granule cell death and peripheral motor axon degeneration. By positional cloning, we identify the mutation in this strain as a retrotransposon insertion in the Clcc1 gene, which encodes a putative chloride channel localized to the ER. Furthermore, we demonstrate that the C3H/HeSnJ inbred strain has late onset cerebellar degeneration due to this mutation. Interestingly, acute knockdown of Clcc1 expression in cultured cells increases sensitivity to ER stress. In agreement, GRP78, the major HSP70 family chaperone in the ER, is upregulated in Clcc1-deficient granule cells in vivo, and ubiquitinated proteins accumulate in these neurons before their degeneration. These data suggest that disruption of chloride homeostasis in the ER disrupts the protein-folding capacity of the ER, leading to eventual neuron death. PMID:25698737

  12. Native-sized recombinant spider silk protein produced in metabolically engineered Escherichia coli results in a strong fiber

    PubMed Central

    Xia, Xiao-Xia; Qian, Zhi-Gang; Ki, Chang Seok; Park, Young Hwan; Kaplan, David L.; Lee, Sang Yup

    2010-01-01

    Spider dragline silk is a remarkably strong fiber that makes it attractive for numerous applications. Much has thus been done to make similar fibers by biomimic spinning of recombinant dragline silk proteins. However, success is limited in part due to the inability to successfully express native-sized recombinant silk proteins (250–320 kDa). Here we show that a 284.9 kDa recombinant protein of the spider Nephila clavipes is produced and spun into a fiber displaying mechanical properties comparable to those of the native silk. The native-sized protein, predominantly rich in glycine (44.9%), was favorably expressed in metabolically engineered Escherichia coli within which the glycyl-tRNA pool was elevated. We also found that the recombinant proteins of lower molecular weight versions yielded inferior fiber properties. The results provide insight into evolution of silk protein size related to mechanical performance, and also clarify why spinning lower molecular weight proteins does not recapitulate the properties of native fibers. Furthermore, the silk expression, purification, and spinning platform established here should be useful for sustainable production of natural quality dragline silk, potentially enabling broader applications. PMID:20660779

  13. Protein

    MedlinePLUS

    ... for the heart. Alternatively, a cup of cooked lentils provides about 18 grams of protein and 15 ... eating approximately one daily serving of beans, chickpeas, lentils or peas can increase fullness, which may lead ...

  14. Effect of a High-Protein Diet on Kidney Function in Healthy Adults: Results From the OmniHeart Trial

    PubMed Central

    Juraschek, Stephen P.; Appel, Lawrence J.; Anderson, Cheryl A.M.; Miller, Edgar R.

    2013-01-01

    Background Consumption of a diet high in protein can cause glomerular hyperfiltration, a potentially maladaptive response, which may accelerate the progression of kidney disease. Study Design An ancillary study of the OmniHeart trial, a randomized 3-period crossover feeding trial testing the effects of partial replacement of carbohydrate with protein on kidney function. Setting & Participants Healthy adults (N=164) with prehypertension or stage 1 hypertension at a community-based research clinic with a metabolic kitchen. Intervention Participants were fed each of 3 diets for 6 weeks. Feeding periods were separated by a 2- to 4-week washout period. Weight was held constant on each diet. The 3 diets emphasized carbohydrate, protein, or unsaturated fat; dietary protein was either 15% (carbohydrate and unsaturated fat diets) or 25% (protein diet) of energy intake. Outcomes Fasting serum creatinine, cystatin C, and ?2-microglobulin levels, estimated glomerular filtration rate (eGFR). Measurements Serum creatinine, cystatin C, and ?2-microglobulin collected at the end of each feeding period. Results Baseline cystatin C-based eGFR was 92.0±16.3 (SD) mL/min/1.73 m2. Compared with the carbohydrate and unsaturated fat diets, the protein diet increased cystatin C-based eGFR by ~4 mL/min/1.73 m2 (P < 0.001). The effects of the protein diet on kidney function were independent of changes in blood pressure. There was no significant difference between the carbohydrate and unsaturated fat diets. Limitations Participants did not have kidney disease at baseline. Conclusions A healthy diet rich in protein increased eGFR. Whether long-term consumption of a high-protein diet leads to kidney disease is uncertain. PMID:23219108

  15. Reduced functionality of PSE-like chicken breast meat batter resulting from alterations in protein conformation.

    PubMed

    Li, K; Zhao, Y Y; Kang, Z L; Wang, P; Han, M Y; Xu, X L; Zhou, G H

    2015-01-01

    The objectives of this study were to evaluate protein thermal stability, water-protein interaction, microstructure, and protein conformation between PSE-like and normal chicken breast meat batters. Sixty pale, soft, and exudative (PSE)-like (L*>53, pH24 h<5.7) and 60 normal (46protein and 2% salt, and they were analyzed for the protein changes and the microstructure using differential scanning calorimetry, low-field (LF)-NMR, SEM, and Raman spectroscopy. PSE-like meat batter had lower gel strength, water-holding capacity, and salt-soluble protein extraction (P < 0.05). Heated PSE-like meat batter formed an aggregated gel matrix, while normal meat batter produced a compact gel network with fine, cross-linked strands by many protein filaments. LF-NMR revealed an increase in the water mobility in heated PSE-like meat batter with an increasing amount of loosely bound water (P < 0.05). No significant changes were observed in the electrophoretic patterns of salt-soluble protein extracts by SDS-PAGE. However, differential scanning calorimetry showed that PSE-like meat had greater myosin and sarcoplasmic proteins/collagen denaturation (P < 0.05). In PSE-like meat, actin denaturation was particular evident after salt addition (P < 0.05) using differential scanning calorimetry. Moreover, Raman spectroscopy indicated that PSE-like meat batter had less unfolded ?-helix and ?-sheet structure formation, reduced exposure of hydrophobic and tyrosine residues (P < 0.05), and changes in the microenvironment of aliphatic residues and tryptophan, which affected salt-soluble protein extraction, gel properties, and water-holding capacity. In conclusion, the inferior functional properties of PSE-like meat were attributed to not only myosin denaturation, but also actin denaturation after salt addition and different protein structural states. PMID:25577798

  16. Allelic variants of the amylose extender mutation of maize demonstrate phenotypic variation in starch structure resulting from modified protein-protein interactions.

    PubMed

    Liu, Fushan; Ahmed, Zaheer; Lee, Elizabeth A; Donner, Elizabeth; Liu, Qiang; Ahmed, Regina; Morell, Matthew K; Emes, Michael J; Tetlow, Ian J

    2012-02-01

    Amylose extender (ae(-)) starches characteristically have modified starch granule morphology resulting from amylopectin with reduced branch frequency and longer glucan chains in clusters, caused by the loss of activity of the major starch branching enzyme (SBE), which in maize endosperm is SBEIIb. A recent study with ae(-) maize lacking the SBEIIb protein (termed ae1.1 herein) showed that novel protein-protein interactions between enzymes of starch biosynthesis in the amyloplast could explain the starch phenotype of the ae1.1 mutant. The present study examined an allelic variant of the ae(-) mutation, ae1.2, which expresses a catalytically inactive form of SBEIIb. The catalytically inactive SBEIIb in ae1.2 lacks a 28 amino acid peptide (Val272-Pro299) and is unable to bind to amylopectin. Analysis of starch from ae1.2 revealed altered granule morphology and physicochemical characteristics distinct from those of the ae1.1 mutant as well as the wild-type, including altered apparent amylose content and gelatinization properties. Starch from ae1.2 had fewer intermediate length glucan chains (degree of polymerization 16-20) than ae1.1. Biochemical analysis of ae1.2 showed that there were differences in the organization and assembly of protein complexes of starch biosynthetic enzymes in comparison with ae1.1 (and wild-type) amyloplasts, which were also reflected in the composition of starch granule-bound proteins. The formation of stromal protein complexes in the wild-type and ae1.2 was strongly enhanced by ATP, and broken by phosphatase treatment, indicating a role for protein phosphorylation in their assembly. Labelling experiments with [?-(32)P]ATP showed that the inactive form of SBEIIb in ae1.2 was phosphorylated, both in the monomeric form and in association with starch synthase isoforms. Although the inactive SBEIIb was unable to bind starch directly, it was strongly associated with the starch granule, reinforcing the conclusion that its presence in the granules is a result of physical association with other enzymes of starch synthesis. In addition, an Mn(2+)-based affinity ligand, specific for phosphoproteins, was used to show that the granule-bound forms of SBEIIb in the wild-type and ae1.2 were phosphorylated, as was the granule-bound form of SBEI found in ae1.2 starch. The data strongly support the hypothesis that the complement of heteromeric complexes of proteins involved in amylopectin synthesis contributes to the fine structure and architecture of the starch granule. PMID:22121198

  17. Mutations in the Myelin Protein Zero result in a spectrum of Charcot-Marie-Tooth phenotypes.

    PubMed

    Kocha?ski, A

    2004-05-01

    Initially the Myelin Protein Zero gene was shown to be mutated in the demyelinating form of Charcot-Marie-Tooth disease (CMT1). The vast majority of the mutations in the Myelin Protein Zero gene have been detected in the Charcot-Marie-Tooth (1B) disease, however, some of them were found in patients suffering from congenital hypomyelinating neuropathy and axonal type Charcot-Marie-Tooth disease. In this study, a Charcot-Marie-Tooth disease phenotype diversity associated with different mutations in the MPZ gene, is described. PMID:15298082

  18. A deep intronic mutation in the ankyrin-1 gene causes diminished protein expression resulting in hemolytic anemia in mice.

    PubMed

    Huang, Hua; Zhao, PengXiang; Arimatsu, Kei; Tabeta, Koichi; Yamazaki, Kazuhisa; Krieg, Lara; Fu, Emily; Zhang, Tian; Du, Xin

    2013-10-01

    Linkage between transmembrane proteins and the spectrin-based cytoskeleton is necessary for membrane elasticity of red blood cells. Mutations of the proteins that mediate this linkage result in various types of hemolytic anemia. Here we report a novel N-ethyl-N-nitrosourea-induced mutation of ankyrin-1, named hema6, which causes hereditary spherocytosis in mice through a mild reduction of protein expression. The causal mutation was traced to a single nucleotide transition located deep into intron 13 of gene Ank1. In vitro minigene splicing assay revealed two abnormally spliced transcripts containing cryptic exons from fragments of Ank1 intron 13. The inclusion of cryptic exons introduced a premature termination codon, which leads to nonsense-mediated decay of the mutant transcripts in vivo. Hence, in homozygous mice, only wild-type ankyrin-1 is expressed, albeit at 70% of the level in wild-type mice. Heterozygotes display a similar hereditary spherocytosis phenotype stemming from intermediate protein expression level, indicating the haploinsufficiency of the mutation. Weakened linkage between integral transmembrane protein, band 3, and underlying cytoskeleton was observed in mutant mice as the result of reduced high-affinity binding sites provided by ankyrin-1. Hema6 is the only known mouse mutant of Ank1 allelic series that expresses full-length canonical ankyrin-1 at a reduced level, a fact that makes it particularly useful to study the functional impact of ankyrin-1 quantitative deficiency. PMID:23934996

  19. Overexpression of Drosophila juvenile hormone esterase binding protein results in anti-JH effects and reduced pheromone abundance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The titer of juvenile hormone (JH), which has wide ranging physiological effects in insects, is regulated in part by JH esterase (JHE). We show that overexpression in Drosophila melanogaster of the JHE binding protein, DmP29 results in a series of apparent anti-JH effects. We hypothesize that DmP29 ...

  20. Analysis of proteins in bronchoalveolar lavage fluids during pulmonary edema resulting from nitrogen dioxide and cadmium exposure

    SciTech Connect

    Gurley, L.R.; London, J.E.; Dethloff, L.A.; Lehnert, B.E.

    1988-01-01

    We have developed a new HPLC method by which quantitative measurements can be made on the biochemical constituents of the extracellular fluid lining of the lung as sampled by bronchoalveolar lavage. Nine of the fractions are proteins, two are phospholipids, and two fractions remained unidentified. Rats were subjected to the intrapulmonary deposition of cadmium, a treatment model known to induce pulmonary edema and cause a translocation of blood compartment proteins into the lung's alveolar space compartment. Resulting pulmonary edema was hallmarked by /approximately/25-fold increases in three major blood compartment-derived HPLC protein fractions, two of which have been identified as albumin and immunoglobulin(s). Analysis of lavage fluid from rats exposed to 100 ppM NO/sub 2/ for 15 min, an exposure regimen which also produces pulmonary edema, indicated that the three blood compartment proteins in the lavage fluids were elevated 35- to 72-fold over controls 24 h after exposure. These results demonstrate that HPLC can be used to provide a highly sensitive method for detection and quantitation of pulmonary edema that can occur in acute lung injuries resulting from environmental insults.

  1. Natriuretic peptide receptor-3 gene (NPR3): nonsynonymous polymorphism results in significant reduction in protein expression because of accelerated degradation.

    PubMed

    Pereira, Naveen L; Lin, Dong; Pelleymounter, Linda; Moon, Irene; Stilling, Gail; Eckloff, Bruce W; Wieben, Eric D; Redfield, Margaret M; Burnett, John C; Yee, Vivien C; Weinshilboum, Richard M

    2013-04-01

    BACKGROUND- The primary role of natriuretic peptide receptor-3 (NPR3) or NPR-C is in the clearance of natriuretic peptides that play an important role in modulating intravascular volume and vascular tone. Genetic variation in NPR3 has been associated with variation in blood pressure and obesity. Despite the importance of NPR3, sequence variation in the gene has not been addressed using DNA from different ethnic populations. We set out to identify and functionally characterize genetic variation in NPR3 in 3 ethnic groups. METHODS AND RESULTS- DNA samples from 96 European American, 96 African American, and 96 Han Chinese American healthy subjects were used to resequence NPR3 exons, splice junctions, and flanking regions. We identified 105 polymorphisms, 50 of which were novel, including 8 nonsynonymous single-nucleotide polymorphisms, 7 were novel. Expression constructs were created for the nonsynonymous single-nucleotide polymorphisms. HEK293 cells were transfected with constructs for wild type and variant allozymes; and recombinant proteins were measured by quantitative Western blot analysis. The most significant change in NPR3 protein was observed for the Arg146 variant allozyme, with 20% of wild-type protein, primarily because of autophagy-dependent degradation. NPR3 structural modeling confirmed that the Arg146 variant protein was not compatible with wild-type conformation and could result in protein misfolding or instability. CONCLUSIONS- Multiple novel NPR3 genetic polymorphisms were identified in 3 ethnic groups. The Arg146 allozyme displayed a significant decrease in protein quantity because of degradation mediated predominantly by autophagy. This genetic variation could have a significant effect on the metabolism of natriuretic peptides with potential clinical implications. PMID:23493048

  2. Lack of the Lysosomal Membrane Protein, GLMP, in Mice Results in Metabolic Dysregulation in Liver

    PubMed Central

    Kong, Xiang Yi; Kase, Eili Tranheim; Herskedal, Anette; Schjalm, Camilla; Damme, Markus; Nesset, Cecilie Kasi; Thoresen, G. Hege; Rustan, Arild C.; Eskild, Winnie

    2015-01-01

    Ablation of glycosylated lysosomal membrane protein (GLMP, formerly known as NCU-G1) has been shown to cause chronic liver injury which progresses into liver fibrosis in mice. Both lysosomal dysfunction and chronic liver injury can cause metabolic dysregulation. Glmpgt/gt mice (formerly known as Ncu-g1gt/gtmice) were studied between 3 weeks and 9 months of age. Body weight gain and feed efficiency of Glmpgt/gt mice were comparable to wild type siblings, only at the age of 9 months the Glmpgt/gt siblings had significantly reduced body weight. Reduced size of epididymal fat pads was accompanied by hepatosplenomegaly in Glmpgt/gt mice. Blood analysis revealed reduced levels of blood glucose, circulating triacylglycerol and non-esterified fatty acids in Glmpgt/gt mice. Increased flux of glucose, increased de novo lipogenesis and lipid accumulation were detected in Glmpgt/gt primary hepatocytes, as well as elevated triacylglycerol levels in Glmpgt/gt liver homogenates, compared to hepatocytes and liver from wild type mice. Gene expression analysis showed an increased expression of genes involved in fatty acid uptake and lipogenesis in Glmpgt/gt liver compared to wild type. Our findings are in agreement with the metabolic alterations observed in other mouse models lacking lysosomal proteins, and with alterations characteristic for advanced chronic liver injury. PMID:26047317

  3. Hepatic overexpression of microsomal triglyceride transfer protein (MTP) results in increased in vivo secretion of VLDL triglycerides and apolipoprotein B

    Microsoft Academic Search

    Uwe J. F. Tietge; Ahmed Bakillah; Cyrille Maugeais; Kazuhisa Tsukamoto; Mahmood Hussain; Daniel J. Rader

    The microsomal triglyceride transfer protein (MTP) is essential for the hepatic secretion of apolipopro- tein (apo) B-containing lipoproteins. Previous studies have indicated that inhibition of MTP results in decreased apoB plasma levels and decreased hepatic triglyceride secretion. However, the metabolic effects of overexpression of MTP have not been investigated. We constructed a recombinant adenovirus expressing MTP (AdhMTP) and used it

  4. Structure of the new mineral sarrabusite, Pb5CuCl4(SeO3)4, solved by manual electron-diffraction tomography.

    PubMed

    Gemmi, Mauro; Campostrini, Italo; Demartin, Francesco; Gorelik, Tatiana E; Gramaccioli, Carlo Maria

    2012-02-01

    The new mineral sarrabusite Pb(5)CuCl(4)(SeO(3))(4) has been discovered in the Sardinian mine of Baccu Locci, near Villaputzu. It occurs as small lemon-yellow spherical aggregates of tabular crystals (< 10 µm) of less than 100 µm in diameter. The crystal structure has been solved from and refined against electron diffraction of a microcrystal. Data sets have been measured by both a manual and an automated version of the new electron-diffraction tomography technique combined with the precession of the electron beam. The sarrabusite structure is monoclinic and consists of (010) layers of straight chains formed by alternating edge-sharing CuO(4)Cl(2) and PbO(8) polyhedra parallel to the c axis, which share corners laterally with two zigzag corner-sharing chains of PbO(6)Cl(2) and PbO(4)Cl(4) bicapped trigonal prisms. These blocks are linked together by SeO(3)(2-) flat-pyramidal groups. PMID:22267554

  5. Tethering of SUUR and HP1 proteins results in delayed replication of euchromatic regions in Drosophila melanogaster polytene chromosomes.

    PubMed

    Pokholkova, Galina V; Koryakov, Dmitry E; Pindyurin, Alexey V; Kozhevnikova, Elena N; Belyakin, Stepan N; Andreyenkov, Oleg V; Belyaeva, Elena S; Zhimulev, Igor F

    2015-06-01

    We analyze how artificial targeting of Suppressor of Under-Replication (SUUR) and HP1 proteins affects DNA replication in the "open," euchromatic regions. Normally these regions replicate early in the S phase and display no binding of either SUUR or HP1. These proteins were expressed as fusions with DNA-binding domain of GAL4 and recruited to multimerized UAS integrated in three euchromatic sites of the polytene X chromosome: 3B, 8D, and 18B. Using PCNA staining as a marker of ongoing replication, we showed that targeting of SUUR(GAL4DBD) and HP1(GAL4DBD) results in delayed replication of appropriate euchromatic regions. Specifically, replication at these regions starts early, much like in the absence of the fusion proteins; however, replication completion is significantly delayed. Notably, delayed replication was insufficient to induce underreplication. Recruitment of SUUR(GAL4DBD) and HP1(GAL4DBD) had distinct effects on expression of a mini-white reporter, found near UAS. Whereas SUUR(GAL4DBD) had no measurable influence on mini-white expression, HP1(GAL4DBD) targeting silenced mini-white, even in the absence of functional SU(VAR)3-9. Furthermore, recruitment of SUUR(GAL4DBD) and HP1(GAL4DBD) had distinct effects on the protein composition of target regions. HP1(GAL4DBD) but not SUUR(GAL4DBD) could displace an open chromatin marker, CHRIZ, from the tethering sites. PMID:25398563

  6. A comprehensive platform to investigate protein-metal ion interactions by affinity capillary electrophoresis.

    PubMed

    Alhazmi, Hassan A; Nachbar, Markus; Albishri, Hassan M; Abd El-Hady, Deia; Redweik, Sabine; El Deeb, Sami; Wätzig, Hermann

    2015-03-25

    In this work, the behavior of several metal ions with different globular proteins was investigated by affinity capillary electrophoresis. Screening was conducted by applying a proper rinsing protocol developed by our group. The use of 0.1M EDTA in the rinsing solution successfully desorbs metal ions from the capillary wall. The mobility ratio was used to evaluate the precision of the method. Excellent precision for repeated runs was achieved for different protein metal ion interactions (RSD% of 0.05-1.0%). Run times were less than 6 min for all of the investigated interactions. The method has been successfully applied for the interaction study of Li(+), Na(+), Mg(2+), Ca(2+), Ba(2+), Al(3+), Ga(3+), La(3+), Pd(2+), Ir(3+), Ru(3+), Rh(3+), Pt(2+), Pt(4+), Os(3+), Au(3+), Au(+), Ag(+), Cu(1+), Cu(2+), Fe(2+), Fe(3+), Co(2+), Ni(2+), Cr(3+), V(3+), MoO4(2-) and SeO3(2-) with bovine serum albumin, ovalbumin, ?-lactoglobulin and myoglobin. Different interaction values were obtained for most of the tested metal ions even for that in the same metal group. Results were discussed and compared in view of metal and semimetal group's interaction behavior with the tested proteins. The calculated normalized difference of mobility ratios for each protein-metal ion interaction and its sign (positive and negative) has been successfully used to detect the interaction and estimate further coordination of the bound metal ion, respectively. The comprehensive platform summarizes all the obtained interaction results, and is valuable for any future protein-metal ion investigation. PMID:25638307

  7. 3D Printing of Li-Ion Microbattery Architectures K. Sun, T.-S. Wei, B.Y. Ahn, J.Y. Seo, S.J. Dillon, and J.A. Lewis, "3D

    E-print Network

    Faraon, Andrei

    3D Printing of Li-Ion Microbattery Architectures K. Sun, T.-S. Wei, B.Y. Ahn, J.Y. Seo, S.J. Dillon, and J.A. Lewis, "3D Prin>ng of Interdigitated Li-Ion Microba) of 3D interdigitated microba8ery (3D-IMA). Scien(fic Achievement 3D prin

  8. A new approach to immobilize poly(vinyl alcohol) on poly(dimethylsiloxane) resulting in low protein adsorption

    NASA Astrophysics Data System (ADS)

    Carneiro, Leandro B.; Ferreira, Jacqueline; Santos, Marcos J. L.; Monteiro, Johny P.; Girotto, Emerson M.

    2011-10-01

    The hydrophobic characteristics of PDMS and non-specific protein adsorption are major drawbacks for its application in biosensing. Here we have combined surface oxidation by plasma and chemical binding of polyvinyl alcohol (PVA) to obtain long-term stability of hydrophilic PDMS surfaces. Mercaptopropyltrimethoxisilane and aminopropyltrimethoxisilane were used as adhesives between the plasma-oxidized PDMS surface and the PVA, immobilized at room temperature. This approach has allowed for fast, uniform, and very stable modification of the PDMS surface, which maintained a hydrophilic character for as long as 30 days. In addition, the modified hydrophilic surface presented minimized protein adsorption when compared to pristine PDMS. The results obtained in this work are important contributions to the growing field of integrated microfluidic biosensors.

  9. Neuronal overexpression of mutant amyloid precursor protein results in prominent deposition of cerebrovascular amyloid

    PubMed Central

    Calhoun, Michael E.; Burgermeister, Patrick; Phinney, Amie L.; Stalder, Martina; Tolnay, Markus; Wiederhold, Karl-Heinz; Abramowski, Dorothee; Sturchler-Pierrat, Christine; Sommer, Bernd; Staufenbiel, Matthias; Jucker, Mathias

    1999-01-01

    Transgenic mice that overexpress mutant human amyloid precursor protein (APP) exhibit one hallmark of Alzheimer’s disease pathology, namely the extracellular deposition of amyloid plaques. Here, we describe significant deposition of amyloid ? (A?) in the cerebral vasculature [cerebral amyloid angiopathy (CAA)] in aging APP23 mice that had striking similarities to that observed in human aging and Alzheimer’s disease. Amyloid deposition occurred preferentially in arterioles and capillaries and within individual vessels showed a wide heterogeneity (ranging from a thin ring of amyloid in the vessel wall to large plaque-like extrusions into the neuropil). CAA was associated with local neuron loss, synaptic abnormalities, microglial activation, and microhemorrhage. Although several factors may contribute to CAA in humans, the neuronal origin of transgenic APP, high levels of A? in cerebrospinal fluid, and regional localization of CAA in APP23 mice suggest transport and drainage pathways rather than local production or blood uptake of A? as a primary mechanism underlying cerebrovascular amyloid formation. APP23 mice on an App-null background developed a similar degree of both plaques and CAA, providing further evidence that a neuronal source of APP/A? is sufficient to induce cerebrovascular amyloid and associated neurodegeneration. PMID:10570203

  10. Synthesis, Crystal Structure, and IR and Mössbauer Spectroscopy of the Isotypic Series M3Fe 2(SeO 3) 6·2H 2O ( M=Mg, Co, Ni)

    NASA Astrophysics Data System (ADS)

    Giester, G.; Beran, A.; Redhammer, G. J.

    1997-06-01

    The representatives of the new isotypic seriesM3Fe2(SeO3)6·2H2O (M=Mg, Co, Ni) were prepared at 500 K from aqueous solutions, kept in Teflon-coated autoclaves. Their crystal structures were determined by direct methods from single-crystal X-ray diffraction data in the space groupPoverline1withZ=1. Mg3Fe2(SeO3)6·2H2O:a=6.494(2),b=7.959(3),c=8.810(4) Ĺ,?=85.88(2),?=79.12(2),?=76.09(2)°,V=433.9 Ĺ3. Co3Fe2(SeO3)6·2H2O:a=6.520(2),b=7.995(3),c=8.774(4) Ĺ,?=85.51(2),?=78.77(1),?=75.84(1)°,V=434.8 Ĺ3. Ni3Fe2(SeO3)6·2H2O:a=6.474(2),b=7.943(3),c=8.736(4) Ĺ,?=85.37(1),?=78.51(1),?=75.91(1)°,V=426.7 Ĺ3. The three differentMsites are commonly occupied byM2+and Fe3+atoms in varying ratios.MO6octahedra are linked to each other by edges to build four-membered groups. Via common corners with furtherMO6octahedra and trigonal pyramidal SeO3groups, complex sheets parallel to (001) are formed which are interconnected by hydrogen bonds only. IR and Mössbauer spectra are discussed.

  11. Heterogeneous N-terminal Acylation of Retinal Proteins Results from the Retina’s Unusual Lipid Metabolism†,§

    PubMed Central

    Bereta, Grzegorz; Palczewski, Krzysztof

    2011-01-01

    Protein N-myristoylation occurs by a covalent attachment of a C14:0 fatty acid to the N-terminal Gly residue. This reaction is catalyzed by a N-myristoyltransferase that uses myristoyl-coenzyme A as substrate. But proteins in the retina also undergo heterogeneous N-acylation with C14:2, C14:1 and C12:0 fatty acids. The basis and the role of this retina-specific phenomenon are poorly understood. We studied guanylate cyclase-activating protein 1 (GCAP1) as an example of retina-specific heterogeneously N-acylated protein. The types and the abundance of fatty acids bound to bovine retinal GCAP1 were: C14:2, 37.0%; C14:0, 32.4%; C14:1, 22.3%; and C12:0, 8.3% as quantified by liquid chromatography coupled mass spectrometry. We also devised a method for N-acylating proteins in vitro and used it to modify GCAP1 with acyl moieties of different lengths. Analysis of these GCAPs both confirmed that N-terminal acylation of GCAP1 is critical for its high activity and proper Ca2+-dependent response and revealed comparable functionality for GCAP1 with acyl moieties of various lengths. We also tested the hypothesis that retinal heterogeneous N-acylation results from retinal enrichment of unusual N-myristoyltransferase substrates. Thus, acyl-coenzyme A esters were purified from both bovine retina and brain and analyzed by liquid chromatography coupled mass spectrometry. Substantial differences in acyl-coenzyme A profiles between the retina and brain were detected. Importantly, the ratios of uncommon N-acylation substrates; C14:2- and C14:1-coenyzme A to C14:0-coenzyme A were higher in the retina than in the brain. Thus, our results suggest that heterogeneous N-acylation, responsible for expansion of retinal proteome, reflects the unique character of retinal lipid metabolism. Additionally, we propose a new hypothesis explaining the physiological relevance of elevated retinal ratios of C14:2- and C14:1-coenzyme A to C14:0-coenzyme A. PMID:21449552

  12. Neonatal presentation of familial glucocorticoid deficiency resulting from a novel splice mutation in the melanocortin 2 receptor accessory protein

    PubMed Central

    Jain, V; Metherell, L A; David, A; Sharma, R; Sharma, P K; Clark, A J L; Chan, L F

    2011-01-01

    Background Familial glucocorticoid deficiency (FGD) is a rare autosomal recessive disorder characterised by isolated glucocorticoid deficiency. Mutations in the ACTH receptor/melanocortin 2 receptor (MC2R), the MC2R accessory protein (MRAP) or the STAR protein (STAR) cause FGD types 1, 2 and 3, respectively, accounting for ?50% of all cases. Patient and methods We report a neonate of Indian origin, who was diagnosed with FGD in the first few days of life. He presented with hypoglycaemic seizures and was noted to have generalised intense hyperpigmentation and normal male genitalia. Biochemical investigations revealed hypocortisolaemia (cortisol 0.223??g/dl; NR 1–23??g/dl) and elevated plasma ACTH (170?pg/ml). Serum electrolytes, aldosterone and plasma renin activity were normal. Peak cortisol following a standard synacthen test was 0.018??g/dl. He responded to hydrocortisone treatment and continues on replacement. Patient DNA was analysed by direct sequencing. The effect of the novel mutation was assessed by an in vitro splicing assay using wild type and mutant heterologous minigenes. Results A novel homozygous mutation c.106+2_3dupTA was found in the MRAP gene. Both parents were heterozygous for the mutation. In an in vitro splicing assay, the mutation resulted in the skipping of exon 3. Conclusion We have identified a novel MRAP mutation where disruption of the intron 3 splice-site results in a prematurely terminated translation product. This protein (if produced) would lack the transmembrane domain that is essential for MC2R interaction. We predict that this would cause complete lack of ACTH response thus explaining the early presentation in this case. PMID:21951701

  13. RNA editing in wheat mitochondria results in the conservation of protein sequences

    Microsoft Academic Search

    José M. Gualberto; Lorenzo Lamattina; Géraldine Bonnard; Jacques-Henry Weil; Jean-Michel Grienenberger

    1989-01-01

    RNA editing is a process that results in the production of a messenger RNA with nucleotide sequences that differ from those of the template DNA1, and provides another mechanism for modulating gene expression. The phenomenon was initially described in the mitochondria of protozoa2, 3. Here we report that RNA editing is also required for the correct expression of plant mitochondria!

  14. Knockdown of Pnpla6 protein results in motor neuron defects in zebrafish.

    PubMed

    Song, Yang; Wang, Molin; Mao, Fei; Shao, Ming; Zhao, Baochang; Song, Zhen; Shao, Changshun; Gong, Yaoqin

    2013-03-01

    Mutations in patatin-like phospholipase domain containing 6 (PNPLA6), also known as neuropathy target esterase (NTE) or SPG39, cause hereditary spastic paraplegia (HSP). Although studies on animal models, including mice and Drosophila, have extended our understanding of PNPLA6, its roles in neural development and in HSP are not clearly understood. Here, we describe the generation of a vertebrate model of PNPLA6 insufficiency using morpholino oligonucleotide knockdown in zebrafish (Danio rerio). Pnpla6 knockdown resulted in developmental abnormalities and motor neuron defects, including axon truncation and branching. The phenotypes in pnpla6 knockdown morphants were rescued by the introduction of wild-type, but not mutant, human PNPLA6 mRNA. Our results also revealed the involvement of BMP signaling in pnpla6 knockdown phenotypes. Taken together, these results demonstrate an important role of PNPLA6 in motor neuron development and implicate overexpression of BMP signaling as a possible mechanism underlying the developmental defects in pnpla6 morphants. PMID:22996643

  15. Disruption of the basal body protein POC1B results in autosomal-recessive cone-rod dystrophy.

    PubMed

    Roosing, Susanne; Lamers, Ideke J C; de Vrieze, Erik; van den Born, L Ingeborgh; Lambertus, Stanley; Arts, Heleen H; Peters, Theo A; Hoyng, Carel B; Kremer, Hannie; Hetterschijt, Lisette; Letteboer, Stef J F; van Wijk, Erwin; Roepman, Ronald; den Hollander, Anneke I; Cremers, Frans P M

    2014-08-01

    Exome sequencing revealed a homozygous missense mutation (c.317C>G [p.Arg106Pro]) in POC1B, encoding POC1 centriolar protein B, in three siblings with autosomal-recessive cone dystrophy or cone-rod dystrophy and compound-heterozygous POC1B mutations (c.199_201del [p.Gln67del] and c.810+1G>T) in an unrelated person with cone-rod dystrophy. Upon overexpression of POC1B in human TERT-immortalized retinal pigment epithelium 1 cells, the encoded wild-type protein localized to the basal body of the primary cilium, whereas this localization was lost for p.Arg106Pro and p.Gln67del variant forms of POC1B. Morpholino-oligonucleotide-induced knockdown of poc1b translation in zebrafish resulted in a dose-dependent small-eye phenotype, impaired optokinetic responses, and decreased length of photoreceptor outer segments. These ocular phenotypes could partially be rescued by wild-type human POC1B mRNA, but not by c.199_201del and c.317C>G mutant human POC1B mRNAs. Yeast two-hybrid screening of a human retinal cDNA library revealed FAM161A as a binary interaction partner of POC1B. This was confirmed in coimmunoprecipitation and colocalization assays, which both showed loss of FAM161A interaction with p.Arg106Pro and p.Gln67del variant forms of POC1B. FAM161A was previously implicated in autosomal-recessive retinitis pigmentosa and shown to be located at the base of the photoreceptor connecting cilium, where it interacts with several other ciliopathy-associated proteins. Altogether, this study demonstrates that POC1B mutations result in a defect of the photoreceptor sensory cilium and thus affect cone and rod photoreceptors. PMID:25018096

  16. Identification of a novel MET mutation in high-grade glioma resulting in an auto-active intracellular protein.

    PubMed

    Navis, Anna C; van Lith, Sanne A M; van Duijnhoven, Sander M J; de Pooter, Maaike; Yetkin-Arik, Bahar; Wesseling, Pieter; Hendriks, Wiljan J A J; Venselaar, Hanka; Timmer, Marco; van Cleef, Patricia; van Bergen En Henegouwen, Paul; Best, Myron G; Wurdinger, Thomas D; Tops, Bastiaan B J; Leenders, William P J

    2015-07-01

    MET has gained interest as a therapeutic target for a number of malignancies because of its involvement in tumorigenesis, invasion and metastasis. At present, a number of inhibitors, both antibodies against MET or its ligand hepatocyte growth factor, and small molecule MET tyrosine kinase inhibitors are in clinical trials. We here describe a novel variant of MET that is expressed in 6 % of high-grade gliomas. Characterization of this mutation in a glioma cell line revealed that it consists of an intronic deletion, resulting in a splice event connecting an intact splice donor site in exon 6 with the next splice acceptor site being that of exon 9. The encoded protein lacks parts of the extracellular IPT domains 1 and 2, encoded by exons 7 and 8, resulting in a novel pseudo-IPT and is named MET(?7-8). MET(?7-8) is located predominantly in the cytosol and is constitutively active. The auto-activating nature of MET(?7-8), in combination with a lack of transmembrane localization, renders MET(?7-8) not targetable using antibodies, although the protein is efficiently deactivated by MET-specific tyrosine kinase inhibitors. Testing of MET-expressing tumors for the presence of this variant may be important for treatment decision making. PMID:25862637

  17. Vitiligo-inducing phenols activate the unfolded protein response in melanocytes resulting in upregulation of IL6 and IL8.

    PubMed

    Toosi, Siavash; Orlow, Seth J; Manga, Prashiela

    2012-11-01

    Vitiligo is characterized by depigmented skin patches caused by loss of epidermal melanocytes. Oxidative stress may have a role in vitiligo onset, while autoimmunity contributes to disease progression. In this study, we sought to identify mechanisms that link disease triggers and spreading of lesions. A hallmark of melanocytes at the periphery of vitiligo lesions is dilation of the endoplasmic reticulum (ER). We hypothesized that oxidative stress results in redox disruptions that extend to the ER, causing accumulation of misfolded peptides, which activates the unfolded protein response (UPR). We used 4-tertiary butyl phenol and monobenzyl ether of hydroquinone, known triggers of vitiligo. We show that expression of key UPR components, including the transcription factor X-box-binding protein 1 (XBP1), is increased following exposure of melanocytes to phenols. XBP1 activation increases production of immune mediators IL6 and IL8. Co-treatment with XBP1 inhibitors reduced IL6 and IL8 production induced by phenols, while overexpression of XBP1 alone increased their expression. Thus, melanocytes themselves produce cytokines associated with activation of an immune response following exposure to chemical triggers of vitiligo. These results expand our understanding of the mechanisms underlying melanocyte loss in vitiligo and pathways linking environmental stressors and autoimmunity. PMID:22696056

  18. GAS41 Amplification Results in Overexpression of a New Spindle Pole Protein

    PubMed Central

    Schmitt, Jana; Fischer, Ulrike; Heisel, Sabrina; Strickfaden, Hilmar; Backes, Christina; Ruggieri, Alessia; Keller, Andreas; Chang, Paul; Meese, Eckart

    2015-01-01

    Amplification is a hallmark of many human tumors but the role of most amplified genes in human tumor development is not yet understood. Previously, we identified a frequently amplified gene in glioma termed glioma-amplified sequence 41 (GAS41). Using the TCGA data portal and performing experiments on HeLa and TX3868, we analyzed the role of GAS41 amplification on GAS41 overexpression and the effect on the cell cycle. Here we show that GAS41 amplification is associated with overexpression in the majority of cases. Both induced and endogenous overexpression of GAS41 leads to an increase in multipolar spindles. We showed that GAS41 is specifically associated with pericentrosome material. As result of an increased GAS41 expression we found bipolar spindles with misaligned chromosomes. This number was even increased by a combined overexpression of GAS41 and a reduced expression of NuMA. We propose that GAS41 amplification may have an effect on the highly altered karyotype of glioblastoma via its role during spindle pole formation. PMID:22619067

  19. Prolonged high light treatment of plant cells results in changes of the amount, the localization and the electrophoretic behavior of several thylakoid membrane proteins.

    PubMed

    Schmid, V; Peter, S; Schäfer, C

    1995-06-01

    The effect of a 30 h high light treatment on the amount and the localization of thylakoid proteins was analysed in low light grown photoautotrophic cells of Marchantia polymorpha and Chenopodium rubrum. High light treatment resulted in a net loss of D1 protein which was accompanied by comparable losses of other proteins of the PS II core (reaction center with inner antenna). LHC II proteins were not reduced correspondingly, indicating that these complexes are less affected by prolonged high light. High light influenced the distribution of PS II components between the grana and the stroma region of the thylakoid membrane, probably by translocation of the respective PS II proteins. Additionally, modifications of several thylakoid proteins were detected in high light treated cells of C. rubrum. These effects are discussed in relation to photoinhibitory damage and repair processes. PMID:24307099

  20. Proton-driven assembly of the Rous Sarcoma virus capsid protein results in the formation of icosahedral particles.

    PubMed

    Hyun, Jae-Kyung; Radjainia, Mazdak; Kingston, Richard L; Mitra, Alok K

    2010-05-14

    In a mature and infectious retroviral particle, the capsid protein (CA) forms a shell surrounding the genomic RNA and the replicative machinery of the virus. The irregular nature of this capsid shell precludes direct atomic resolution structural analysis. CA hexamers and pentamers are the fundamental building blocks of the capsid, however the pentameric state, in particular, remains poorly characterized. We have developed an efficient in vitro protocol for studying the assembly of Rous sarcoma virus (RSV) CA that involves mild acidification and produces structures modeling the authentic viral capsid. These structures include regular spherical particles with T = 1 icosahedral symmetry, built from CA pentamers alone. These particles were subject to cryoelectron microscopy (cryo-EM) and image processing, and a pseudo-atomic model of the icosahedron was created by docking atomic structures of the constituent CA domains into the cryo-EM-derived three-dimensional density map. The N-terminal domain (NTD) of CA forms pentameric turrets, which decorate the surface of the icosahedron, while the C-terminal domain (CTD) of CA is positioned underneath, linking the pentamers. Biophysical analysis of the icosahedral particle preparation reveals that CA monomers and icosahedra are the only detectable species and that these exist in reversible equilibrium at pH 5. These same acidic conditions are known to promote formation of a RSV CA CTD dimer, present within the icosahedral particle, which facilitates capsid assembly. The results are consistent with a model in which RSV CA assembly is a nucleation-limited process driven by very weak protein-protein interactions. PMID:20228062

  1. Proton-driven Assembly of the Rous Sarcoma Virus Capsid Protein Results in the Formation of Icosahedral Particles*

    PubMed Central

    Hyun, Jae-Kyung; Radjainia, Mazdak; Kingston, Richard L.; Mitra, Alok K.

    2010-01-01

    In a mature and infectious retroviral particle, the capsid protein (CA) forms a shell surrounding the genomic RNA and the replicative machinery of the virus. The irregular nature of this capsid shell precludes direct atomic resolution structural analysis. CA hexamers and pentamers are the fundamental building blocks of the capsid, however the pentameric state, in particular, remains poorly characterized. We have developed an efficient in vitro protocol for studying the assembly of Rous sarcoma virus (RSV) CA that involves mild acidification and produces structures modeling the authentic viral capsid. These structures include regular spherical particles with T = 1 icosahedral symmetry, built from CA pentamers alone. These particles were subject to cryoelectron microscopy (cryo-EM) and image processing, and a pseudo-atomic model of the icosahedron was created by docking atomic structures of the constituent CA domains into the cryo-EM-derived three-dimensional density map. The N-terminal domain (NTD) of CA forms pentameric turrets, which decorate the surface of the icosahedron, while the C-terminal domain (CTD) of CA is positioned underneath, linking the pentamers. Biophysical analysis of the icosahedral particle preparation reveals that CA monomers and icosahedra are the only detectable species and that these exist in reversible equilibrium at pH 5. These same acidic conditions are known to promote formation of a RSV CA CTD dimer, present within the icosahedral particle, which facilitates capsid assembly. The results are consistent with a model in which RSV CA assembly is a nucleation-limited process driven by very weak protein-protein interactions. PMID:20228062

  2. Accumulation of a Brazil nut albumin in seeds of transgenic canola results in enhanced levels of seed protein methionine

    Microsoft Academic Search

    Susan B. Altenbach; Chiung-Chi Kuo; Lisa C. Staraci; Karen W. Pearson; Connie Wainwright; Anca Georgescu; Jeffrey Townsend

    1992-01-01

    We have increased the methionine content of the seed proteins of a commercial winter variety of canola by expressing a chimeric gene encoding a methionine-rich seed protein from Brazil nut in the seeds of transgenic plants. Transgenic canola seeds accumulate the heterologous methionine-rich protein at levels which range from 1.7% to 4.0% of the total seed protein and contain up

  3. Th(VO3)2(SeO3) and Ln(VO3)2(IO3) (Ln = Ce, Pr, Nd, Sm, and Eu): unusual cases of aliovalent substitution.

    PubMed

    Eaton, Teresa; Lin, Jian; Cross, Justin N; Stritzinger, Jared T; Albrecht-Schmitt, Thomas E

    2014-04-11

    Th(VO3)2(SeO3) and Ln(VO3)2(IO3) (Ln = Ce, Pr, Nd, Sm, and Eu) have been prepared and characterized. Surprisingly, these compounds are isotypic and rather extreme examples of aliovalent substitution (Th(IV)vs. Ln(III); Se(IV)O3(2-)vs. I(V)O3(-)) are possible in this structure type. PMID:24590373

  4. Perinatal Protein Malnutrition Affects Mitochondrial Function in Adult and Results in a Resistance to High Fat Diet-Induced Obesity

    PubMed Central

    Jousse, Céline; Muranishi, Yuki; Parry, Laurent; Montaurier, Christophe; Even, Patrick; Launay, Jean-Marie; Carraro, Valérie; Maurin, Anne-Catherine; Averous, Julien; Chaveroux, Cédric; Bruhat, Alain; Mallet, Jacques; Morio, Béatrice; Fafournoux, Pierre

    2014-01-01

    Epidemiological findings indicate that transient environmental influences during perinatal life, especially nutrition, may have deleterious heritable health effects lasting for the entire life. Indeed, the fetal organism develops specific adaptations that permanently change its physiology/metabolism and that persist even in the absence of the stimulus that initiated them. This process is termed “nutritional programming”. We previously demonstrated that mothers fed a Low-Protein-Diet (LPD) during gestation and lactation give birth to F1-LPD animals presenting metabolic consequences that are different from those observed when the nutritional stress is applied during gestation only. Compared to control mice, adult F1-LPD animals have a lower body weight and exhibit a higher food intake suggesting that maternal protein under-nutrition during gestation and lactation affects the energy metabolism of F1-LPD offspring. In this study, we investigated the origin of this apparent energy wasting process in F1-LPD and demonstrated that minimal energy expenditure is increased, due to both an increased mitochondrial function in skeletal muscle and an increased mitochondrial density in White Adipose Tissue. Importantly, F1-LPD mice are protected against high-fat-diet-induced obesity. Clearly, different paradigms of exposure to malnutrition may be associated with differences in energy expenditure, food intake, weight and different susceptibilities to various symptoms associated with metabolic syndrome. Taken together these results demonstrate that intra-uterine environment is a major contributor to the future of individuals and disturbance at a critical period of development may compromise their health. Consequently, understanding the molecular mechanisms may give access to useful knowledge regarding the onset of metabolic diseases. PMID:25118945

  5. The Identification of Two Germ-line Mutations in the Human Breast Cancer Resistance Protein Gene that Result in the Expression of a Low\\/Non-functional Protein

    Microsoft Academic Search

    Sho Yoshioka; Kazuhiro Katayama; Chikako Okawa; Sachiko Takahashi; Satomi Tsukahara; Junko Mitsuhashi; Yoshikazu Sugimoto

    2007-01-01

    Purpose  We examined the effects of the nine nonsynonymous germ-line mutations\\/SNPs in the breast cancer resistance protein (BCRP\\/ABCG2) gene on the expression and function of the protein.\\u000a \\u000a \\u000a \\u000a Materials and Methods  We generated cDNAs for each of these mutants (G151T, C458T, C496G, A616C, T623C, T742C, T1291C, A1768T, and G1858A BCRP) and compared the effects of their exogenous expression in PA317 cells with a

  6. The Relation between Dietary Protein, Calcium and Bone Health in Women: Results from the EPIC-Potsdam Cohort

    Microsoft Academic Search

    Cornelia Weikert; Dietmar Walter; Kurt Hoffmann; Anja Kroke; Manuela M. Bergmann; Heiner Boeing

    2005-01-01

    Background\\/Aims: The role of dietary protein in bone health is controversial. The objective of the present study was to examine the association between protein intake, dietary calcium, and bone structure measured by broadband ultrasound attenuation (BUA). Methods: Our analysis includes 8,178 female study participants of the European Prospective Investigation into Cancer and Nutrition (EPIC) Potsdam Study. Ultrasound bone measurements were

  7. Different Changes in Protein and Phosphoprotein Levels Result from Serum Starvation of High-Grade Glioma and Adenocarcinoma Cell Lines

    PubMed Central

    Levin, Victor A.; Panchabhai, Sonali C.; Shen, Li; Kornblau, Steven M.; Qiu, Yihua; Baggerly, Keith A.

    2012-01-01

    Tumor cells undergoing serum starvation in vitro partially mimic metabolically stressed cells trying to adjust to a changed environment in vivo by inducing signal transduction and gene expression so that the tumor continues to grow. Our hypothesis is that the changes in protein and phosphoprotein levels after serum starvation may reflect the adapted phenotype of the tumor, which could be targeted for therapy. We used reverse-phase protein microarrays to interrogate five high-grade glioma cell lines and seven adenocarcinoma cell lines for differences in the level of 81 proteins and 25 phosphoproteins. All cell lines were studied in the well-fed condition of growth with 10% FBS and the starved condition of 0.5% FBS. Protein expression levels were normalized to ?-actin and trichotomized as increased (+1, upper 75th quartile), decreased (?1, lowest 25th quartile), or unchanged (0, others) to focus on the patterns of the biggest (and hopefully most robust) changes in protein and phosphoprotein levels. We examined these trichotomized values to better understand Starved-Fed differences among the cell lines and thereby gain better/clearer insight into the effects of serum starvation on potential cellular responses. In general, the expression of proteins and phosphoproteins 24 h after FBS starvation increased more often in glioma lines than in adenocarcinoma lines, which appeared to have fewer increased protein scores and more decreased scores. Many of the proteins increased in gliomas were downstream targets of the PTEN-PI-3 kinase-AKT, EGFR-MAPK-Stat, and transcription activator-polyamine signaling pathways. In adenocarcinomas, the expression of proteins and phosphoproteins generally increased in apoptosis pathways, while there were minor fluctuations in the other pathways above. Contrawise, gliomas become resistant to apoptosis after 24 h of serum starvation and upregulate transcription activators and polyamines more so than adenocarciomas. PMID:19894763

  8. Hypomorphic CEP290/NPHP6 mutations result in anosmia caused by the selective loss of G proteins in cilia of olfactory sensory neurons.

    PubMed

    McEwen, Dyke P; Koenekoop, Robert K; Khanna, Hemant; Jenkins, Paul M; Lopez, Irma; Swaroop, Anand; Martens, Jeffrey R

    2007-10-01

    Cilia regulate diverse functions such as motility, fluid balance, and sensory perception. The cilia of olfactory sensory neurons (OSNs) compartmentalize the signaling proteins necessary for odor detection; however, little is known regarding the mechanisms of protein sorting/entry into olfactory cilia. Nephrocystins are a family of ciliary proteins likely involved in cargo sorting during transport from the basal body to the ciliary axoneme. In humans, loss-of-function of the cilia-centrosomal protein CEP290/NPHP6 is associated with Joubert and Meckel syndromes, whereas hypomorphic mutations result in Leber congenital amaurosis (LCA), a form of early-onset retinal dystrophy. Here, we report that CEP290-LCA patients exhibit severely abnormal olfactory function. In a mouse model with hypomorphic mutations in CEP290 [retinal dystrophy-16 mice (rd16)], electro-olfactogram recordings revealed an anosmic phenotype analogous to that of CEP290-LCA patients. Despite the loss of olfactory function, cilia of OSNs remained intact in the rd16 mice. As in wild type, CEP290 localized to dendritic knobs of rd16 OSNs, where it was in complex with ciliary transport proteins and the olfactory G proteins G(olf) and Ggamma(13). Interestingly, we observed defective ciliary localization of G(olf) and Ggamma(13) but not of G protein-coupled odorant receptors or other components of the odorant signaling pathway in the rd16 OSNs. Our data implicate distinct mechanisms for ciliary transport of olfactory signaling proteins, with CEP290 being a key mediator involved in G protein trafficking. The assessment of olfactory function can, therefore, serve as a useful diagnostic tool for genetic screening of certain syndromic ciliary diseases. PMID:17898177

  9. NCYM, a Cis-Antisense Gene of MYCN, Encodes a De Novo Evolved Protein That Inhibits GSK3? Resulting in the Stabilization of MYCN in Human Neuroblastomas

    PubMed Central

    Suenaga, Yusuke; Islam, S. M. Rafiqul; Alagu, Jennifer; Kaneko, Yoshiki; Kato, Mamoru; Tanaka, Yukichi; Kawana, Hidetada; Hossain, Shamim; Matsumoto, Daisuke; Yamamoto, Mami; Shoji, Wataru; Itami, Makiko; Shibata, Tatsuhiro; Nakamura, Yohko; Ohira, Miki; Haraguchi, Seiki; Takatori, Atsushi; Nakagawara, Akira

    2014-01-01

    The rearrangement of pre-existing genes has long been thought of as the major mode of new gene generation. Recently, de novo gene birth from non-genic DNA was found to be an alternative mechanism to generate novel protein-coding genes. However, its functional role in human disease remains largely unknown. Here we show that NCYM, a cis-antisense gene of the MYCN oncogene, initially thought to be a large non-coding RNA, encodes a de novo evolved protein regulating the pathogenesis of human cancers, particularly neuroblastoma. The NCYM gene is evolutionally conserved only in the taxonomic group containing humans and chimpanzees. In primary human neuroblastomas, NCYM is 100% co-amplified and co-expressed with MYCN, and NCYM mRNA expression is associated with poor clinical outcome. MYCN directly transactivates both NCYM and MYCN mRNA, whereas NCYM stabilizes MYCN protein by inhibiting the activity of GSK3?, a kinase that promotes MYCN degradation. In contrast to MYCN transgenic mice, neuroblastomas in MYCN/NCYM double transgenic mice were frequently accompanied by distant metastases, behavior reminiscent of human neuroblastomas with MYCN amplification. The NCYM protein also interacts with GSK3?, thereby stabilizing the MYCN protein in the tumors of the MYCN/NCYM double transgenic mice. Thus, these results suggest that GSK3? inhibition by NCYM stabilizes the MYCN protein both in vitro and in vivo. Furthermore, the survival of MYCN transgenic mice bearing neuroblastoma was improved by treatment with NVP-BEZ235, a dual PI3K/mTOR inhibitor shown to destabilize MYCN via GSK3? activation. In contrast, tumors caused in MYCN/NCYM double transgenic mice showed chemo-resistance to the drug. Collectively, our results show that NCYM is the first de novo evolved protein known to act as an oncopromoting factor in human cancer, and suggest that de novo evolved proteins may functionally characterize human disease. PMID:24391509

  10. Mechanisms of cross-talk between G-protein-coupled receptors resulting in enhanced release of intracellular Ca2+.

    PubMed Central

    Werry, Tim D; Wilkinson, Graeme F; Willars, Gary B

    2003-01-01

    Alteration in [Ca(2+)](i) (the intracellular concentration of Ca(2+)) is a key regulator of many cellular processes. To allow precise regulation of [Ca(2+)](i) and a diversity of signalling by this ion, cells possess many mechanisms by which they are able to control [Ca(2+)](i) both globally and at the subcellular level. Among these are many members of the superfamily of GPCRs (G-protein-coupled receptors), which are characterized by the presence of seven transmembrane domains. Typically, those receptors able to activate PLC (phospholipase C) enzymes cause release of Ca(2+) from intracellular stores and influence Ca(2+) entry across the plasma membrane. It has been well documented that Ca(2+) signalling by one type of GPCR can be influenced by stimulation of a different type of GPCR. Indeed, many studies have demonstrated heterologous desensitization between two different PLC-coupled GPCRs. This is not surprising, given our current understanding of negative-feedback regulation and the likely shared components of the signalling pathway. However, there are also many documented examples of interactions between GPCRs, often coupling preferentially to different signalling pathways, which result in a potentiation of Ca(2+) signalling. Such interactions have important implications for both the control of cell function and the interpretation of in vitro cell-based assays. However, there is currently no single mechanism that adequately accounts for all examples of this type of cross-talk. Indeed, many studies either have not addressed this issue or have been unable to determine the mechanism(s) involved. This review seeks to explore a range of possible mechanisms to convey their potential diversity and to provide a basis for further experimental investigation. PMID:12790797

  11. Deletion of the D domain of the human parainfluenza virus type 3 (HPIV3) PD protein results in decreased viral RNA synthesis and beta interferon (IFN-?) expression.

    PubMed

    Roth, Jason P; Li, Joseph K-K; Morrey, John D; Barnard, Dale L; Vollmer, Almut H

    2013-08-01

    The human parainfluenza virus type 3 (HPIV3) phosphoprotein (P) gene is unusual as it contains an editing site where nontemplated ribonucleotide residues can be inserted. This RNA editing can lead to the expression of the viral P, PD, putative W, and theoretical V protein from a single gene. Although the HPIV3 PD protein has been detected, its function and those of the W and V proteins are poorly understood. Therefore, we first used reverse genetics techniques to construct and rescue a recombinant (r)HPIV3 clone with a polyhistidine sequence at the 5' end of the P gene for tagged protein detection. Western blot analysis demonstrated the presence of the P, PD, and W proteins, but no V protein was detected. Then, we functionally studied the D domain of the PD protein by constructing two rHPIV3 knockout clones that are deficient in the expression of the D domain. Results from growth kinetic studies with infected MA-104 and A596 cells showed that viral replication of the two knockout viruses (rHPIV3-?ES and rHPIV3-?D) was comparable to that of the parental virus in both cell lines. However, viral mRNA transcription and genomic replication was significantly reduced. Furthermore, cytokine/chemokine profiles of A549 cells infected with either knockout virus were unchanged or showed lower levels compared to those from cells infected with the parental virus. These data suggest that the D domain of the PD protein may play a luxury role in HPIV3 RNA synthesis and may also be involved in disrupting the expression of beta interferon. PMID:23686695

  12. Ultrasonic Energy as a New Tool for Fast Isotopic 18O Labeling of Proteins for Mass Spectrometry-Based Techniques: Preliminary Results

    SciTech Connect

    Carreira, R.J.; Rial-Otero, R.; Lopez-Ferrer, Dani; Lodeiro, C.; Capelo, J.L.

    2008-07-15

    Preliminary results regarding fast isotopic labeling of proteins with 18O in conjunction with matrix assisted laser desorption ionization time of flight mass spectrometry technique are presented. Similar 16O/18O isotopic labeling ratios were found for the overnight procedure (12 h) and the new fast ultrasonic one (30 min) for the BSA, ovalbumin and ?-lactalbumin proteins. The procedure, however, failed to promote double 18O isotopic labeling for the proteins ovalbumin and ?-lactalbumin. Two different sonication frequencies, 35 kHz and 130 kHz, were studied at two different sonication times of 15 min and 30 min, being best results obtained with the procedure at 130 kHz of sonication frequency and 30 min of sonication time. For comparative purposes the overnight isotopic 18O labeling procedure was done. In addition, the new fast isotopic labeling procedure was also studied without ultrasonication, in a water bath at 60 şC.

  13. Serological diagnosis of hantavirus infections by an enzyme-linked immunosorbent assay based on detection of immunoglobulin G and M responses to recombinant nucleocapsid proteins of five viral serotypes.

    PubMed Central

    Elgh, F; Lundkvist, A; Alexeyev, O A; Stenlund, H; Avsic-Zupanc, T; Hjelle, B; Lee, H W; Smith, K J; Vainionpää, R; Wiger, D; Wadell, G; Juto, P

    1997-01-01

    Worldwide, hantaviruses cause more than 100,000 human infections annually. Rapid and accurate methods are important both in monitoring acute infections and for epidemiological studies. We and others have shown that the amino termini of hantavirus nucleocapsid proteins (Ns) are sensitive tools for the detection of specific antibodies in hantavirus disease. Accordingly, we expressed truncated Ns (amino acids 1 to 117) in Escherichia coli from the five hantaviruses known to be pathogenic to man; Hantaan (HTN), Seoul (SEO), Dobrava (DOB), Sin Nombre (SN), and Puumala (PUU) viruses. In order to obtain pure antigens for use in an enzyme-linked immunosorbent assay (ELISA), the recombinant proteins were purified by polyhistidine-metal chelate affinity chromatography. Polyclonal animal antisera and a panel of serum specimens from hantavirus-infected individuals from Scandinavia, Slovenia, Russia, Korea, China, and the United States were used to evaluate the usefulness of the method. With both human and animal sera, it was possible to designate the antibody response into two groups: those with HTN, SEO, and DOB virus reactivity on the one hand and those with SN and PUU virus reactivity on the other. In sera from Scandinavia, European Russia, and the United States, the antibody response was directed mainly to the PUU and SN virus group. The sera from Asia reacted almost exclusively with the HTN, SEO, and DOB types of viruses. This was true for both the immunoglobulin M (IgM) and IgG antibody responses, indicating that this type of discrimination can be done during the acute phase of hantavirus infections. Both the HTN, SEO, and DOB virus and the PUU and SN virus types of antibody response patterns were found in patients from the Balkan region (Solvenia). PMID:9114393

  14. Long-term clinical results of microsomal triglyceride transfer protein inhibitor use in a patient with homozygous familial hypercholesterolemia.

    PubMed

    Raper, Anna; Kolansky, Daniel M; Sachais, Bruce S; Meagher, Emma A; Baer, Amanda L; Cuchel, Marina

    2015-01-01

    We report the case of a 49-year-old woman with homozygous familial hypercholesterolemia and a complicated cardiovascular history, treated for 5 years with a microsomal triglyceride transfer protein inhibitor in addition to her other lipid-lowering therapy. PMID:25670368

  15. An encoded N-terminal extension results in low levels of heterologous protein production in Escherichia coli

    Microsoft Academic Search

    Samantha S Orchard; Heidi Goodrich-Blair

    2005-01-01

    BACKGROUND: The tdk gene (encoding deoxythymidine kinase) of the gamma-proteobacterium Xenorhabdus nematophila has two potential translation start sites. The promoter-distal start site was predicted to be functional based on amino acid sequence alignment with closely related Tdk proteins. However, to experimentally determine if either of the two possible start codons allows production of a functional Tdk, we expressed the \\

  16. Mutations in the major DNA-binding protein gene of herpes simplex virus type 1 result in increased levels of viral gene expression.

    PubMed Central

    Godowski, P J; Knipe, D M

    1983-01-01

    We have examined the effect of temperature-sensitive mutations in the herpes simplex virus 1 DNA-binding protein gene on viral gene expression. We have found that at the nonpermissive temperature, the synthesis of certain immediate early, early, and late viral polypeptides was greater in cells infected with the temperature-sensitive mutants than in cells infected with the wild-type virus. This effect was independent of the requirement for this viral protein for viral DNA replication. The altered rate of synthesis of viral proteins was due to a thermolabile gene product. Cells infected with these mutants at the permissive temperature and then shifted to the nonpermissive temperature exhibited enhanced levels of viral gene expression. The addition of actinomycin D at the time of the temperature shift prevented the alteration in viral protein synthesis. Therefore, continuing transcription is required for this change in gene expression. Northern blot analysis of cytoplasmic RNA showed that the steady-state level of specific viral transcripts expressed from parental virus genomes was greater in cells infected by these mutants at the nonpermissive temperature. These results indicate that the major DNA-binding protein of herpes simplex virus type 1 acts as a negative regulator of viral gene expression by affecting the abundance of cytoplasmic viral mRNAs. Images PMID:6312079

  17. Protease Inhibitor Coinfusion with Amyloid b-Protein Results in Enhanced Deposition and Toxicity in Rat Brain

    Microsoft Academic Search

    Sally A. Frautschy; David L. Horn; Jason J. Sigel; Marni E. Harris-White; John J. Mendoza; Fusheng Yang; T. C. Saido; Gregory M. Cole

    1998-01-01

    Amyloid b-protein, Ab, is normally produced in brain and is cleared by unknown mechanisms. In Alzheimer's disease (AD), Ab accumulates in plaque-like deposits and is implicated ge- netically in neurodegeneration. Here we investigate mecha- nisms for Ab degradation and Ab toxicity in vivo, focusing on the effects of Ab40, which is the peptide that accumulates in apolipoprotein E4-associated AD. Chronic

  18. Reduced amount of the glucose-regulated protein GRP94 in skeletal myoblasts results in loss of fusion competence

    Microsoft Academic Search

    LUISA GORZA; MAURIZIO VITADELLO

    We previously showed that skeletal myocytes of the adult rabbit do not accumulate the endoplasmic reticulum glucose-regulated protein GRP94, neither constitutively nor inducibly, at vari- ance with skeletal myocytes during perinatal devel- opment (5). Here we show that C2C12 cells up- regulate GRP94 during differentiation and, similarly to primary cultures of murine skeletal myocytes, specifically display GRP94 immunoreactivity on the

  19. Nd 5O 4Cl[AsO 3] 2 and Gd 5O 4Br 3[SeO 3] 2: Two lanthanoid oxide halides with complex "lone-pair" oxoanions

    NASA Astrophysics Data System (ADS)

    Kang, Dong-Hee; Wontcheu, Joseph; Schleid, Thomas

    2009-02-01

    Both compounds, neodymium oxide chloride oxoarsenate(III) Nd 5O 4Cl[AsO 3] 2 and gadolinium oxide bromide oxoselenate(IV) Gd 5O 4Br 3[SeO 3] 2, were prepared by solid-state reactions from mixtures of the corresponding binary oxides and halides, and their crystal structures have been determined by X-ray diffraction of single crystals. They crystallize monoclinically ( a = 1241.62(9) pm, b = 565.78(4) pm, c = 902.03(7) pm, ? = 116.454(3)° for Nd 5O 4Cl[AsO 3] 2 and a = 1243.70(9) pm, b = 549.91(4) pm, c = 1005.28(8) pm, ? = 91.869(3)° for Gd 5O 4Br 3[SeO 3] 2) in space group C2/ m with two formula units per unit cell. The non-isotypic crystal structures contain three crystallographically different M 3+ cations (M = Nd and Gd). The coordination sphere of (M1) 3+ consists of eight oxygen atoms (CN = 8) exclusively, whereas (M2) 3+ carries six oxygen atoms and one X - anion (X = Cl and Br, CN = 7) in each case. For (M3) 3+, however, CN = 8 is realized by six oxygen atoms and two Cl - anions in Nd 5O 4Cl[AsO 3] 2, but five oxygen atoms and three Br - anions in Gd 5O 4Br 3[SeO 3] 2. The isolated pyramidal [AsO 3] 3-/[SeO 3] 2- anions ( d(As 3+-O 2-) = 175-179; d(Se 4+-O 2-) = 165-174 pm) originate from three oxygen atoms (O2 and two O3), which surround the As 3+/Se 4+ cations together with the stereochemically active non-bonding electron pair ( lone pair) ? 1-tetrahedrally (?(O-As-O) = 95-102; ?(O-Se-O) = 95-96°). Both crystal structures are built up of corrugated two-dimensional lanthanoid-oxygen layers {[}?2 consisting of edge- and corner-shared [OM 4] 10+ tetrahedra ( d(O 2--Nd 3+) = 228-242; d(O 2--Gd 3+) = 226-235 pm). The single Cl - anion in the neodymium and the two crystallographically independent Br - anions in the gadolinium compound reside in between these sheets, where the lone-pair electrons at the As 3+/Se 4+ cations point into the center of channels, which are formed by lanthanoid-oxygen layers and halide chains.

  20. Single amino acid exchange in bacteriophage HK620 tailspike protein results in thousand-fold increase of its oligosaccharide affinity.

    PubMed

    Broeker, Nina K; Gohlke, Ulrich; Müller, Jürgen J; Uetrecht, Charlotte; Heinemann, Udo; Seckler, Robert; Barbirz, Stefanie

    2013-01-01

    Bacteriophage HK620 recognizes and cleaves the O-antigen polysaccharide of Escherichia coli serogroup O18A1 with its tailspike protein (TSP). HK620TSP binds hexasaccharide fragments with low affinity, but single amino acid exchanges generated a set of high-affinity mutants with submicromolar dissociation constants. Isothermal titration calorimetry showed that only small amounts of heat were released upon complex formation via a large number of direct and solvent-mediated hydrogen bonds between carbohydrate and protein. At room temperature, association was both enthalpy- and entropy-driven emphasizing major solvent rearrangements upon complex formation. Crystal structure analysis showed identical protein and sugar conformers in the TSP complexes regardless of their hexasaccharide affinity. Only in one case, a TSP mutant bound a different hexasaccharide conformer. The extended sugar binding site could be dissected in two regions: first, a hydrophobic pocket at the reducing end with minor affinity contributions. Access to this site could be blocked by a single aspartate to asparagine exchange without major loss in hexasaccharide affinity. Second, a region where the specific exchange of glutamate for glutamine created a site for an additional water molecule. Side-chain rearrangements upon sugar binding led to desolvation and additional hydrogen bonding which define this region of the binding site as the high-affinity scaffold. PMID:22923442

  1. The copper-transporting capacity of ATP7A mutants associated with Menkes disease is ameliorated by COMMD1 as a result of improved protein expression.

    PubMed

    Vonk, Willianne I M; de Bie, Prim; Wichers, Catharina G K; van den Berghe, Peter V E; van der Plaats, Rozemarijn; Berger, Ruud; Wijmenga, Cisca; Klomp, Leo W J; van de Sluis, Bart

    2012-01-01

    Menkes disease (MD) is an X-linked recessive disorder characterized by copper deficiency resulting in a diminished function of copper-dependent enzymes. Most MD patients die in early childhood, although mild forms of MD have also been described. A diversity of mutations in the gene encoding of the Golgi-resident copper-transporting P(1B)-type ATPase ATP7A underlies MD. To elucidate the molecular consequences of the ATP7A mutations, various mutations in ATP7A associated with distinct phenotypes of MD (L873R, C1000R, N1304S, and A1362D) were analyzed in detail. All mutants studied displayed changes in protein expression and intracellular localization parallel to a dramatic decline in their copper-transporting capacity compared to ATP7A the wild-type. We restored these observed defects in ATP7A mutant proteins by culturing the cells at 30°C, which improves the quality of protein folding, similar to that which as has recently has been demonstrated for misfolded ATP7B, a copper transporter homologous to ATP7A. Further, the effect of the canine copper toxicosis protein COMMD1 on ATP7A function was examined as COMMD1 has been shown to regulate the proteolysis of ATP7B proteins. Interestingly, in addition to adjusted growth temperature, binding of COMMD1 partially restored the expression, subcellular localization, and copper-exporting activities of the ATP7A mutants. However, no effect of pharmacological chaperones was observed. Together, the presented data might provide a new direction for developing therapies to improve the residual exporting activity of unstable ATP7A mutant proteins, and suggests a potential role for COMMD1 in this process. PMID:21667063

  2. Cyclophilin A Binds to the Viral RNA and Replication Proteins, Resulting in Inhibition of Tombusviral Replicase Assembly

    PubMed Central

    Kovalev, Nikolay

    2013-01-01

    Replication of plus-stranded RNA viruses is greatly affected by numerous host-encoded proteins that act as restriction factors. Cyclophilins, which are a large family of cellular prolyl isomerases, have been found to inhibit Tomato bushy stunt tombusvirus (TBSV) replication in a Saccharomyces cerevisiae model based on genome-wide screens and global proteomics approaches. In this report, we further characterize single-domain cyclophilins, including the mammalian cyclophilin A and plant Roc1 and Roc2, which are orthologs of the yeast Cpr1p cyclophilin, a known inhibitor of TBSV replication in yeast. We found that recombinant CypA, Roc1, and Roc2 strongly inhibited TBSV replication in a cell-free replication assay. Additional in vitro studies revealed that CypA, Roc1, and Roc2 cyclophilins bound to the viral replication proteins, and CypA and Roc1 also bound to the viral RNA. These interactions led to inhibition of viral RNA recruitment, the assembly of the viral replicase complex, and viral RNA synthesis. A catalytically inactive mutant of CypA was also able to inhibit TBSV replication in vitro due to binding to the replication proteins and the viral RNA. Overexpression of CypA and its mutant in yeast or plant leaves led to inhibition of tombusvirus replication, confirming that CypA is a restriction factor for TBSV. Overall, the current work has revealed a regulatory role for the cytosolic single-domain Cpr1-like cyclophilins in RNA virus replication. PMID:24089553

  3. Exposure to vehicle emissions results in altered blood brain barrier permeability and expression of matrix metalloproteinases and tight junction proteins in mice

    PubMed Central

    2013-01-01

    Background Traffic-generated air pollution-exposure is associated with adverse effects in the central nervous system (CNS) in both human exposures and animal models, including neuroinflammation and neurodegeneration. While alterations in the blood brain barrier (BBB) have been implicated as a potential mechanism of air pollution-induced CNS pathologies, pathways involved have not been elucidated. Objectives To determine whether inhalation exposure to mixed vehicle exhaust (MVE) mediates alterations in BBB permeability, activation of matrix metalloproteinases (MMP) -2 and ?9, and altered tight junction (TJ) protein expression. Methods Apolipoprotein (Apo) E?/? and C57Bl6 mice were exposed to either MVE (100 ?g/m3 PM) or filtered air (FA) for 6 hr/day for 30 days and resulting BBB permeability, expression of ROS, TJ proteins, markers of neuroinflammation, and MMP activity were assessed. Serum from study mice was applied to an in vitro BBB co-culture model and resulting alterations in transport and permeability were quantified. Results MVE-exposed Apo E?/? mice showed increased BBB permeability, elevated ROS and increased MMP-2 and ?9 activity, compared to FA controls. Additionally, cerebral vessels from MVE-exposed mice expressed decreased levels of TJ proteins, occludin and claudin-5, and increased levels of inducible nitric oxide synthase (iNOS) and interleukin (IL)-1? in the parenchyma. Serum from MVE-exposed animals also resulted in increased in vitro BBB permeability and altered P-glycoprotein transport activity. Conclusions These data indicate that inhalation exposure to traffic-generated air pollutants promotes increased MMP activity and degradation of TJ proteins in the cerebral vasculature, resulting in altered BBB permeability and expression of neuroinflammatory markers. PMID:24344990

  4. Placing the RPL32 Promoter Upstream of a Second Promoter Results in a Strongly Increased Number of Stably Transfected Mammalian Cell Lines That Display High Protein Expression Levels

    PubMed Central

    Hoeksema, F.; Hamer, K.; Siep, M.; Verhees, J. A.; Otte, A. P.

    2011-01-01

    The use of high stringency selection systems commonly results in a strongly diminished number of stably transfected mammalian cell lines. Here we placed twelve different promoters upstream of an adjacent primary promoter and tested whether this might result in an increased number of colonies; this is in the context of a stringent selection system. We found that only the promoter of the human ribosomal protein, RPL32, induced a high number of colonies in CHO-DG44 cells. This phenomenon was observed when the RPL32 promoter was combined with the CMV, SV40, EF1-?, and the ?-actin promoters. In addition, these colonies displayed high protein expression levels. The RPL32 promoter had to be functionally intact, since the deletion of a small region upstream of the transcription start site demolished its positive action. We conclude that adding the RPL32 promoter to an expression cassette in cis may be a powerful tool to augment gene expression levels. PMID:21350661

  5. Placing the RPL32 Promoter Upstream of a Second Promoter Results in a Strongly Increased Number of Stably Transfected Mammalian Cell Lines That Display High Protein Expression Levels.

    PubMed

    Hoeksema, F; Hamer, K; Siep, M; Verhees, J A; Otte, A P

    2011-01-01

    The use of high stringency selection systems commonly results in a strongly diminished number of stably transfected mammalian cell lines. Here we placed twelve different promoters upstream of an adjacent primary promoter and tested whether this might result in an increased number of colonies; this is in the context of a stringent selection system. We found that only the promoter of the human ribosomal protein, RPL32, induced a high number of colonies in CHO-DG44 cells. This phenomenon was observed when the RPL32 promoter was combined with the CMV, SV40, EF1-?, and the ?-actin promoters. In addition, these colonies displayed high protein expression levels. The RPL32 promoter had to be functionally intact, since the deletion of a small region upstream of the transcription start site demolished its positive action. We conclude that adding the RPL32 promoter to an expression cassette in cis may be a powerful tool to augment gene expression levels. PMID:21350661

  6. 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

  7. Accumulation of PrLeg, a Perilla legumin protein in potato tuber results in enhanced level of sulphur-containing amino acids.

    PubMed

    Goo, Young-Min; Kim, Tae-Won; Lee, Min-Kyung; Lee, Shin-Woo

    2013-09-01

    Potato is the fourth staple food in the world, following rice, wheat, and maize, whereas tubers contain high quality of starch, relatively high amounts of vitamin C and many other important substances. It also contains relatively good quality of protein (about 3 to 6% of the dried weight) and patatin, and 11S globulin is a major storage protein with high level of lysine. However, tuber protein contains relatively low amounts of sulphur-containing amino acids, which may result in low nutritional value. Recently, we cloned a gene encoding PrLeg polypeptide, a seed storage protein from Perilla, which contains relatively higher levels of sulphur-containing amino acids. We transformed PrLeg cDNA into a potato plant to over-express under the direction of the tuber-specific promoter, patatin. Most of the transgenic lines identified through PCR and RT-PCR analyses were able to accumulate high amount of prLeg transcript in their tuber tissue, while very little or no transcript that were detected in their leaf tissues. The level of methionine content was elevated up to three-fold compared to non-transgenic parental line, without any significant changes in other amino acids, suggesting that further research is required to get a deeper insight into their nutritional value. PMID:24161240

  8. Accumulation of radium in ferruginous protein bodies formed in lung tissue: association of resulting radiation hotspots with malignant mesothelioma and other malignancies

    PubMed Central

    Nakamura, Eizo; Makishima, Akio; Hagino, Kyoko; Okabe, Kazunori

    2009-01-01

    While exposure to fibers and particles has been proposed to be associated with several different lung malignancies including mesothelioma, the mechanism for the carcinogenesis is not fully understood. Along with mineralogical observation, we have analyzed forty-four major and trace elements in extracted asbestos bodies (fibers and proteins attached to them) with coexisting fiber-free ferruginous protein bodies from extirpative lungs of individuals with malignant mesothelioma. These observations together with patients’ characteristics suggest that inhaled iron-rich asbestos fibers and dust particles, and excess iron deposited by continuous cigarette smoking would induce ferruginous protein body formation resulting in ferritin aggregates in lung tissue. Chemical analysis of ferruginous protein bodies extracted from lung tissues reveals anomalously high concentrations of radioactive radium, reaching millions of times higher concentration than that of seawater. Continuous and prolonged internal exposure to hotspot ionizing radiation from radium and its daughter nuclides could cause strong and frequent DNA damage in lung tissue, initiate different types of tumour cells, including malignant mesothelioma cells, and may cause cancers. PMID:19644223

  9. In vivo detection and characterization of protein adducts resulting from bioactivation of haloethene cysteine S-conjugates by 19F NMR: chlorotrifluoroethene and tetrafluoroethene.

    PubMed

    Harris, J W; Dekant, W; Anders, M W

    1992-01-01

    Several haloalkenes are selective nephrotoxins. The bioactivation of nephrotoxic haloalkenes involves hepatic glutathione S-conjugate formation, peptidase-catalyzed metabolism of the glutathione S-conjugates to the corresponding cysteine S-conjugates, uptake of cysteine S-conjugates by the kidneys, and renal cysteine conjugate beta-lyase-catalyzed beta-elimination of a thiol. The haloalkyl and haloalkenyl thiols thus released are unstable and yield reactive intermediates whose interactions with cellular constituents are though to contribute to the observed toxicity of S-conjugates. Tetrafluoroethene and chlorotrifluoroethene are metabolized to the cysteine S-conjugates S-(1,1,2,2-tetrafluoroethyl)-L-cysteine (TFEC) and S-(2-chloro-1,1,2-trifluoroethyl)-L-cysteine (CTFC), respectively. Administration of TFEC (1.0 mmol/kg) or CTFC (1.0 mmol/kg) to rats resulted in acylation of renal proteins, as demonstrated with 19F nuclear magnetic resonance spectroscopy. Single, broad resonances near 41 or 56 ppm were found in spectra of renal proteins from TFEC- or CTFC-treated rats, respectively, and these resonances were not lost on dialysis. Renal protein incubated with 2-chloro-1,1,2-trifluoroethyl-2-nitrophenyl disulfide, a proreactive intermediate that yields 2-chloro-1,1,2-trifluoroethanethiol, showed the same 19F NMR spectrum as was found with CTFC-treated rats. In vitro incubation of various N alpha-blocked amino acids with this proreactive intermediate indicated that only lysine is stably adducted, whereas histidine is transiently acylated. In each case, proteolysis of modified protein converted a single broad NMR resonance to a doublet with little change in chemical shift and with clearly resolved, characteristic H-F couplings. The single, stable amino acid adduct formed with renal proteins of rats given CTFC or TFEC was N epsilon-(chlorofluorothioacetyl)lysine and N epsilon-(difluorothioacetyl)lysine, respectively. PMID:1581534

  10. Partial loss of the DNA repair scaffolding protein, Xrcc1, results in increased brain damage and reduced recovery from ischemic stroke in mice.

    PubMed

    Ghosh, Somnath; Canugovi, Chandrika; Yoon, Jeong Seon; Wilson, David M; Croteau, Deborah L; Mattson, Mark P; Bohr, Vilhelm A

    2015-07-01

    Oxidative DNA damage is mainly repaired by base excision repair (BER). Previously, our laboratory showed that mice lacking the BER glycosylases 8-oxoguanine glycosylase 1 (Ogg1) or nei endonuclease VIII-like 1 (Neil1) recover more poorly from focal ischemic stroke than wild-type mice. Here, a mouse model was used to investigate whether loss of 1 of the 2 alleles of X-ray repair cross-complementing protein 1 (Xrcc1), which encodes a nonenzymatic scaffold protein required for BER, alters recovery from stroke. Ischemia and reperfusion caused higher brain damage and lower functional recovery in Xrcc1(+/-) mice than in wild-type mice. Additionally, a greater percentage of Xrcc1(+/-) mice died as a result of the stroke. Brain samples from human individuals who died of stroke and individuals who died of non-neurological causes were assayed for various steps of BER. Significant losses of thymine glycol incision, abasic endonuclease incision, and single nucleotide incorporation activities were identified, as well as lower expression of XRCC1 and NEIL1 proteins in stroke brains compared with controls. Together, these results suggest that impaired BER is a risk factor in ischemic brain injury and contributes to its recovery. PMID:25971543

  11. Comparison of AOD retrieved from Brewer spectrophotometer in SeoComparison of AOD retrieved from Brewer spectrophotometer in Seoulul JaJa--Ho Koo,Ho Koo, YunYun--Mi Kim,Mi Kim, Jhoon Kim, Hi Ku ChoJhoon Kim, Hi Ku Cho

    E-print Network

    Wang, Yuhang

    Comparison of AOD retrieved from Brewer spectrophotometer in SeoComparison of AOD retrieved from Brewer spectrophotometer in Seoulul JaJa--Ho Koo,Ho Koo, YunYun--Mi Kim,Mi Kim, Jhoon Kim, Hi Ku Cho-GAW Brewer Users Group Meeting : 28 Oct ­ 3 Nov 2007 Summary 1. AOD retrieved from Brewer spectrophotometer

  12. Neuroprotection resulting from insufficiency of RANBP2 is associated with the modulation of protein and lipid homeostasis of functionally diverse but linked pathways in response to oxidative stress

    PubMed Central

    Cho, Kyoung-in; Yi, Haiqing; Tserentsoodol, Nomingerel; Searle, Kelly; Ferreira, Paulo A.

    2010-01-01

    SUMMARY Oxidative stress is a deleterious stressor associated with a plethora of disease and aging manifestations, including neurodegenerative disorders, yet very few factors and mechanisms promoting the neuroprotection of photoreceptor and other neurons against oxidative stress are known. Insufficiency of RAN-binding protein-2 (RANBP2), a large, mosaic protein with pleiotropic functions, suppresses apoptosis of photoreceptor neurons upon aging and light-elicited oxidative stress, and promotes age-dependent tumorigenesis by mechanisms that are not well understood. Here we show that, by downregulating selective partners of RANBP2, such as RAN GTPase, UBC9 and ErbB-2 (HER2; Neu), and blunting the upregulation of a set of orphan nuclear receptors and the light-dependent accumulation of ubiquitylated substrates, light-elicited oxidative stress and Ranbp2 haploinsufficiency have a selective effect on protein homeostasis in the retina. Among the nuclear orphan receptors affected by insufficiency of RANBP2, we identified an isoform of COUP-TFI (Nr2f1) as the only receptor stably co-associating in vivo with RANBP2 and distinct isoforms of UBC9. Strikingly, most changes in proteostasis caused by insufficiency of RANBP2 in the retina are not observed in the supporting tissue, the retinal pigment epithelium (RPE). Instead, insufficiency of RANBP2 in the RPE prominently suppresses the light-dependent accumulation of lipophilic deposits, and it has divergent effects on the accumulation of free cholesterol and free fatty acids despite the genotype-independent increase of light-elicited oxidative stress in this tissue. Thus, the data indicate that insufficiency of RANBP2 results in the cell-type-dependent downregulation of protein and lipid homeostasis, acting on functionally interconnected pathways in response to oxidative stress. These results provide a rationale for the neuroprotection from light damage of photosensory neurons by RANBP2 insufficiency and for the identification of novel therapeutic targets and approaches promoting neuroprotection. PMID:20682751

  13. Positive Selection Results in Frequent Reversible Amino Acid Replacements in the G Protein Gene of Human Respiratory Syncytial Virus

    PubMed Central

    Botosso, Viviane F.; de A. Zanotto, Paolo M.; Ueda, Mirthes; Arruda, Eurico; Gilio, Alfredo E.; Vieira, Sandra E.; Stewien, Klaus E.; Peret, Teresa C. T.; Jamal, Leda F.; Pardini, Maria I. de M. C.; Pinho, Joăo R. R.; Massad, Eduardo; Sant'Anna, Osvaldo A.; Holmes, Eddie C.; Durigon, Edison L.

    2009-01-01

    Human respiratory syncytial virus (HRSV) is the major cause of lower respiratory tract infections in children under 5 years of age and the elderly, causing annual disease outbreaks during the fall and winter. Multiple lineages of the HRSVA and HRSVB serotypes co-circulate within a single outbreak and display a strongly temporal pattern of genetic variation, with a replacement of dominant genotypes occurring during consecutive years. In the present study we utilized phylogenetic methods to detect and map sites subject to adaptive evolution in the G protein of HRSVA and HRSVB. A total of 29 and 23 amino acid sites were found to be putatively positively selected in HRSVA and HRSVB, respectively. Several of these sites defined genotypes and lineages within genotypes in both groups, and correlated well with epitopes previously described in group A. Remarkably, 18 of these positively selected tended to revert in time to a previous codon state, producing a “flip-flop” phylogenetic pattern. Such frequent evolutionary reversals in HRSV are indicative of a combination of frequent positive selection, reflecting the changing immune status of the human population, and a limited repertoire of functionally viable amino acids at specific amino acid sites. PMID:19119418

  14. Copper deficiency results in AMP-activated protein kinase activation and acetylCoA carboxylase phosphorylation in rat cerebellum

    PubMed Central

    Gybina, Anna A.; Prohaska, Joseph R.

    2008-01-01

    Copper (Cu) deficiency impairs cerebellar development including biosynthetic processes like myelination and synaptogenesis. The activity of cerebellar mitochondrial cuproenzyme cytochrome c oxidase is markedly lower in Cu deficient rat pups and is accompanied by higher lactate levels indicating mitochondrial inhibition. Cu deficiency impaired energy metabolism is thought to contribute to developmental delays, but specific mechanisms linking these phenomena have remained unexplored. AMP activated protein kinase (AMPK) is a cellular energy sensor that is activated during mitochondrial inhibition and shuts down biosynthetic processes to help conserve cellular ATP levels. Activated AMPK phosphorylates and inhibits acetyl CoA carboxylase (ACC), the first enzyme in fatty acid biosynthesis. We hypothesize AMPK is activated and ACC inhibited in Cu deficient cerebella. Perinatal copper deficiency was studied in young rats in rapidly frozen cerebella. Compared to copper adequate (Cu+) pups, copper deficient (Cu?) pups were hypothermic, had lower brain copper levels and markedly higher cerebellar lactate. Concentration of phosphorylated AMPK (pAMPK), indicating AMPK activation, was robustly higher in Cu? cerebella of rat pups at two ages and in two separate experiments. Compared to Cu+ cerebella, pACC content was significantly higher in all Cu? samples. Mechanisms leading to AMPK activation remain elusive. Higher AMP/ATP ratios and increased reactive nitrogen species (RNS) can lead to AMPK activation. ATP and AMP concentrations were unaltered and nitric oxide metabolites and 3-nitrotyrosine peptide levels remained unchanged in Cu? cerebella. AMPK activation may explain how ATP levels can be maintained even with a severe mitochondrial loss of CCO function. PMID:18339363

  15. IL-4 upregulates Ig? and Ig? protein, resulting in augmented IgM maturation and BCR-triggered B cell activation1

    PubMed Central

    Guo, Benchang; Rothstein, Thomas L.

    2015-01-01

    Interleukin 4 (IL-4) is critical for optimal B cell activation and germinal center B cell expansion in T-dependent immune responses; however, the underlying mechanism remains elusive. In the present study, we found that primary B cells express little Ig? and Ig? protein despite substantial levels of messenger RNA. IL-4 markedly up-regulates Ig? and Ig? protein expression that requires STAT6. Elevated Ig? and Ig? protein form heterodimers that associate with IgM and significantly promote IgM maturation and surface IgM expression, resulting in amplified BCR-initiated signaling that is Lyn-dependent. In vivo, we found that pre-germinal center B cells express upregulated Ig?, Ig?, and surface IgM expression, in conjunction with elevated BCR-triggered pERK ex vivo, that are dependent on IL-4 and reversed by in vivo administration of neutralizing anti-IL-4 antibody. Thus, this study elucidates a novel mechanism for crosstalk between the IL-4 and B cell receptors that programs enhancement of subsequent BCR signaling. PMID:23776171

  16. 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

  17. Feeding soy protein isolate prevents impairment of bone acquisition by western diets as a result of insulin signaling in bone

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Excessive consumption of high fat/high cholesterol “Western” diets during postnatal life results in increased energy intake, development of obesity and systemic insulin resistance. However, how this diet impairs bone development and remodeling is not well understood, and no effective dietary interve...

  18. Search Engine Optimization: SEO Book

    NSDL National Science Digital Library

    Wall, Aaron

    The strange and wondrous ways in which search engines gather their indexes is made a little clearer in this tutorial. Written by a web designer disappointed with how difficult his pages were to find with the standard search engines, this page gives insight into how Infoseek, Lycos, Alta Vista, Excite, Web Crawler, and Open Text catalog web pages. The search strategy of each engine is described, along with tips for how web designers can increase their site's chances of being among the hits returned when users enter relevant search criteria. Although indexing algorithms are constantly being updated, this site presents common-sense guidelines that web designers interested in reaching a wider audience will find useful.

  19. Adenomatous polyposis coli tumor suppressor protein has signaling activity in Xenopus laevis embryos resulting in the induction of an ectopic dorsoanterior axis.

    PubMed

    Vleminckx, K; Wong, E; Guger, K; Rubinfeld, B; Polakis, P; Gumbiner, B M

    1997-01-27

    Mutations in the adenomatous polyposis coli (APC) tumor suppressor gene are linked to both familial and sporadic human colon cancer. So far, a clear biological function for the APC gene product has not been determined. We assayed the activity of APC in the early Xenopus embryo, which has been established as a good model for the analysis of the signaling activity of the APC-associated protein beta-catenin. When expressed in the future ventral side of a four-cell embryo, full-length APC induced a secondary dorsoanterior axis and the induction of the homeobox gene Siamois. This is similar to the phenotype previously observed for ectopic beta-catenin expression. In fact, axis induction by APC required the availability of cytosolic beta-catenin. These results indicate that APC has signaling activity in the early Xenopus embryo. Signaling activity resides in the central domain of the protein, a part of the molecule that is missing in most of the truncating APC mutations in colon cancer. Signaling by APC in Xenopus embryos is not accompanied by detectable changes in expression levels of beta-catenin, indicating that it has direct positive signaling activity in addition to its role in beta-catenin turnover. From these results we propose a model in which APC acts as part of the Wnt/beta-catenin signaling pathway, either upstream of, or in conjunction with, beta-catenin. PMID:9015311

  20. Vascular remodeling alters adhesion protein and cytoskeleton reactions to inflammatory stimuli resulting in enhanced permeability increases in rat venules

    PubMed Central

    Yuan, Dong

    2012-01-01

    Vascular remodeling has been implicated in many inflammation-involved diseases. This study aims to investigate the microvascular remodeling-associated alterations in cell-cell adhesion and cytoskeleton reactions to inflammatory stimuli and their impact on microvessel permeability. Experiments were conducted in individually perfused rat mesenteric venules. Microvessel permeability was determined by measuring hydraulic conductivity (Lp), and endothelial intracellular calcium concentration, [Ca2+]i, was measured in fura-2-perfused vessels. Alterations in VE-cadherin and F-actin arrangement were examined by confocal imaging. Vascular wall cellular composition and structural changes were evaluated by electron microscopy. Vessels exposed to platelet activating factor (PAF) on day 1 were reevaluated 3 days later in rats that had undergone survival surgery. Initial PAF exposure and surgical disturbance increased microvascular wall thickness along with perivascular cell proliferation and altered F-actin arrangement. Although basal permeability was not changed, upon reexposure to PAF, peak endothelial [Ca2+]i was augmented and the peak Lp was 9.3 ± 1.7 times higher than that of day 1. In contrast to patterns of PAF-induced stress fiber formation and VE-cadherin redistribution observed in day 1 vessels, the day 4 vessels at the potentiated Lp peak exhibited wide separations of VE-cadherin between endothelial cells and striking stress fibers throughout the vascular walls. Confocal images and ultrastructural micrographs also revealed that the largely separated VE-cadherin and endothelial gaps were completely covered by F-actin bundles in extended pericyte processes at the PAF-induced Lp peak. These results indicate that inflammation-induced vascular remodeling increased endothelial susceptibility to inflammatory stimuli with augmented Ca2+ response resulting in upregulated contractility and potentiated permeability increase. Weakened adhesions between the endothelial cells and contractile mechanisms are both involved in increasing permeability in the intact microvessels and are aggravated during remodeling. The perivascular cells play important roles in stabilizing the microvessel wall, while lessening an otherwise much greater magnitude of leakage during cytoskeletal contraction. PMID:22837164

  1. Gene fusions of signal sequences with a modified beta-glucuronidase gene results in retention of the beta-glucuronidase protein in the secretory pathway/plasma membrane.

    PubMed Central

    Yan, X; Gonzales, R A; Wagner, G J

    1997-01-01

    Signal sequences and endoplasmic reticulum (ER) retention signals are known to play central roles in targeting and translocation in the secretory pathway, but molecular aspects about their involvement are poorly understood. We tested the effectiveness of deduced signal sequences from various genes (hydroxyproline-rich glycoprotein [HRGP] from Phaseolus vulgaris; Serpin from Manduca sexta) to direct a modified beta-glucuronidase (GUS) protein into the secretory pathway in transgenic tobacco (Nicotiana tabacum L.). The reporter protein was not secreted to the cell wall/extracellular space as monitored using extracellular fluid analysis (low- or high-ionic-strength conditions) but occurred in membranes with a density of 1.16 to 1.20 g/mL. Membrane-bound GUS equilibrated with the plasma membrane (PM) and the ER on linear sucrose gradients with or without ethylenediaminetetraacetic acid, suggesting that GUS associates with the ER and the PM. Confocal microscopy of fixed cultured cells prepared from GUS control and HRGP signal peptide (SP)-GUS-expressing plants suggested only cytosolic localization in GUS-expressing plants but substantial peripheral localization in HRGP SP-GUS plants, which is consistent with GUS being associated with the PM. Aqueous two-phase partitioning of microsomal membranes from HRGP SP-GUS and Serpin SP-GUS transgenic leaves also indicated that GUS activity was enriched in the ER and the PM. These observations, together with hydrophobic moment plot analysis, suggest that properties of the SP-GUS protein result in its retention in the secretory pathway and PM. PMID:9390428

  2. Protein-protein interaction databases.

    PubMed

    Szklarczyk, Damian; Jensen, Lars Juhl

    2015-01-01

    Years of meticulous curation of scientific literature and increasingly reliable computational predictions have resulted in creation of vast databases of protein interaction data. Over the years, these repositories have become a basic framework in which experiments are analyzed and new directions of research are explored. Here we present an overview of the most widely used protein-protein interaction databases and the methods they employ to gather, combine, and predict interactions. We also point out the trade-off between comprehensiveness and accuracy and the main pitfall scientists have to be aware before adopting protein interaction databases in any single-gene or genome-wide analysis. PMID:25859942

  3. Mutations in the TIR1 auxin receptor that increase affinity for auxin/indole-3-acetic acid proteins result in auxin hypersensitivity.

    PubMed

    Yu, Hong; Moss, Britney L; Jang, Seunghee S; Prigge, Michael; Klavins, Eric; Nemhauser, Jennifer L; Estelle, Mark

    2013-05-01

    The phytohormone auxin regulates virtually every aspect of plant development. The hormone directly mediates the interaction between the two members of the auxin coreceptor complex, a TRANSPORT INHIBITOR RESPONSE (TIR1)/AUXIN SIGNALING F-BOX protein and an AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) transcriptional repressor. To learn more about the interaction between these proteins, a mutant screen was performed using the yeast (Saccharomyces cerevisiae) two-hybrid system in Arabidopsis (Arabidopsis thaliana). Two tir1 mutations were identified that increased interaction with Aux/IAAs. The D170E and M473L mutations increase affinity between TIR1 and the degron motif of Aux/IAAs and enhance the activity of the SCF(TIR1) complex. This resulted in faster degradation of Aux/IAAs and increased transcription of auxin-responsive genes in the plant. Plants carrying the pTIR1:tir1 D170E/M473L-Myc transgene exhibit diverse developmental defects during plant growth and display an auxin-hypersensitive phenotype. This work demonstrates that changes in the leucine-rich repeat domain of the TIR1 auxin coreceptor can alter the properties of SCF(TIR1). PMID:23539280

  4. Intracellular accumulation of a 46 kDa species of mouse prion protein as a result of loss of glycosylation in cultured mammalian cells.

    PubMed

    Biswas, Subhabrata; Langeveld, Jan P M; Tipper, Donald; Lu, Shan

    2006-10-13

    Prion diseases are fatal neurodegenerative disorders characterized by the accumulation of an abnormal isoform (PrPSc) of the normal cellular prion protein (PrPC) in the brain. Reportedly, abnormal N-linked glycosylation patterns in PrPC are associated with disease susceptibility; thus, we compared the glycosylation status of normal and several mutant forms of the murine prion protein (MuPrP) in cultured mammalian cells. Substitution of the N-terminal signal sequence of normal MuPrP with a heterologous signal peptide did not alter glycosylation. When expressed without the C-terminal glycophosphatidylinositol anchor signal, the majority of MuPrP remained intracellular and unglycosylated, and a 46 kDa species (p46) of the unglycosylated PrPC was detected on reducing gels. p46 was also observed when wild-type MuPrP was expressed in the presence of tunicamycin or enzymatically deglycosylated in vitro. A rabbit polyclonal anti-serum raised against dimeric MuPrP cross-reacted with p46 and localized the signal within the Golgi apparatus. We propose that the 46 kDa signal is a dimeric form of MuPrP and in the light of recent studies, it can be argued that a relatively stable, non-glycosylated, cytoplasmic PrPC dimer, produced as a result of compromised glycosylation is an intermediate in initiating conversion of PrPC to PrPSc in sporadic transmissible spongiform encephalopathies. PMID:16935263

  5. Disruption of the protein kinase N gene of Drosophila melanogaster Results in the Recessive delorean Allele (pkndln) With a Negative Impact on Wing Morphogenesis

    PubMed Central

    Sass, Georgette L.; Ostrow, Bruce D.

    2014-01-01

    We describe the delorean mutation of the Drosophila melanogaster protein kinase N gene (pkndln) with defects in wing morphology. Flies homozygous for the recessive pkndln allele have a composite wing phenotype that exhibits changes in relative position and shape of the wing blade as well as loss of specific vein and bristle structures. The pkndln allele is the result of a P-element insertion in the first intron of the pkn locus, and the delorean wing phenotype is contingent upon the interaction of insertion-bearing alleles in trans. The presence of the insertion results in production of a novel transcript that initiates from within the 3? end of the P-element. The delorean-specific transcript is predicted to produce a wild-type PKN protein. The delorean phenotype is not the result of a reduction in pkn expression, as it could not be recreated using a variety of wing-specific drivers of pkn-RNAi expression. Rather, it is the presence of the delorean-specific transcript that correlates with the mutant phenotype. We consider the delorean wing phenotype to be due to a pairing-dependent, recessive mutation that behaves as a dosage-sensitive, gain of function. Our analysis of genetic interactions with basket and nemo reflects an involvement of pkn and Jun-terminal kinase signaling in common processes during wing differentiation and places PKN as a potential effector of Rho1’s involvement in the Jun-terminal kinase pathway. The delorean phenotype, with its associated defects in wing morphology, provides evidence of a role for PKN in adult morphogenetic processes. PMID:24531729

  6. Heat shock protein 70 and nitric oxide concentrations in non-tumorous and neoplastic canine mammary tissues: preliminary results - Short communication.

    PubMed

    Szczubia?, Marek; Urban-Chmiel, Renata; ?opuszy?ski, Wojciech

    2015-06-01

    The concentrations of heat shock protein 70 (Hsp70) and nitric oxide ions (NO), measured as nitrite, were determined in canine mammary tumours and nontumorous mammary gland tissues. The concentrations of Hsp70 and NO were significantly higher in both benign and malignant tumours than in non-tumorous mammary tissues. Hsp70 concentration decreased with the increase in the grade of histological malignancy. A strong positive correlation was found between the concentrations of Hsp70 and NO in the benign tumours as well as in grade I and grade II malignant tumours. The results indicate that the process of neoplastic transformation in the canine mammary gland is related to a significant increase in Hsp70 and NO concentration in tumour tissues, and an interdependence between Hsp70 and nitrite ion production can be observed. PMID:26051259

  7. Evidence that the glutamine-stimulated loss of nitrate reductase protein from the yeast Candida nitratophila is not the result of inducer exclusion.

    PubMed Central

    Hipkin, C R; Kau, D A; Cannons, A C

    1993-01-01

    Synthesis of nitrate reductase protein and increases in nitrate reductase activity occurred in cultures of the yeast Candida nitratophila when they were incubated in medium containing ammonium nitrate. Similar treatment with glutamine plus nitrate resulted in little increase in nitrate reductase activity, in cultures grown previously with reduced nitrogen compounds, and decreases in enzyme activity, in cultures adapted to nitrate. Labelling studies conducted in vivo revealed a rapid cessation of de novo nitrate reductase synthesis when glutamine was supplied to nitrate-adapted cultures in the presence of nitrate. Intracellular glutamine concentrations increased rapidly under these conditions and these cultures exhibited high glutamine: glutamate ratios. As nitrate was taken up in the presence of glutamine in these experiments, it is concluded that the glutamine-stimulated inhibition of nitrate reductase synthesis is a consequence of repression and rapid turnover of nitrate reductase mRNA and not inducer (nitrate) exclusion. Images Figure 3 Figure 5 PMID:8240265

  8. Lipid Raft- and Protein Kinase C-mediated Synergism between Glucocorticoid- and Gonadotropin-releasing Hormone Signaling Results in Decreased Cell Proliferation*

    PubMed Central

    Wehmeyer, Lancelot; Du Toit, Andrea; Lang, Dirk M.; Hapgood, Janet P.

    2014-01-01

    Cross-talk between the glucocorticoid receptor (GR) and other receptors is emerging as a mechanism for fine-tuning cellular responses. We have previously shown that gonadotropin-releasing hormone (GnRH) ligand-independently activates the GR and synergistically modulates glucocorticoid-induced transcription of an endogenous gene in L?T2 pituitary gonadotrope precursor cells. Here, we investigated GR and GnRH receptor (GnRHR) cross-talk that involves co-localization with lipid rafts in L?T2 cells. We report that the GnRHR and a small population of the GR co-localize with the lipid raft protein flotillin-1 (Flot-1) at the plasma membrane and that the GR is present in a complex with Flot-1, independent of the presence of ligands. We found that the SGK-1 gene is up-regulated by Dex and GnRH alone, whereas a combination of both ligands resulted in a synergistic increase in SGK-1 mRNA levels. Using siRNA-mediated knockdown and antagonist strategies, we show that the gene-specific synergistic transcriptional response requires the GR, GnRHR, and Flot-1 as well as the protein kinase C pathway. Interestingly, although several GR cofactors are differentially recruited to the SGK-1 promoter in the presence of Dex and GnRH, GR levels remain unchanged compared with Dex treatment alone, suggesting that lipid raft association of the GR has a role in enhancing its transcriptional output in the nucleus. Finally, we show that Dex plus GnRH synergistically inhibit cell proliferation in a manner dependent on SGK-1 and Flot-1. Collectively the results support a mechanism whereby GR and GnRHR cross-talk within Flot-1-containing lipid rafts modulates cell proliferation via PKC activation and SGK-1 up-regulation. PMID:24558046

  9. Active immunization against riboflavin carrier protein results in peri-implantation embryonic loss leading to pregnancy termination in rats: use of alternate adjuvants.

    PubMed

    Rao, J; Seshagiri, P B; Shetty, G; Ramesh, G; Adiga, P R

    2000-09-01

    To investigate the mechanism of pregnancy termination following immuno-neutralization of riboflavin carrier protein (RCP) and to use acceptable adjuvants, we actively immunized female rats with reduced and carboxymethylated RCP (RCM-RCP) using various adjuvants (during primary immunization) such as sodium phthalylated lipopolysaccharide (SPLPS), purified S. typhi outer membrane proteins (porins) and a combination of them. Rats (5-14 per group) were immunized with alugel adsorbed RCM-RCP (100 microg/dose) either alone or with SPLPS or porins or SPLPS+porins. Control animals received RCM-RCP emulsified with Fruend's completelincomplete adjuvants (FCA/FIA). All animals received five boosters at intervals of 21 days. The lowest (4 X 10(-3)) and the highest (> 70 X 10(-3)) anti-RCM-RCP antibody titers were observed in alugel adsorbed-RCM-RCP group and control groups, respectively. Immunized animals showed reduced fertility following 3rd, 4th and 5th boosters. Reduction in fertility was 30-60% in alugel adsorbed RCM-RCP group, 90-100% in FCA-RCM-RCP group and 80-90% in SPLPS+porins group. Fertility reduction was not strictly correlatable with the serum antibody titers. RCP-specific IgG could be localized in the uterine endometrial glands and luminal epithelial cells in the immunized animals. Animals in the FCA/FIA group showed abnormal implantation/resorption sites and their histological sections showed degenerated embryos. But, day 5 preimplantation embryos were normal. These results show that (a) SPLPS+porins can be used as adjuvants in place of FCA/FIA for active immunization against RCM-RCP and (b) early termination of pregnancy in the immunized animals is due largely to the failure of normal embryo implantation. PMID:12561942

  10. Downregulation of Cellular c-Jun N-Terminal Protein Kinase and NF-?B Activation by Berberine May Result in Inhibition of Herpes Simplex Virus Replication

    PubMed Central

    Song, Siwei; Qiu, Min; Chu, Ying; Chen, Deyan; Wang, Xiaohui; Su, Airong

    2014-01-01

    Berberine is a quaternary ammonium salt from the protoberberine group of isoquinoline alkaloids. Some reports show that berberine exhibits anti-inflammatory, antitumor, and antiviral properties by modulating multiple cellular signaling pathways, including p53, nuclear factor ?B (NF-?B), and mitogen-activated protein kinase. In the present study, we investigated the antiviral effect of berberine against herpes simplex virus (HSV) infection. Current antiherpes medicines such as acyclovir can lessen the recurring activation when used early at infection but are unable to prevent or cure infections where treatment has selected for resistant mutants. In searching for new antiviral agents against herpesvirus infection, we found that berberine reduced viral RNA transcription, protein synthesis, and virus titers in a dose-dependent manner. To elucidate the mechanism of its antiviral activity, the effect of berberine on the individual steps of viral replication cycle of HSV was investigated via time-of-drug addition assay. We found that berberine acted at the early stage of HSV replication cycle, between viral attachment/entry and genomic DNA replication, probably at the immediate-early gene expression stage. We further demonstrated that berberine significantly reduced HSV-induced NF-?B activation, as well as I?B-? degradation and p65 nuclear translocation. Moreover, we found that berberine also depressed HSV-induced c-Jun N-terminal kinase (JNK) phosphorylation but had little effect on p38 phosphorylation. Our results suggest that the berberine inhibition of HSV infection may be mediated through modulating cellular JNK and NF-?B pathways. PMID:24913175

  11. Moderate Hypoxia Followed by Reoxygenation Results in Blood-Brain Barrier Breakdown via Oxidative Stress-Dependent Tight-Junction Protein Disruption

    PubMed Central

    Zehendner, Christoph M.; Librizzi, Laura; Hedrich, Jana; Bauer, Nina M.; Angamo, Eskedar A.; de Curtis, Marco; Luhmann, Heiko J.

    2013-01-01

    Re-canalization of cerebral vessels in ischemic stroke is pivotal to rescue dysfunctional brain areas that are exposed to moderate hypoxia within the penumbra from irreversible cell death. Goal of the present study was to evaluate the effect of moderate hypoxia followed by reoxygenation (MHR) on the evolution of reactive oxygen species (ROS) and blood-brain barrier (BBB) integrity in brain endothelial cells (BEC). BBB integrity was assessed in BEC in vitro and in microvessels of the guinea pig whole brain in situ preparation. Probes were exposed to MHR (2 hours 67-70 mmHg O2, 3 hours reoxygenation, BEC) or towards occlusion of the arteria cerebri media (MCAO) with or without subsequent reperfusion in the whole brain preparation. In vitro BBB integrity was evaluated using trans-endothelial electrical resistance (TEER) and transwell permeability assays. ROS in BEC were evaluated using 2?,7?-dichlorodihydrofluorescein diacetate (DCF), MitoSox and immunostaining for nitrotyrosine. Tight-junction protein (TJ) integrity in BEC, stainings for nitrotyrosine and FITC-albumin extravasation in the guinea pig brain preparation were assessed by confocal microscopy. Diphenyleneiodonium (DPI) was used to investigate NADPH oxidase dependent ROS evolution and its effect on BBB parameters in BEC. MHR impaired TJ proteins zonula occludens 1 (ZO-1) and claudin 5 (Cl5), decreased TEER, and significantly increased cytosolic ROS in BEC. These events were blocked by the NADPH oxidase inhibitor DPI. MCAO with or without subsequent reoxygenation resulted in extravasation of FITC-albumin and ROS generation in the penumbra region of the guinea pig brain preparation and confirmed BBB damage. BEC integrity may be impaired through ROS in MHR on the level of TJ and the BBB is also functionally impaired in moderate hypoxic conditions followed by reperfusion in a complex guinea pig brain preparation. These findings suggest that the BBB is susceptible towards MHR and that ROS play a key role in this process. PMID:24324834

  12. Increased adenosine levels in mice expressing mutant glial fibrillary acidic protein in astrocytes result in failure of induction of LTP reversal (depotentiation) in hippocampal CA1 neurons.

    PubMed

    Fujii, Satoshi; Tanaka, Kenji F; Ikenaka, Kazuhiro; Yamazaki, Yoshihiko

    2014-08-26

    Astrocytes regulate the activity of neighboring neurons by releasing chemical transmitters, including ATP. Adenosine levels in the cerebrospinal fluid of mice that express a mutant human glial fibrillary acidic protein in astrocytes are slightly elevated compared to those in wild type mice and this might result from the observed increased release by mutant astrocytes of ATP, which can be used to produce adenosine. Using hippocampal slices from these mutant mice, we examined whether the increased endogenous adenosine levels in the hippocampus modulate the reversal of long-term potentiation (LTP), i.e. depotentiation (DP), in CA1 neurons. In hippocampal slices from wild type mice, a stable LTP was induced by tetanic stimulation consisting of 100 pulses at 100 Hz, and this was reversed by a train of low frequency stimulation (LFS) of 500 pulses at 1 Hz applied 30 min later. This induction of DP was inhibited by application of either 100 nM adenosine or 0.5 nM N(6)-cyclopentyladenosine, an adenosine A1 receptor agonist, during LFS, indicating that the increase in extracellular adenosine levels attenuated DP induction by acting on adenosine A1 receptors. In contrast, although a stable LTP was also induced in hippocampal slices from mutant mice, induction of DP was inhibited, but DP could be induced by application, during LFS, of 50 nM 8-cyclopentyltheophylline, an adenosine A1 receptor antagonist. These results suggest that a small increase in extracellular adenosine levels resulting from increased ATP release by astrocytes results in attenuation of DP in hippocampal CA1 neurons in the mutant mice. PMID:25017946

  13. Staphylococcus aureus protein A binding to osteoblast tumour necrosis factor receptor 1 results in activation of nuclear factor kappa B and release of interleukin-6 in bone infection.

    PubMed

    Claro, Tânia; Widaa, Amro; McDonnell, Cormac; Foster, Timothy J; O'Brien, Fergal J; Kerrigan, Steven W

    2013-01-01

    Staphylococcus aureus is the major pathogen among the staphylococci and the most common cause of bone infections. These infections are mainly characterized by bone destruction and inflammation, and are often debilitating and very difficult to treat. Previously we demonstrated that S. aureus protein A (SpA) can bind to osteoblasts, which results in inhibition of osteoblast proliferation and mineralization, apoptosis, and activation of osteoclasts. In this study we used small interfering RNA (siRNA) to demonstrate that osteoblast tumour necrosis factor receptor-1 (TNFR-1) is responsible for the recognition of and binding to SpA. TNFR-1 binding to SpA results in the activation of nuclear factor kappa B (NF?B). In turn, NF?B translocates to the nucleus of the osteoblast, which leads to release of interleukin 6 (IL-6). Silencing TNFR-1 in osteoblasts or disruption of the spa gene in S. aureus prevented both NF?B activation and IL-6 release. As well as playing a key role in proinflammatory reactions, IL-6 is also an important osteotropic factor. Release of IL-6 from osteoblasts results in the activation of the bone-resorbing cells, the osteoclasts. Consistent with our results described above, both silencing TNFR-1 in osteoblasts and disruption of spa in S. aureus prevented osteoclast activation. These studies are the first to demonstrate the importance of the TNFR-1-SpA interaction in bone infection, and may help explain the mechanism through which osteoclasts become overactivated, leading to bone destruction. Anti-inflammatory drug therapy could be used either alone or in conjunction with antibiotics to treat osteomyelitis or for prophylaxis in high-risk patients. PMID:23154968

  14. Rats fed soy protein isolate (SPI) have impaired hepatic CYP1A1 induction by polycyclic aromatic hydrocarbons as a result of interference with aryl hydrocarbon receptor signaling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Consumption of soy diet has been found to reduce cancer incidence in animals and is associated with reduced cancer risk in humans. Previously, we have demonstrated that female Sprague-Dawley rats fed purified AIN-93G diets with soy protein isolate (SPI) as the sole protein source had reduced CYP1A1 ...

  15. Absence of myotonic dystrophy protein kinase (DMPK) mRNA as a result of a triplet repeat expansion in myotonic dystrophy

    SciTech Connect

    Carango, P.; Noble, J.E.; Funanage, V.L.; Marks, H.G. (Alfred I. duPont Institute, Wilmington, DE (United States))

    1993-11-01

    Myotonic dystrophy is an autosomally dominant inherited disease in which system-wide abnormalities are caused by a triplet repeat expansion within the 3[prime] untranslated region of the myotonic dystrophy protein kinase (DMPK) gene. To determine the effect an expanded repeat region has on DMPK expression, the authors have separated the chromosome 19 homologues from a 36-year-old woman with myotonic dystrophy into different cell lines by way of somatic cell hybridization. Hybrid DM9101 contains the normal DMPK allele (13 repeats), whereas hybrid DM1115 harbors the mutant allele ([approximately]133 repeats). Reverse transcription/polymerase chain reaction (RT/PCR) amplification of coding sequences from the DMPK gene has shown both reduced levels of primary DMPK transcripts and impaired processing of these transcripts in hybrid cell line DM1115. These findings suggest that the presence of a large number of repeats in the 3[prime] untranslated region of the DMPK gene reduces both the synthesis and the processing of DMPK mRNA, resulting in undetectable levels of processed DMPK mRNA from the mutant allele. 41 refs., 6 figs., 1 tab.

  16. Oral immune regulation using colitis extracted proteins for treatment of Crohn’s disease: Results of a phase I clinical trial

    PubMed Central

    Israeli, Eran; Goldin, Eran; Shibolet, Oren; Klein, Athalia; Hemed, Nilla; Engelhardt, Dean; Rabbani, Elazar; Ilan, Yaron

    2005-01-01

    AIM: To evaluate safety and possible efficacy of induction of oral immune regulation using colitis extracted proteins (CEP) in Crohn’s disease (CD) subjects. METHODS: Ten CDs were treated orally with autologous CEP thrice weekly for 16 wk. Subjects were monitored for CDAI and IBDQ. Immune modulatory effect was assessed by T-lymphocyte FACS analysis, CEP-specific IFN? ELISPOT assay and cytokine levels. RESULTS: Induction of oral immune regulation significantly ameliorated disease activity. All (10/10) subjects had clinical response (CDAI ? 70) and 7/10 achieved clinical remission (CDAI ? 150). Significant increase in mean IBDQ score was noted (134±9 vs 164±12). No treatment-related adverse events were noted. High levels of CEP-specific IFN? spot forming colonies were detected in five subjects prior to treatment and in all five, a marked decrease was observed. The CD4+/CD8+ lymphocyte ratio and peripheral NKT cell numbers increased significantly, in 7/10 and in 5/10 subjects, respectively. Significant increase in serum IL-10 and IL-4 levels was observed in 7/10 subjects during treatment period. CONCLUSION: Immune regulation via oral administration of CEP is a safe and possibly effective treatment for subjects with moderate CD and may provide means of antigen-specific immune modulation. PMID:15918198

  17. Insulin inhibition of 5' adenosine monophosphate-activated protein kinase in the heart results in activation of acetyl coenzyme A carboxylase and inhibition of fatty acid oxidation.

    PubMed

    Gamble, J; Lopaschuk, G D

    1997-11-01

    Acetyl coenzyme A (CoA) carboxylase (ACC) is an important regulator of fatty acid oxidation in the heart, since it produces malonyl CoA, a potent inhibitor of mitochondrial fatty acid uptake. Under conditions of metabolic stress, 5'adenosine monophosphate-activated protein kinase (AMPK), which is highly expressed in cardiac muscle, can phosphorylate and decrease ACC activity. In this study, we determined if fatty acid oxidation in the heart could be regulated by insulin, due to alterations in AMPK regulation of ACC activity. Isolated working rat hearts were perfused with Krebs-Henseleit solution containing 11 mmol/L glucose, 0.4 mmol/L [9,10(-3)H]palmitate, and either 100 microU/mL insulin or 1,000 microU/mL insulin. Increasing insulin concentration resulted in a decrease in fatty acid oxidation rates (P < .05), a decrease in AMPK activity (P < .05), and an increase in ACC activity (P < .05) compared with the low-insulin group. A negative correlation was observed between AMPK and ACC activity (r = -.76). We conclude that insulin, acting through inhibition of AMPK and stimulation of ACC, is capable of inhibiting myocardial fatty acid oxidation. PMID:9361684

  18. Slow Proton Transfer Coupled to Unfolding Explains the Puzzling Results of Single-Molecule Experiments on BBL, a Paradigmatic Downhill Folding Protein

    PubMed Central

    Cerminara, Michele; Campos, Luis A.; Ramanathan, Ravishankar; Muńoz, Victor

    2013-01-01

    A battery of thermodynamic, kinetic, and structural approaches has indicated that the small ?-helical protein BBL folds-unfolds via the one-state downhill scenario. Yet, single-molecule fluorescence spectroscopy offers a more conflicting view. Single-molecule experiments at pH 6 show a unique half-unfolded conformational ensemble at mid denaturation, whereas other experiments performed at higher pH show a bimodal distribution, as expected for two-state folding. Here we use thermodynamic and laser T-jump kinetic experiments combined with theoretical modeling to investigate the pH dependence of BBL stability, folding kinetics and mechanism within the pH 6–11 range. We find that BBL unfolding is tightly coupled to the protonation of one of its residues with an apparent pKa of ?7. Therefore, in chemical denaturation experiments around neutral pH BBL unfolds gradually, and also converts in binary fashion to the protonated species. Moreover, under the single-molecule experimental conditions (denaturant midpoint and 279 K), we observe that proton transfer is much slower than the ?15 microseconds folding-unfolding kinetics of BBL. The relaxation kinetics is distinctly biphasic, and the overall relaxation time (i.e. 0.2–0.5 ms) becomes controlled by the proton transfer step. We then show that a simple theoretical model of protein folding coupled to proton transfer explains quantitatively all these results as well as the two sets of single-molecule experiments, including their more puzzling features. Interestingly, this analysis suggests that BBL unfolds following a one-state downhill folding mechanism at all conditions. Accordingly, the source of the bimodal distributions observed during denaturation at pH 7–8 is the splitting of the unique conformational ensemble of BBL onto two slowly inter-converting protonation species. Both, the unprotonated and protonated species unfold gradually (one-state downhill), but they exhibit different degree of unfolding at any given condition because the native structure is less stable for the protonated form. PMID:24205082

  19. Identification of Protein Instability Determinants in the Carboxy-Terminal Region of c-Myb Removed as a Result of Retroviral Integration in Murine Monocytic Leukemias

    PubMed Central

    Bies, Juraj; Nazarov, Viktor; Wolff, Linda

    1999-01-01

    The c-myb oncogene has been a target of retroviral insertional mutagenesis in murine monocytic leukemias. One mechanism by which c-myb can be activated is through the integration of a retroviral provirus into the central portion of the locus, causing premature termination of c-myb transcription and translation. We had previously shown that a leukemia-specific c-Myb protein, truncated at the site of proviral integration by 248 amino acids, had approximately a fourfold-increased half-life compared to the normal c-Myb protein, due to its ability to escape rapid degradation by the ubiquitin-26S proteasome pathway. Here we provide evidence for the existence of more than one instability determinant in the carboxy-terminal region of the wild-type protein, which appear to act independently of each other. The data were derived from examination of premature termination mutants and deletion mutants of the normal protein, as well as analysis of another carboxy-terminally truncated protein expressed in leukemia. Evidence is provided that one instability determinant is located in the terminal 87 amino acids of the protein and another is located in the vicinity of the internal region that has leucine zipper homology. In leukemias, different degrees of protein stability are attained following proviral integration depending upon how many determinants are removed. Interestingly, although PEST sequences (rich in proline, glutamine, serine, and threonine), often associated with degradation, are found in c-Myb, deletion of PEST-containing regions had no effect on protein turnover. This study provides further insight into how inappropriate expression of c-Myb may contribute to leukemogenesis. In addition, it will facilitate further studies aimed at characterizing the specific role of individual regions of the normal protein in targeting to the 26S proteasome. PMID:9971784

  20. Downregulation of uncoupling protein-3 in vivo is linked to changes in muscle mitochondrial energy metabolism as a result of capsiate administration.

    PubMed

    Faraut, B; Giannesini, B; Matarazzo, V; Marqueste, T; Dalmasso, C; Rougon, G; Cozzone, P J; Bendahan, D

    2007-05-01

    Although it has been suggested that the skeletal muscle mitochondrial uncoupling protein-3 (UCP3) is involved in regulating energy expenditure, its role is still poorly understood. In the present study, we aimed at investigating noninvasively, using magnetic resonance techniques, metabolic changes occurring in exercising muscle as a result of capsiate treatment, which has been previously linked to UCP3 upregulation. We showed that capsiate ingestion strongly reduced UCP3 gene expression in rat gastrocnemius muscle. This large underexpression was accompanied by a significant increase in the rate of mitochondrial ATP production and phosphocreatine level both at rest and during muscle stimulation. Similarly, the stimulation-induced ATP fall and ADP accumulation were significantly less after capsiate administration than in untreated rats. The larger oxidative ATP production rate could not be explained by a proportional decrease in the anaerobic component, i.e., glycolysis and phosphocreatine breakdown. In addition, the mechanical performance was not affected by capsiate administration. Finally, the plasma free fatty acid (FFA) level increased in capsiate-treated rats, whereas no significant change was observed after muscle stimulation in the control group. Considering the corresponding enhanced UCP3 mRNA expression occurring in the control group after muscle stimulation, one can suggest that changes in FFA level and UCP3 mRNA expression are not mechanistically correlated. Overall, we have shown that capsiate administration induced a UCP3 downregulation coupled with an increased mitochondrial ATP synthesis, whereas the muscle force-generating capacity was unchanged. This suggests that a decrease in muscle efficiency and/or additional noncontractile ATP-consuming mechanisms result from UCP3 downregulation. PMID:17264228

  1. Liver-specific reduction of Mfn2 protein by RNAi results in impaired glycometabolism and lipid homeostasis in BALB/c mice.

    PubMed

    Chen, Xiaolin; Xu, Yancheng

    2009-12-01

    Mitofusin-2 (Mfn2) gene expression is positively correlated with insulin sensitivity in patients with type 2 diabetes. However, it is unclear if Mfn2 is involved in carbohydrate metabolism and lipid homeostasis. In order to investigate the specific functions of Mfn2 in glycometabolism and lipid homeostasis in BALB/c mice, a RNA interference technique-mediated hydrodynamic injection was developed, in which short hairpin RNAs (shRNAs) were used to inhibit the Mfn2 expression in vivo. Seventy-two mice were randomly divided into two groups: the Mfn2 reduction group (Mfn2/shRNA) and the negative control group (NC). Intraperitoneal glucose tolerance tests and intraperitoneal insulin tolerance tests were used to evaluate glycometabolism and insulin sensitivity. D-(3-3H) glucose or 3H2O was injected into the tail vein or intraperitoneally to facilitate the calculation of the rate of hepatic glucose production and fatty acid synthesis in vivo. The results showed that, in Mfn2/shRNA mice, the liver Mfn2 protein was significantly decreased, and fasting blood glucose concentrations were increased by approximately 48%, when compared with the NC mice. In parallel with the changes in fasting glucose levels, hepatic glucose production was significantly elevated in Mfn2/shRNA mice. When insulin was administrated, these mice exhibited impaired insulin tolerance. It was also found that the reduction of Mfn2 markedly decreased the rate of fatty acid synthesis in the liver, and the Mfn2/shRNA mice exhibited hypertriglyceridema. Taken together, our results indicate that Mfn2 plays an important role in maintaining glucose and lipid homeostasis, and in the development of insulin resistance in vivo. PMID:20037808

  2. N-Octanoyl Dopamine Treatment of Endothelial Cells Induces the Unfolded Protein Response and Results in Hypometabolism and Tolerance to Hypothermia

    PubMed Central

    Stamellou, Eleni; Fontana, Johann; Wedel, Johannes; Ntasis, Emmanouil; Sticht, Carsten; Becker, Anja; Pallavi, Prama; Wolf, Kerstin; Krämer, Bernhard K.; Hafner, Mathias; van Son, Willem J.; Yard, Benito A.

    2014-01-01

    Aim N-acyl dopamines (NADD) are gaining attention in the field of inflammatory and neurological disorders. Due to their hydrophobicity, NADD may have access to the endoplasmic reticulum (ER). We therefore investigated if NADD induce the unfolded protein response (UPR) and if this in turn influences cell behaviour. Methods Genome wide gene expression profiling, confirmatory qPCR and reporter assays were employed on human umbilical vein endothelial cells (HUVEC) to validate induction of UPR target genes and UPR sensor activation by N-octanoyl dopamine (NOD). Intracellular ATP, apoptosis and induction of thermotolerance were used as functional parameters to assess adaptation of HUVEC. Results NOD, but not dopamine dose dependently induces the UPR. This was also found for other synthetic NADD. Induction of the UPR was dependent on the redox activity of NADD and was not caused by selective activation of a particular UPR sensor. UPR induction did not result in cell apoptosis, yet NOD strongly impaired cell proliferation by attenuation of cells in the S-G2/M phase. Long-term treatment of HUVEC with low NOD concentration showed decreased intracellular ATP concentration paralleled with activation of AMPK. These cells were significantly more resistant to cold inflicted injury. Conclusions We provide for the first time evidence that NADD induce the UPR in vitro. It remains to be assessed if UPR induction is causally associated with hypometabolism and thermotolerance. Further pharmacokinetic studies are warranted to address if the NADD concentrations used in vitro can be obtained in vivo and if this in turn shows therapeutic efficacy. PMID:24926788

  3. Elevated levels of synthesis of over 20 proteins results after mutation of the Rhizobium leguminosarum exopolysaccharide synthesis gene pssA.

    PubMed

    Guerreiro, N; Ksenzenko, V N; Djordjevic, M A; Ivashina, T V; Rolfe, B G

    2000-08-01

    The protein expression profiles of Rhizobium leguminosarum strains in response to specific genetic perturbations in exopolysaccharide (EPS) biosynthesis genes were examined using two-dimensional gel electrophoresis. Lesions in either pssA, pssD, or pssE of R. leguminosarum bv. viciae VF39 or in pssA of R. leguminosarum bv. trifolii ANU794 not only abolished the capacity of these strains to synthesize EPS but also had a pleiotropic effect on protein synthesis levels. A minimum of 22 protein differences were observed for the two pssA mutant strains. The differences identified in the pssD and pssE mutants of strain VF39 were a distinct subset of the same protein synthesis changes that occurred in the pssA mutant. The pssD and pssE mutant strains shared identical alterations in the proteins synthesized, suggesting that they share a common function in the biosynthesis of EPS. In contrast, a pssC mutant that produces 38% of the EPS level of the parental strain showed no differences in its protein synthesis patterns, suggesting that the absence of EPS itself was contributing to the changes in protein synthesis and that there may be a complex interconnection of the EPS biosynthetic pathway with other metabolic pathways. Genetic complementation of pssA can restore wild-type protein synthesis levels, indicating that many of the observed differences in protein synthesis are also a specific response to a dysfunctional PssA. The relevance of these proteins, which are grouped as members of the pssA mutant stimulon, remains unclear, as the majority lacked a homologue in the current sequence databases and therefore possibly represent a novel functional network(s). These findings have illustrated the potential of proteomics to reveal unexpected higher-order processes of protein function and regulation that arise from mutation. In addition, it is evident that enzymatic pathways and regulatory networks are more interconnected and more sensitive to structural changes in the cell than is often appreciated. In these cases, linking the observed phenotype directly to the mutated gene can be misleading, as the phenotype could be attributable to downstream effects of the mutation. PMID:10913086

  4. Intracellular accumulation of a 46 kDa species of mouse prion protein as a result of loss of glycosylation in cultured mammalian cells

    Microsoft Academic Search

    Subhabrata Biswas; Jan P. M. Langeveld; Donald Tipper; Shan Lu

    2006-01-01

    Prion diseases are fatal neurodegenerative disorders characterized by the accumulation of an abnormal isoform (PrPSc) of the normal cellular prion protein (PrPC) in the brain. Reportedly, abnormal N-linked glycosylation patterns in PrPC are associated with disease susceptibility; thus, we compared the glycosylation status of normal and several mutant forms of the murine prion protein (MuPrP) in cultured mammalian cells. Substitution

  5. Depletion of the RNA-binding protein RBP33 results in increased expression of silenced RNA polymerase II transcripts in Trypanosoma brucei

    E-print Network

    Fernández-Moya, Sandra M.; Carrington, Mark; Estévez, Antonio M.

    2014-09-12

    obtained from two biological replicates using the Illumina mRNA-seq kit according to manufacturer’s instructions. Libraries were se- quenced on an Illumina platform at EASIH (Eastern Sequencing and Informatics Hub, Cambridge, UK) and the reads of 36... protein sequences for globularity and disorder. Nucleic Acids Res 31: 3701– 3708. 32. Dosztanyi Z, Csizmok V, Tompa P, Simon I (2005) IUPred: web server for the prediction of intrinsically unstructured regions of proteins based on estimated energy content...

  6. Low-dietary protein intake induces problems with glucose homeostasis and results in hepatic steatosis in heavy milk-fed calves

    Microsoft Academic Search

    W. J. J. Gerrits; J. J. G. C. van den Borne; J. W. Blum

    2008-01-01

    We studied effects of protein intake at two protein-free energy intake levels on plasma glucose and insulin concentrations, urinary glucose excretion and on liver and intestinal fat content in milk-fed veal calves. Two experiments were performed at body weights (BW) of 80–160kg (mean 120kg; Exp. 1) and 160–240kg (mean 200kg; Exp. 2). In each experiment, 36 calves were allocated to

  7. Inverse relationship between urinary markers of animal protein intake and blood pressure in Chinese: results from the WHO Cardiovascular Diseases and Alimentary Comparison (CARDIAC) Study

    Microsoft Academic Search

    Longjian Liu

    Longjian Liu,a,b Katsumi Ikeda,a Yukio Yamoria on behalf of the WHO-CARDIAC Study Group Background This issue of the relationship between animal protein intake and blood pressure (BP) is unsolved. We examined the associations between urinary 3-methylhistidine (3MH) excretion (a biological marker of animal protein intake) and BP in 11 Chinese population samples (Urumqi, Altai, Lhasa, Tulufan, Hetian, Guiyang, Guangzhou, Shanghai,

  8. Identification of precursor to cytomegalovirus capsid assembly protein and evidence that processing results in loss of its carboxy-terminal end.

    PubMed Central

    Gibson, W; Marcy, A I; Comolli, J C; Lee, J

    1990-01-01

    The 37-kilodalton (kDa) assembly protein of cytomegalovirus (strain Colburn) B capsids is shown to have a 40-kDa precursor. Pulse-chase radiolabeling experiments revealed that conversion of the precursor to the product was slow, requiring over 6 h for completion, and correlated with movement from the cytoplasmic to the nuclear fraction of Nonidet P-40-disrupted cells. Of these two proteins, only the 40-kDa precursor was synthesized in vitro from infected-cell RNA, consistent with its being the primary translation product. Amino acid sequence data obtained from CNBr-treated, high-performance liquid chromatography-purified assembly protein indicated that precursor translation begins at the first of two closely spaced potential initiation sites and that precursor maturation involves the loss of at least 32 amino acids from its carboxy-terminal end. It is also shown by immunological cross-reactivity and peptide similarity that three low-abundance B-capsid proteins (i.e., the 45-kilodalton [45K], 39K, and 38K proteins) are closely related to the assembly protein; the nature of this relatedness is discussed. Images PMID:2154607

  9. Technical decision making with higher order structure data: utilization of differential scanning calorimetry to elucidate critical protein structural changes resulting from oxidation.

    PubMed

    Arthur, Kelly K; Dinh, Nikita; Gabrielson, John P

    2015-04-01

    Differential scanning calorimetry (DSC) is a useful tool for monitoring thermal stability of the molecular conformation of proteins. Here, we present an example of the sensitivity of DSC to changes in stability arising from a common chemical degradation pathway, oxidation. This Note is part of a series of industry case studies demonstrating the application of higher order structure data for technical decision making. For this study, six protein products from three structural classes were evaluated at multiple levels of oxidation. For each protein, the melting temperature (Tm ) decreased linearly as a function of oxidation; however, differences in the rate of change in Tm , as well as differences in domain Tm stability were observed across and within structural classes. For one protein, analysis of the impact of oxidation on protein function was also performed. For this protein, DSC was shown to be a leading indicator of decreased antigen binding suggesting a subtle conformation change may be underway that can be detected using DSC prior to any observable impact on product potency. Detectable changes in oxidized methionine by mass spectrometry (MS) occurred at oxidation levels below those with a detectable conformational or functional impact. Therefore, by using MS, DSC, and relative potency methods in concert, the intricate relationship between a primary structural modification, changes in conformational stability, and functional impact can be elucidated. PMID:25561411

  10. Association between serum insulin-like growth factor I or IGF-binding protein 3 and estimated glomerular filtration rate: results of a population-based sample

    PubMed Central

    2012-01-01

    Background Insulin-like growth factor I (IGF-I), which is mostly carried in blood by IGF-binding protein 3 (IGFBP-3), was associated to the glomerular filtration rate and chronic kidney disease in a multiethnic study among US adults. The aim of the present study was to investigate whether serum IGF-I or IGFBP-3 are associated with estimated glomerular filtration rate (eGFR) in a population-based study of Caucasian adults. Methods Data from 4028 subjects (2048 women) aged 20 to 81 years from the Study of Health in Pomerania (SHIP) were analyzed. Total serum IGF-I and IGFBP-3 concentrations were determined by chemiluminescence immunoassays and categorized into sex- and age-specific quartiles. Results After adjusting for age, waist circumference and type 2 diabetes mellitus, analysis of variance (ANOVA) revealed inverse associations between serum IGF-I concentrations and eGFR in men as well as between serum IGFBP-3 concentrations and eGFR in men and women. Logistic regression analyses confirmed these findings and showed that high IGF-I or IGFBP-3 concentrations were associated with an increased risk of decreased eGFR (<60 mL/min/1.73 m2) in men or women. These relations became stronger when lower eGFR cut-offs were used for the analyses. Conclusion Our data revealed associations of increased serum IGF-I concentrations and decreased eGFR in men but not in women and an association of increased serum IGFBP-3 concentrations and decreased eGFR in both sexes. PMID:23237568

  11. Estimation of local cerebral protein synthesis rates with L-(1-/sup 11/C)leucine and PET: Methods, model, and results in animals and humans

    SciTech Connect

    Hawkins, R.A.; Huang, S.C.; Barrio, J.R.; Keen, R.E.; Feng, D.; Mazziotta, J.C.; Phelps, M.E.

    1989-08-01

    We have estimated the cerebral protein synthesis rates (CPSR) in a series of normal human volunteers and monkeys using L-(1-/sup 11/C)leucine and positron emission tomography (PET) using a three-compartment model. The model structure, consisting of a tissue precursor, metabolite, and protein compartment, was validated with biochemical assay data obtained in rat studies. The CPSR values estimated in human hemispheres of about 0.5 nmol/min/g agree well with hemispheric estimates in monkeys. The sampling requirements (input function and scanning sequence) for accurate estimates of model parameters were investigated in a series of computer simulation studies.

  12. Bacteriophage Protein–Protein Interactions

    PubMed Central

    Häuser, Roman; Blasche, Sonja; Dokland, Terje; Haggĺrd-Ljungquist, Elisabeth; von Brunn, Albrecht; Salas, Margarita; Casjens, Sherwood; Molineux, Ian

    2012-01-01

    Bacteriophages T7, ?, P22, and P2/P4 (from Escherichia coli), as well as ?29 (from Bacillus subtilis), are among the best-studied bacterial viruses. This chapter summarizes published protein interaction data of intraviral protein interactions, as well as known phage–host protein interactions of these phages retrieved from the literature. We also review the published results of comprehensive protein interaction analyses of Pneumococcus phages Dp-1 and Cp-1, as well as coliphages ? and T7. For example, the ?55 proteins encoded by the T7 genome are connected by ?43 interactions with another ?15 between the phage and its host. The chapter compiles published interactions for the well-studied phages ? (33 intra-phage/22 phage-host), P22 (38/9), P2/P4 (14/3), and ?29 (20/2). We discuss whether different interaction patterns reflect different phage lifestyles or whether they may be artifacts of sampling. Phages that infect the same host can interact with different host target proteins, as exemplified by E. coli phage ? and T7. Despite decades of intensive investigation, only a fraction of these phage interactomes are known. Technical limitations and a lack of depth in many studies explain the gaps in our knowledge. Strategies to complete current interactome maps are described. Although limited space precludes detailed overviews of phage molecular biology, this compilation will allow future studies to put interaction data into the context of phage biology. PMID:22748812

  13. Effects of proteins on protein diffusion

    PubMed Central

    Wang, Yaqiang; Li, Conggang; Pielak, Gary J.

    2010-01-01

    Despite increased attention, little is known about how the crowded intracellular environment affects basic phenomena like protein diffusion. Here, we use NMR to quantify the rotational and translational diffusion of a 7.4-kDa test protein, chymotrypsin inhibitor 2 (CI2), in solutions of glycerol, synthetic polymers, proteins, and cell lysates. As expected, translational diffusion and rotational diffusion decrease with increasing viscosity. In glycerol, for example, the decrease follows the Stokes-Einstein and Stokes-Einstein-Debye laws. Synthetic polymers cause negative deviation from the Stokes Laws and affect translation more than rotation. Surprisingly, however, protein crowders have the opposite effect, causing positive deviation and reducing rotational diffusion more than translational diffusion. Indeed, bulk proteins severely attenuate the rotational diffusion of CI2 in crowded protein solutions. Similarly, CI2 diffusion in cell lysates is comparable to its diffusion in crowded protein solutions, supporting the biological relevance of the results. The rotational attenuation is independent of the size and total charge of the crowding protein, suggesting that the effect is general. The difference between the behavior of synthetic polymers and protein crowders suggests that synthetic polymers may not be suitable mimics of the intracellular environment. NMR relaxation data reveal that the source of the difference between synthetic polymers and proteins is the presence of weak interactions between the proteins and CI2. In summary, weak but non-specific, non-covalent chemical interactions between proteins appear to fundamentally impact protein diffusion in cells. PMID:20560582

  14. MART-1 peptide vaccination plus IMP321 (LAG-3Ig fusion protein) in patients receiving autologous PBMCs after lymphodepletion: results of a Phase I trial

    PubMed Central

    2014-01-01

    Background Immunotherapy offers a promising novel approach for the treatment of cancer and both adoptive T-cell transfer and immune modulation lead to regression of advanced melanoma. However, the potential synergy between these two strategies remains unclear. Methods We investigated in 12 patients with advanced stage IV melanoma the effect of multiple MART-1 analog peptide vaccinations with (n?=?6) or without (n?=?6) IMP321 (LAG-3Ig fusion protein) as an adjuvant in combination with lymphodepleting chemotherapy and adoptive transfer of autologous PBMCs at day (D) 0 (Trial registration No: NCT00324623). All patients were selected on the basis of ex vivo detectable MART-1-specific CD8 T-cell responses and immunized at D0, 8, 15, 22, 28, 52, and 74 post-reinfusion. Results After immunization, a significant expansion of MART-1-specific CD8 T cells was measured in 83% (n?=?5/6) and 17% (n?=?1/6) of patients from the IMP321 and control groups, respectively (P?2-, >4- and >6-fold higher at D15, D30 and D60 (P?

  15. A single amino acid substitution in the hemagglutinin-neuraminidase protein of Newcastle disease virus results in increased fusion and decreased neuraminidase activities without changes in virus pathotype

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Newcastle disease virus (NDV) attachment to the host cell is mediated by the hemagglutinin-neuraminidase (HN), a multifunctional protein that has receptor recognition, neuraminidase and fusion promotion activities. The process that correlates receptor binding and fusion triggering is poorly understo...

  16. Feeding soy protein isolate (SPI) does not result in an estrogenic gene expression profile in the mammary of ovariectomized (OVX) female rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Concerns of increased breast cancer risk in women consuming soy exist because of the perceived estrogenicity of soy isoflavones. Female Sprague-Dawley rats (N equals 20/group) were fed AIN-93G diets with casein or SPI as the protein from PND30. On PND50 rats were OVX and 10/group infused s.c. with 5...

  17. S1. Sample applications of the PRS method In this section we provide results from the PRS method on different protein systems. Each

    E-print Network

    Yanikoglu, Berrin

    ) structures have been determined at 1.8 and 1.9 Ĺ resolution, respectively, by x-ray crystallography [1 of the protein yield high correlations (Ci in equation 19) with the displacements obtained from x- ray structures the displacement profiles from PRS and x-ray structures, for the apo (black curve), and the holo form (gray curve

  18. FEEDING DIETS CONTAINING RICE PROTEIN ISOLATE (RPI) RESULTS IN ACTIVATION OF HEPATIC PPAR AND LXR-REGULATED GENES IN WEANLING RATS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Feeding rice protein isolate (RPI) has been reported to prevent the development of atherosclerotic plaques in apoE knockout mice and to improve body composition indices in rats. The mechanisms remain unknown. The current study examined the effects of feeding (RPI) on expression of genes and transc...

  19. Binding of selenium-75 to blood and liver cytosolic proteins in the preruminant calf

    SciTech Connect

    Jenkins, K.J.; Hidiroglou, M.

    1988-02-01

    Labeled selenite (75Se) administered to calves in milk replacer, containing .2 or 5 ppm Se, was rapidly absorbed with peak blood 75Se at 6 h. Gel filtration and dialysis treatment of plasma and erythrocyte hemolysates showed that initially 75Se was transported in blood as 75SeO3= or loosely bound to plasma and erythrocyte proteins. At high Se intake, albumin became a transport protein for some of the plasma 75Se, and proportionately more blood radioactivity was carried in the erythrocytes. At 72 h after dosing, most plasma 75Se was tightly bound to protein in glutathione peroxidase fraction with low peroxidase activity, possibly Se transport protein. At 72 h, distribution of 75Se in erythrocyte was 35 to 40% in glutathione peroxidase, 50% in hemoglobin, and 5% in a selenite plus selenopolypeptide fraction. Erythrocyte peroxidase activity was mostly in the glutathione peroxidase fraction (57%) and hemoglobin (38%). Molecular weight estimate for erythrocyte glutathione peroxidase was 84,200 daltons; about 90% of blood peroxidase activity was in erythrocytes. High Se intake had no marked effect on distribution of 75Se among liver cytosolic proteins. About 35% of 75Se was in glutathione peroxidase fraction, having most of the peroxidase activity, 25% in void volume, 11 to 18% in a selenite plus selenopolypeptide fraction, and small amounts in selenoproteins of about 12,000 and 50,000 daltons.

  20. Rhizobium selenitireducens proteins involved in the reduction of selenite to elemental selenium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microbial based bioremediation barriers can remove the metalloid selenite (SeO3–2) from flowing groundwater. The organisms associated with the process include microorganisms from within the bacterial and archaeal domains that can reduce soluble SeO3–2 to the insoluble and reddish-colored elemental ...

  1. Perinatal Oxygen Restriction Does Not Result in Reduced Rat Frontal Cortex Synaptophysin Protein Levels at Adulthood as Opposed to Postmortem Findings in Schizophrenia

    Microsoft Academic Search

    Carmit Nadri; Galila Agam

    2009-01-01

    Synaptophysin, a synaptic vesicle protein and a marker for synaptic density has been found to be reduced in postmortem prefrontal\\u000a cortex of schizophrenia patients, consistent with evidence for synaptic deficits in schizophrenia. The contribution of both\\u000a genetic and environmental factors to the etiology of schizophrenia is well established, and obstetric complications have been\\u000a suggested as a non-genetic risk factor of

  2. Caloric Restriction Results in Decreased Expression of Peroxisome Proliferator-Activated Receptor Superfamily in Muscle of Normal and Long-Lived Growth Hormone Receptor\\/ Binding Protein Knockout Mice

    Microsoft Academic Search

    Michal M. Masternak; Khalid A. Al-Regaiey; Marc Michael Del Rosario Lim; Michael S. Bonkowski; Jacob A. Panici; Grzegorz K. Przybylski; Andrzej Bartke

    Resistance to growth hormone, reduced insulin-like growth factor 1 (IGF1) action, and enhanced insulin sensitivity are likely mediators of extended life span and delayed aging process in growth hormone receptor\\/binding protein knockout (GHR-KO) mice. Fat metabolism and genes involved in fatty acid oxidation are strongly involved in insulin action. Using real-time polymerase chain reaction and western blot we have examined

  3. Disruption of Protein Kinase C Results in Impairment of Wound Healing and Enhancement of Tumor Formation in Mouse Skin Carcinogenesis1

    Microsoft Academic Search

    Kazuhiro Chida; Takeshi Hara; Takaaki Hirai; Chieko Konishi; Kenji Nakamura; Kazuki Nakao; Atsu Aiba; Motoya Katsuki; Toshio Kuroki

    2003-01-01

    We have generated a mouse strain lacking protein kinase C (PKC) to evaluate its significance in epithelial organization and tumor formation. The PKC-deficient mice exhibited increased susceptibility to tumor formation in two-stage skin carcinogenesis by single application of 7,12- dimethylbenz(a)anthracene (DMBA) for tumor initiation and repeated applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) for tumor promotion. The tumor formation was not enhanced by

  4. Single-Site Mutations in a Hyperthermophilic Variant of the B1 Domain of Protein G Result in Self-Assembled Oligomers †

    Microsoft Academic Search

    Scott C. Meyer; Carmen Huerta; Indraneel Ghosh

    2005-01-01

    We have characterized two homologous, single-point core mutants of a 57-residue, hyper- thermophilic variant of the B1 domain of protein G (HTB1). These single-point mutations in HTB1 replace a Phe residue in the hydrophobic core with either a Glu or Asp residue. Both of these homologous core- variant mutants undergo significant structural rearrangement from the native, monomeric fold and exist

  5. Loss of mitogen-activated protein kinase kinase kinase 4 (MEKK4) results in enhanced apoptosis and defective neural tube development

    Microsoft Academic Search

    Hongbo Chi; Matthew R. Sarkisian; Pasko Rakic; Richard A. Flavell

    2005-01-01

    Neural tube defects (NTDs) are prevalent human birth defects. Mitogen-activated protein kinases (MAPKs), such as c-Jun N-terminal kinase (JNK), are implicated in facilitating neural tube closure, yet upstream regulators remain to be identified. Here, we show that MAP kinase kinase kinase 4 (MEKK4) is strongly expressed in the developing neuroepithelium. Mice deficient in MEKK4 develop highly penetrant NTDs that cannot

  6. Inhibition of CCAAT/enhancer binding protein ? expression by chrysin in microglial cells results in anti-inflammatory and neuroprotective effects.

    PubMed

    Gresa-Arribas, Núria; Serratosa, Joan; Saura, Josep; Solŕ, Carme

    2010-10-01

    The control of neuroinflammation is a potential target to be considered in the treatment of neurodegenerative diseases. It is therefore important to find anti-inflammatory drugs and study new targets that inhibit neuroinflammation. We designed an experimental model of neuroinflammation in vitro to study the anti-inflammatory and neuroprotective effects of the flavonoid chrysin and the involvement of nuclear factor-?B p65 and CCAAT/enhancer binding proteins (C/EBPs) ? and ? transcription factors in its mechanism of action. We used primary cultures of mouse embryonic cortical neurons and cultures of BV2 (murine microglial cell line) or mouse primary microglia. We induced neuronal death in neuronal-BV2/microglial co-cultures using lipopolysaccharide of Escherichia coli and interferon-?. Chrysin pre-treatment inhibited nitric oxide and tumor necrosis factor-? production, as well as inducible nitric oxide synthase expression in lipopolysaccharide E. coli and interferon-?-treated microglial cells, but did not affect cyclooxygenase-2 expression. Chrysin pre-treatment also protected neurons against the neurotoxicity induced by reactive microglial cells. These effects were associated to a decrease in C/EBP? protein level, mRNA expression, and DNA-binding activity, with no effect on C/EBP? and p65 nuclear protein levels or DNA-binding activity, pointing out C/EBP? as a possible mediator of chrysin effects. Consequently, C/EBP? is a possible target to act against neuroinflammation in neurodegenerative processes. PMID:20722966

  7. A single amino acid change resulting in loss of fluorescence of eGFP in a viral fusion protein confers fitness and growth advantage to the recombinant vesicular stomatitis virus

    SciTech Connect

    Dinh, Phat X.; Panda, Debasis; Das, Phani B.; Das, Subash C.; Das, Anshuman [School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, Nebraska 68583-0900 (United States) [School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, Nebraska 68583-0900 (United States); The Nebraska Center for Virology, University of Nebraska-Lincoln, Lincoln, Nebraska 68583-0900 (United States); Pattnaik, Asit K., E-mail: apattnaik2@unl.edu [School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, Nebraska 68583-0900 (United States); The Nebraska Center for Virology, University of Nebraska-Lincoln, Lincoln, Nebraska 68583-0900 (United States)

    2012-10-25

    Using a recombinant vesicular stomatitis virus encoding eGFP fused in-frame with an essential viral replication protein, the phosphoprotein P, we show that during passage in culture, the virus mutates the nucleotide C289 within eGFP of the fusion protein PeGFP to A or T, resulting in R97S/C amino acid substitution and loss of fluorescence. The resultant non-fluorescent virus exhibits increased fitness and growth advantage over its fluorescent counterpart. The growth advantage of the non-fluorescent virus appears to be due to increased transcription and replication activities of the PeGFP protein carrying the R97S/C substitution. Further, our results show that the R97S/C mutation occurs prior to accumulation of mutations that can result in loss of expression of the gene inserted at the G-L gene junction. These results suggest that fitness gain is more important for the recombinant virus than elimination of expression of the heterologous gene.

  8. Rats fed soy protein isolate (SPI) have impaired hepatic CYP1A1 induction by polycyclic aromatic hydrocarbons as a result of interference with aryl hydrocarbon receptor signaling

    SciTech Connect

    Singhal, Rohit [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Badger, Thomas M. [Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Arkansas Children's Nutrition Center, Little Rock, AR 72202 (United States); Ronis, Martin J. [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Arkansas Children's Nutrition Center, Little Rock, AR 72202 (United States)], E-mail: RonisMartinJ@uams.edu

    2008-03-01

    Consumption of soy diets has been found to reduce cancer incidence in animals and is associated with reduced cancer risk in humans. Previously, we have demonstrated that female Sprague-Dawley rats fed purified AIN-93G diets with soy protein isolate (SPI) as the sole protein source had reduced CYP1A1 induction and basal aryl hydrocarbon receptor (AhR) levels relative to those fed the same diet containing casein (CAS). In the present study, the molecular mechanisms underlying reduced AhR expression have been studied. The SPI-effect on AhR was not observed after feeding diets containing the purified soy isoflavones genistein or daidzein. Rat hepatoma FGC-4 cells were treated with the serum obtained from rats fed CAS- or SPI-containing diets. Reduced AhR levels (P < 0.05) were observed after 24 h exposure to SPI-serum without any changes in the overall expression of chaperone proteins-HSP90 and XAP2. SPI-serum-stimulated AhR degradation was inhibited by treating the cells with the proteasome inhibitor, MG132, and was observed to be preceded by ubiquitination of the receptor. A reduced association of XAP2 with the immunoprecipitated AhR complex was observed. SPI-serum-mediated AhR degradation was preceded by nuclear translocation of the receptor. However, the translocated receptor was found to be unable to heterodimerize with ARNT or to bind to XRE elements on the CYP1A1 enhancer. These data suggest that feeding SPI-containing diets antagonizes AhR signaling by a novel mechanism which differs from those established for known AhR antagonists.

  9. Calcium-energized motor protein forisome controls damage in phloem: potential applications as biomimetic "smart" material.

    PubMed

    Srivastava, Vineet Kumar; Tuteja, Renu; Tuteja, Narendra

    2015-06-01

    Forisomes are ATP independent, mechanically active proteins from the Fabaceae family (also called Leguminosae). These proteins are located in sieve tubes of phloem and function to prevent loss of nutrient-rich photoassimilates, upon mechanical injury/wounding. Forisomes are SEO (sieve element occlusion) gene family proteins that have recently been shown to be involved in wound sealing mechanism. Recent findings suggest that forisomes could act as an ideal model to study self assembly mechanism for the development of nanotechnological devices like microinstruments, the microfluidic system frequently used in space exploration missions. Technology enabling improvement in micro instruments has been identified as a key technology by NASA in future space exploration missions. Forisomes are designated as biomimetic smart materials which are calcium-energized motor proteins. Since forisomes are biomolecules from plant systems it can be doctored through genetic engineering. In contrast, "smart" materials which are not derived from plants are difficult to modify in their properties. Current levels of understanding about forisomes conformational shifts with respect to calcium ions and pH changes requires supplement of future advances with relation to its 3D structure to understand self assembly processes. In plant systems it forms blood clots in the form of occlusions to prevent nutrient fluid leakage and thus proves to be a unique damage control system of phloem tissue. PMID:24020505

  10. Hepatitis B virus surface protein mutations clustered mainly in CTL immune epitopes in chronic carriers: results of an Iranian nationwide study.

    PubMed

    Khedive, A; Norouzi, M; Ramezani, F; Karimzadeh, H; Alavian, S M; Malekzadeh, R; Montazeri, G; Nejatizadeh, A; Ziaee, M; Abedi, F; Ataei, B; Yaran, M; Sayad, B; Somi, M H; Sarizadeh, G; Sanei-Moghaddam, I; Mansour-Ghanaei, F; Rafatpanah, H; Pourhosseingholi, M A; Keyvani, H; Kalantari, E; Saberifiroozi, M; Judaki, M A; Ghamari, S; Daram, M; Mahabadi, M; Fazeli, Z; Goodarzi, Z; Poortahmasebi, V; Jazayeri, S M

    2013-07-01

    Mutations within the coding region of hepatitis B surface antigen (HBsAg) have been found naturally in chronic carriers. To characterize the mutations of HBsAg from Iranian chronic carriers who were vaccine and/or medication naive. The surface genes from 360 patients were amplified and directly sequenced. The distribution of amino acid substitutions was classified according to different immune epitopes of the surface protein. All isolates belonged to genotype D. 222 (61.6%) of 360 patients contained at least one amino acid substitution. 404 (74.5%) of 542 amino acid changes occurred in different immune epitopes of HBsAg, of which 112 (27.7%) in 32 residues of B-cell epitopes (62 in the 'a' determinant); 111 (27.4%) in 32 residues of T helper; and 197 (48.7%) in 32 residues inside cytotoxic T lymphocyte (CTL) epitopes. One Th (186-197) and two CTL (28-51 and 206-215) epitopes were found to be hotspot motifs for the occurrence of 213 (52.7%) substitutions. 20 stop codons were identified in different epitopes. There was a significant association between amino acid substitutions and anti-HBe seropositivity; however, the correlation between such changes with viral load and ALT levels was not significant. In chronic hepatitis B virus(HBV) carriers, positive selection in particular outside the 'a' determinant appeared to exert influence on the surface proteins. These changes could be immune escape mutations naturally occurring due to the host immune surveillance especially at the T-cell level. PMID:23730843

  11. Protein Condensation

    NASA Astrophysics Data System (ADS)

    Gunton, James D.; Shiryayev, Andrey; Pagan, Daniel L.

    2007-09-01

    Preface; 1. Introduction; 2. Globular protein structure; 3. Experimental methods; 4. Thermodynamics and statistical mechanics; 5. Protein-protein interactions; 6. Theoretical studies of equilibrium; 7. Nucleation theory; 8. Experimental studies of nucleation; 9. Lysozyme; 10. Some other globular proteins; 11. Membrane proteins; 12. Crystallins and cataracts; 13. Sickle hemoglobin and sickle cell anemia; 14, Alzheimer's disease; Index.

  12. Aberrant CCND1 copies and cyclin D1 mRNA expression do not result in the production of functional cyclin D1 protein in anaplastic large cell lymphoma.

    PubMed

    Bobos, Mattheos; Kotoula, Vassiliki; Kaloutsi, Vassiliki; Karayannopoulou, Georgia; Papadimitriou, Constantine S; Kostopoulos, Ioannis

    2009-08-01

    Scattered reports in the literature have shown that Cyclin D1 mRNA and protein may be expressed in anaplastic large cell lymphoma (ALCL). ALCLs are characterized by the presence of ALK translocations. Aberrant Cyclin D1 expression seems to promote proliferation in other types of lymphoma, while a growth promoting CCND1/TACSD1(TROP2) fusion product has also been described in tumors. Herein, we investigated 44 ALCL cases for chromosome 11 and CCND1 status (by FISH), cyclin D1 mRNA expression (by in situ hybridization and RT-PCR) and Cyclin D1 protein (by immunohistochemistry with two different monoclonal antibodies), as well as for the expression of Trop-2/GA733-1 (by immunohistochemistry). Polysomy of CCND1 (11q13) and chromosome 11 was found in 15/38 evaluated cases (39.5%). This change was specific for CD30+ neoplastic cells, as shown by double fluorescent staining. Neoplastic cells in the majority of ALCL expressed cyclin D1 mRNA (29/41 [70.7%]), in association with the presence of ALK translocations (p=0.024) and systemic, rather than cutaneous disease (p=0.021). Remarkably, however, Cyclin D1 protein was not detected in neoplastic cells (0/44 cases), neither were these found positive for Trop-2. In conclusion, aberrant copies of CCND1 / chromosome 11 may be observed in ALCL, probably as a consequence of the reported ploidy changes in these tumors. ALCL may often express cyclin D1 mRNA, which, however, does not result in the production of functional Cyclin D1 protein or Trop-2, suggesting that these proteins do not play a role in the pathogenesis of ALCL. PMID:19554511

  13. Research Results

    NASA Astrophysics Data System (ADS)

    2011-12-01

    Research on Global Carbon Emission and Sequestration NSFC Funded Project Made Significant Progress in Quantum Dynamics Functional Human Blood Protein Obtained from Rice How Giant Pandas Thrive on a Bamboo Diet New Evidence of Interpersonal Violence from 129,000 Years Ago Found in China Aptamer-Mediated Efficient Capture and Release of T Lymphocytes on Nanostructured Surfaces BGI Study Results on Resequencing 50 Accessions of Rice Cast New Light on Molecular Breeding BGI Reports Study Results on Frequent Mutation of Genes Encoding UMPP Components in Kidney Cancer Research on Habitat Shift Promoting Species Diversification

  14. Topology Independent Protein Structural Alignment

    Microsoft Academic Search

    Joe Dundas; T. Andrew Binkowski; Bhaskar Dasgupta; Jie Liang

    2007-01-01

    Protein structural alignment is an indispensable tool used for many differ- ent studies in bioinformatics. Most structural alignment algorithms assume that the structural units of two similar proteins will align sequentially. This assumption may not be true for all similar proteins and as a result, proteins with similar structure but with permuted sequence arrangement are often missed. We present a

  15. Functional clustering of yeast proteins from the protein-protein interaction network

    Microsoft Academic Search

    Taner Z. Sen; Andrzej Kloczkowski; Robert L. Jernigan

    2006-01-01

    BACKGROUND: The abundant data available for protein interaction networks have not yet been fully understood. New types of analyses are needed to reveal organizational principles of these networks to investigate the details of functional and regulatory clusters of proteins. RESULTS: In the present work, individual clusters identified by an eigenmode analysis of the connectivity matrix of the protein-protein interaction network

  16. Computational Prediction of Protein–Protein Interaction Networks: Algo-rithms and Resources

    PubMed Central

    Zahiri, Javad; Bozorgmehr, Joseph Hannon; Masoudi-Nejad, Ali

    2013-01-01

    Protein interactions play an important role in the discovery of protein functions and pathways in biological processes. This is especially true in case of the diseases caused by the loss of specific protein-protein interactions in the organism. The accuracy of experimental results in finding protein-protein interactions, however, is rather dubious and high throughput experimental results have shown both high false positive beside false negative information for protein interaction. Computational methods have attracted tremendous attention among biologists because of the ability to predict protein-protein interactions and validate the obtained experimental results. In this study, we have reviewed several computational methods for protein-protein interaction prediction as well as describing major databases, which store both predicted and detected protein-protein interactions, and the tools used for analyzing protein interaction networks and improving protein-protein interaction reliability. PMID:24396273

  17. Computational Prediction of Protein-Protein Interaction Networks: Algo-rithms and Resources.

    PubMed

    Zahiri, Javad; Bozorgmehr, Joseph Hannon; Masoudi-Nejad, Ali

    2013-09-01

    Protein interactions play an important role in the discovery of protein functions and pathways in biological processes. This is especially true in case of the diseases caused by the loss of specific protein-protein interactions in the organism. The accuracy of experimental results in finding protein-protein interactions, however, is rather dubious and high throughput experimental results have shown both high false positive beside false negative information for protein interaction. Computational methods have attracted tremendous attention among biologists because of the ability to predict protein-protein interactions and validate the obtained experimental results. In this study, we have reviewed several computational methods for protein-protein interaction prediction as well as describing major databases, which store both predicted and detected protein-protein interactions, and the tools used for analyzing protein interaction networks and improving protein-protein interaction reliability. PMID:24396273

  18. A bivalent Neisseria meningitidis recombinant lipidated factor H binding protein vaccine in young adults: results of a randomised, controlled, dose-escalation phase 1 trial.

    PubMed

    Richmond, P C; Nissen, M D; Marshall, H S; Lambert, S B; Roberton, D; Gruber, W C; Jones, T R; Arora, A

    2012-09-21

    Neisseria meningitidis is a leading cause of meningitis and septicaemia, but a broadly-protective vaccine against endemic serogroup B disease is not licensed and available. The conserved, outer-membrane lipoprotein factor H binding protein (fHBP, also known as LP2086) is expressed as one of two subfamily variants in virtually all meningococci. This study investigated the safety, tolerability, and immunogenicity of a recombinant-expressed bivalent fHBP (r-fHBP) vaccine in healthy adults. Participants (N=103) aged 18-25 years were recruited into three ascending dose level cohorts of 20, 60, and 200?g of a bivalent r-fHBP vaccine formulation and randomised to receive vaccine or placebo at 0, 1, and 6 months. The vaccine was well tolerated. Geometric mean titres (GMTs) for r-fHBP subfamily-specific IgG antibodies increased 19-168-fold from pre-vaccination to post-dose 2 in a dose level-dependent manner. In addition, robust serum bactericidal assay using human complement (hSBA) responses for strains expressing both homologous and heterologous fHBP variants were observed. After three vaccinations, 16-52% of the placebo group and 47-90%, 75-100%, and 88-100%, of the 20, 60, and 200?g dose levels, respectively, had seroprotective (? 1:4) hSBA titres against six serogroup B strains. The bivalent r-fHBP vaccine was well tolerated and induced robust bactericidal activity against six diverse serogroup B strains in young adults at the 60 and 200?g dose levels. PMID:22871351

  19. Toxicity of selenium (Na sub 2 SeO sub 3 ) and mercury (HgCl sub 2 ) on the planarian Dugesia gonocephala

    SciTech Connect

    Congiu, A.M.; Casu, S.; Ugazio, G. (Istituto di Genetica (Italy))

    1989-10-01

    The toxicity of selenium (Na{sub 2}SeO{sub 3}) and mercury (HgCl{sub 2}) was determined by using a freshwater planarian which is particularly sensitive to pollution, and belongs to a fissiparous breed of Dugesia gonocephala. The mortality and fissiparity frequency of the subjects were studied. They were exposed to intense treatments (48 hours) or for medium to long periods of time (21 days) to either the single compounds or a combination of both, and were fed or fasting. The lethal effect of sodium selenite is correlated to the food intake, whereas the toxicity of mercurous chloride is probably the result of a fixative effect which does not depend on feeding. The 21-day treatment with the first compound has a non-negligible lethal effect which is probably due to an accumulation phenomenon. At doses where an antioxidant effect prevails, fissiparity is stimulated. On the other hand, the second compound reduces reproduction frequency to half the base values. Compared to the Paracentrotus lividus, the Dugesia gonocephala offers various advantages concerning toxicological experiments; besides being easier to handle in the laboratory, it is available all year round and is not subject to seasonal cycles. It is also more susceptible to the toxic effect of mercury, which is a common and highly toxic pollutant, than the sea urchin.

  20. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2012 BINF 731 Protein Engineering Protein Engineering Engineering Protein Engineering #12;Protein Engineering Protein Engineering Protein Engineering Protein Engineering Protein Engineering #12;

  1. Protein Complexes in Bacteria

    PubMed Central

    Caufield, J. Harry; Abreu, Marco; Wimble, Christopher; Uetz, Peter

    2015-01-01

    Large-scale analyses of protein complexes have recently become available for Escherichia coli and Mycoplasma pneumoniae, yielding 443 and 116 heteromultimeric soluble protein complexes, respectively. We have coupled the results of these mass spectrometry-characterized protein complexes with the 285 “gold standard” protein complexes identified by EcoCyc. A comparison with databases of gene orthology, conservation, and essentiality identified proteins conserved or lost in complexes of other species. For instance, of 285 “gold standard” protein complexes in E. coli, less than 10% are fully conserved among a set of 7 distantly-related bacterial “model” species. Complex conservation follows one of three models: well-conserved complexes, complexes with a conserved core, and complexes with partial conservation but no conserved core. Expanding the comparison to 894 distinct bacterial genomes illustrates fractional conservation and the limits of co-conservation among components of protein complexes: just 14 out of 285 model protein complexes are perfectly conserved across 95% of the genomes used, yet we predict more than 180 may be partially conserved across at least half of the genomes. No clear relationship between gene essentiality and protein complex conservation is observed, as even poorly conserved complexes contain a significant number of essential proteins. Finally, we identify 183 complexes containing well-conserved components and uncharacterized proteins which will be interesting targets for future experimental studies. PMID:25723151

  2. Forces Stabilizing Proteins

    PubMed Central

    Pace, C. Nick; Scholtz, J. Martin; Grimsley, Gerald R.

    2014-01-01

    The goal of this article is to summarize what has been learned about the major forces stabilizing proteins since the late 1980s when site-directed mutagenesis became possible. The following conclusions are derived from experimental studies of hydrophobic and hydrogen bonding variants. 1. Based on studies of 138 hydrophobic interaction variants in 11 proteins, burying a –CH2– group on folding contributes 1.1 ± 0.5 kcal/mol to protein stability. 2. The burial of nonpolar side chains contributes to protein stability in two ways: first, a term that depends on the removal of the side chains from water and, more importantly, the enhanced London dispersion forces that result from the tight packing in the protein interior. 3. Based on studies of 151 hydrogen bonding variants in 15 proteins, forming a hydrogen bond on folding contributes 1.1 ± 0.8 kcal/mol to protein stability. 4. The contribution of hydrogen bonds to protein stability is strongly context dependent. 5. Hydrogen bonds by side chains and peptide groups make similar contributions to protein stability. 6. Polar group burial can make a favorable contribution to protein stability even if the polar group is not hydrogen bonded. 7. Hydrophobic interactions and hydrogen bonds both make large contributions to protein stability. PMID:24846139

  3. Folding scene investigation: membrane proteins

    PubMed Central

    Booth, Paula J; Curnow, Paul

    2009-01-01

    Investigations into protein folding have concentrated on experimentally tractable proteins with the result that membrane protein folding remains unsolved. New evidence is providing insight into the nature of the interactions stabilising the folded state of ?-helical membrane proteins as well as giving hints on the character of the folding transition state. These developments show that classical methods used for water-soluble proteins can be successfully adapted for membrane proteins. The advances, coupled with increasing numbers of solved crystal structures, augur well for future research into the mechanisms of membrane protein folding. PMID:19157854

  4. Constitutive or seed-specific overexpression of Arabidopsis G-protein ? subunit 3 (AGG3) results in increased seed and oil production and improved stress tolerance in Camelina sativa.

    PubMed

    Roy Choudhury, Swarup; Riesselman, Adam J; Pandey, Sona

    2014-01-01

    Heterotrimeric G-proteins consisting of G?, G? and G? subunits play an integral role in mediating multiple signalling pathways in plants. A novel, recently identified plant-specific G? protein, AGG3, has been proposed to be an important regulator of organ size and mediator of stress responses in Arabidopsis, whereas its potential homologs in rice are major quantitative trait loci for seed size and panicle branching. To evaluate the role of AGG3 towards seed and oil yield improvement, the gene was overexpressed in Camelina sativa, an oilseed crop of the Brassicaceae family. Analysis of multiple homozygous T4 transgenic Camelina lines showed that constitutive overexpression of AGG3 resulted in faster vegetative as well as reproductive growth accompanied by an increase in photosynthetic efficiency. Moreover, when expressed constitutively or specifically in seed tissue, AGG3 was found to increase seed size, seed mass and seed number per plant by 15%-40%, effectively resulting in significantly higher oil yield per plant. AGG3 overexpressing Camelina plants also exhibited improved stress tolerance. These observations draw a strong link between the roles of AGG3 in regulating two critical yield parameters, seed traits and plant stress responses, and reveal an effective biotechnological tool to dramatically increase yield in agricultural crops. PMID:24102738

  5. Concurrent Breakpoints Chang-Seo Park

    E-print Network

    Sen, Koushik

    breakpoints, a light-weight and programmatic way to make a concurrency bug reproducible. We describe the bug. In this paper, we propose a simple light-weight technique called concurrent breakpoints, reproducibility of bugs is a key requirement. Unfortunately, bugs in concurrent programs are notoriously diffi

  6. Protein- protein interaction detection system using fluorescent protein microdomains

    DOEpatents

    Waldo, Geoffrey S. (Santa Fe, NM); Cabantous, Stephanie (Los Alamos, NM)

    2010-02-23

    The invention provides a protein labeling and interaction detection system based on engineered fragments of fluorescent and chromophoric proteins that require fused interacting polypeptides to drive the association of the fragments, and further are soluble and stable, and do not change the solubility of polypeptides to which they are fused. In one embodiment, a test protein X is fused to a sixteen amino acid fragment of GFP (.beta.-strand 10, amino acids 198-214), engineered to not perturb fusion protein solubility. A second test protein Y is fused to a sixteen amino acid fragment of GFP (.beta.-strand 11, amino acids 215-230), engineered to not perturb fusion protein solubility. When X and Y interact, they bring the GFP strands into proximity, and are detected by complementation with a third GFP fragment consisting of GFP amino acids 1-198 (strands 1-9). When GFP strands 10 and 11 are held together by interaction of protein X and Y, they spontaneous association with GFP strands 1-9, resulting in structural complementation, folding, and concomitant GFP fluorescence.

  7. Protein folds and functions

    Microsoft Academic Search

    Andrew CR Martin; Christine A Orengo; E Gail Hutchinson; Susan Jones; Maria Karmirantzou; Roman A Laskowski; John BO Mitchell; Chiara Taroni; Janet M Thornton

    1998-01-01

    Background: The recent rapid increase in the number of available three-dimensional protein structures has further highlighted the necessity to understand the relationship between biological function and structure. Using structural classification schemes such as SCOP, CATH and DALI, it is now possible to explore global relationships between protein fold and function, something which was previously impractical.Results: Using a relational database of

  8. Debittering of protein hydrolyzates

    Microsoft Academic Search

    Badal C Saha; Kiyoshi Hayashi

    2001-01-01

    Enzymatic hydrolysis of proteins frequently results in bitter taste, which is due to the formation of low molecular weight peptides composed of mainly hydrophobic amino acids. Methods for debittering of protein hydrolyzates include selective separation such as treatment with activated carbon, extraction with alcohol, isoelectric precipitation, chromatography on silica gel, hydrophobic interaction chromatography, and masking of bitter taste. Bio-based methods

  9. Beyond proteins.

    PubMed

    Robson, B

    1999-08-01

    Increased understanding of the biological principles of protein structure and folding, combined with advances in protein-synthetic chemistry, should not only allow us to borrow from biology but also to depart from it and so produce protein-like, but non-protein, molecules and molecular devices. However, radical departures from protein-like forms into more-robust and truly novel 'smart' polymers and materials first require a solution to the protein-folding problem using only fundamental physicochemical principles. Any such practical solution may not come from raw computing power alone but rather from a deeper understanding of topological principles. PMID:10407402

  10. Physics of protein motility and motor proteins

    NASA Astrophysics Data System (ADS)

    Kolomeisky, Anatoly B.

    2013-09-01

    Motor proteins are enzymatic molecules that transform chemical energy into mechanical motion and work. They are critically important for supporting various cellular activities and functions. In the last 15 years significant progress in understanding the functioning of motor proteins has been achieved due to revolutionary breakthroughs in single-molecule experimental techniques and strong advances in theoretical modelling. However, microscopic mechanisms of protein motility are still not well explained, and the collective efforts of many scientists are needed in order to solve these complex problems. In this special section the reader will find the latest advances on the difficult road to mapping motor proteins dynamics in various systems. Recent experimental developments have allowed researchers to monitor and to influence the activity of single motor proteins with a high spatial and temporal resolution. It has stimulated significant theoretical efforts to understand the non-equilibrium nature of protein motility phenomena. The latest results from all these advances are presented and discussed in this special section. We would like to thank the scientists from all over the world who have reported their latest research results for this special section. We are also grateful to the staff and editors of Journal of Physics: Condensed Matter for their invaluable help in handling all the administrative and refereeing activities. The field of motor proteins and protein motility is fast moving, and we hope that this collection of articles will be a useful source of information in this highly interdisciplinary area. Physics of protein motility and motor proteins contents Physics of protein motility and motor proteinsAnatoly B Kolomeisky Identification of unique interactions between the flexible linker and the RecA-like domains of DEAD-box helicase Mss116 Yuan Zhang, Mirkó Palla, Andrew Sun and Jung-Chi Liao The load dependence of the physical properties of a molecular motor Xianchao Meng, Min Yu and Yunxin Zhang Microtubule organization by kinesin motors and microtubule crosslinking protein MAP65 Joshua Pringle, Amutha Muthukumar, Amanda Tan, Laura Crankshaw, Leslie Conway and Jennifer L Ross Backtracking dynamics of RNA polymerase: pausing and error correction Mamata Sahoo and Stefan Klumpp First-passage problems in DNA replication: effects of template tension on stepping and exonuclease activities of a DNA polymerase motor Ajeet K Sharma and Debashish Chowdhury

  11. (Protein engineering)

    SciTech Connect

    Mural, R.J.

    1987-04-21

    The traveler attended an international meeting, Protein Engineering 87','' held at the University of Oxford. Speakers of international standing addressed 425 conferees on topics ranging from the theoretical aspects of protein structure to the therapeutic uses of engineered'' proteins. The traveler presented work of the Protein Engineering Program of the Biology Division at a poster session at this Conference. There were a number of opportunities to interact with colleagues and exchange information concerning this new and rapidly growing field.

  12. Ribosomal proteins

    Microsoft Academic Search

    E. Kaltschmidt; G. Stöffler; M. Dzionara; H. G. Wittmann

    1970-01-01

    The ribosomal proteins fromE. coli strains B, C, K12 (A19), and MRE600 were extracted and analyzed by two-dimensional polyacrylamide gel electrophoresis. All four strains were found to be indistinguishable in their 50S ribosomal protein components, while there were differences among the 30S proteins. Strains K and B differ in protein S5 and S7. Strain C differs from strain B in

  13. Protein Microarrays

    NASA Astrophysics Data System (ADS)

    Ricard-Blum, S.

    Proteins are key actors in the life of the cell, involved in many physiological and pathological processes. Since variations in the expression of messenger RNA are not systematically correlated with variations in the protein levels, the latter better reflect the way a cell functions. Protein microarrays thus supply complementary information to DNA chips. They are used in particular to analyse protein expression profiles, to detect proteins within complex biological media, and to study protein-protein interactions, which give information about the functions of those proteins [3-9]. They have the same advantages as DNA microarrays for high-throughput analysis, miniaturisation, and the possibility of automation. Section 18.1 gives a brief overview of proteins. Following this, Sect. 18.2 describes how protein microarrays can be made on flat supports, explaining how proteins can be produced and immobilised on a solid support, and discussing the different kinds of substrate and detection method. Section 18.3 discusses the particular format of protein microarrays in suspension. The diversity of protein microarrays and their applications are then reported in Sect. 18.4, with applications to therapeutics (protein-drug interactions) and diagnostics. The prospects for future developments of protein microarrays are then outlined in the conclusion. The bibliography provides an extensive list of reviews and detailed references for those readers who wish to go further in this area. Indeed, the aim of the present chapter is not to give an exhaustive or detailed analysis of the state of the art, but rather to provide the reader with the basic elements needed to understand how proteins are designed and used.

  14. Ultrafiltration of pegylated proteins

    NASA Astrophysics Data System (ADS)

    Molek, Jessica R.

    There is considerable clinical interest in the use of "second-generation" therapeutics produced by conjugation of a native protein with various polymers including polyethylene glycol (PEG). PEG--protein conjugates, so-called PEGylated proteins, can exhibit enhanced stability, half-life, and bioavailability. One of the challenges in the commercial production of PEGylated proteins is the purification required to remove unreacted polymer, native protein, and in many cases PEGylated proteins with nonoptimal degrees of conjugation. The overall objective of this thesis was to examine the use of ultrafiltration for the purification of PEGylated proteins. This included: (1) analysis of size-based separation of PEGylated proteins using conventional ultrafiltration membranes, (2) use of electrically-charged membranes to exploit differences in electrostatic interactions, and (3) examination of the effects of PEGylation on protein fouling. The experimental results were analyzed using appropriate theoretical models, with the underlying physical properties of the PEGylated proteins evaluated using size exclusion chromatography, capillary electrophoresis, dynamic light scattering, and reverse phase chromatography. PEGylated proteins were produced by covalent attachment of activated PEG to a protein via primary amines on the lysine residues. A simple model was developed for the reaction kinetics, which was used to explore the effect of reaction conditions and mode of operation on the distribution of PEGylated products. The effective size of the PEGylated proteins was evaluated using size exclusion chromatography, with appropriate correlations developed for the size in terms of the molecular weight of the native protein and attached PEG. The electrophoretic mobility of the PEGylated proteins were evaluated by capillary electrophoresis with the data in good agreement with a simple model accounting for the increase in protein size and the reduction in the number of protonated amine groups in the PEGylated proteins. Ultrafiltration experiments were performed using PEGylated alpha-lactalbumin, ovalbumin, and bovine serum albumin. In contrast to the size exclusion chromatography data, the sieving coefficient of the PEGylated proteins depended upon both the number and size of the attached PEG chains due to the elongation or deformation of the PEG associated with the filtrate flux. Sieving coefficients at low filtrate flux were in good agreement with predictions of available hydrodynamic models, with significant elongation occurring when the Deborah number for the PEG chain exceeded 0.001. The effects of electrostatic interactions on the ultrafiltration of PEGylated proteins were examined using electrically-charged membranes generated by covalent attachment of sulphonic acid groups to the base cellulosic membrane. Transmission of PEGylated proteins through charged membranes was dramatically reduced at low ionic strength due to strong electrostatic interactions, despite the presence of the neutral PEG. The experimental results were in good agreement with model calculations developed for the partitioning of charged spheres into charged cylindrical pores. The experimental and theoretical results provide the first quantitative analysis of the effects of PEGylation on transport through semipermeable ultrafiltration membranes. The results from small-scale ultrafiltration experiments were used to develop a two-stage diafiltration process to purify PEGylated alpha-lactalbumin. The first-stage used a neutral membrane to remove the unreacted protein by exploiting differences in size. The second stage used a negatively-charged membrane to remove hydrolyzed PEG, with the PEGylated product retained by strong electrostatic interactions. This process provided a purification factor greater than 1000 with respect to the unreacted protein and greater than 20-fold with respect to the PEG with an overall yield of PEGylated alpha-lactalbumin of 78%. These results provide the first demonstration of the potential of using ultrafiltration for the purificatio

  15. Effects of 4 weight-loss diets differing in fat, protein, and carbohydrate on fat mass, lean mass, visceral adipose tissue, and hepatic fat: results from the POUNDS LOST trial123

    PubMed Central

    de Souza, Russell J; Carey, Vincent J; Hall, Kevin D; LeBoff, Meryl S; Loria, Catherine M; Laranjo, Nancy M; Sacks, Frank M; Smith, Steven R

    2012-01-01

    Background: Weight loss reduces body fat and lean mass, but whether these changes are influenced by macronutrient composition of the diet is unclear. Objective: We determined whether energy-reduced diets that emphasize fat, protein, or carbohydrate differentially reduce total, visceral, or hepatic fat or preserve lean mass. Design: In a subset of participants in a randomized trial of 4 weight-loss diets, body fat and lean mass (n = 424; by using dual-energy X-ray absorptiometry) and abdominal and hepatic fat (n = 165; by using computed tomography) were measured after 6 mo and 2 y. Changes from baseline were compared between assigned amounts of protein (25% compared with 15%) and fat (40% compared with 20%) and across 4 carbohydrate amounts (35% through 65%). Results: At 6 mo, participants lost a mean (±SEM) of 4.2 ± 0.3 kg (12.4%) fat and 2.1 ± 0.3 kg (3.5%) lean mass (both P < 0.0001 compared with baseline values), with no differences between 25% and 15% protein (P ? 0.10), 40% and 20% fat (P ? 0.34), or 65% and 35% carbohydrate (P ? 0.27). Participants lost 2.3 ± 0.2 kg (13.8%) abdominal fat: 1.5 ± 0.2 kg (13.6%) subcutaneous fat and 0.9 ± 0.1 kg (16.1%) visceral fat (all P < 0.0001 compared with baseline values), with no differences between the diets (P ? 0.29). Women lost more visceral fat than did men relative to total-body fat loss. Participants regained ?40% of these losses by 2 y, with no differences between diets (P ? 0.23). Weight loss reduced hepatic fat, but there were no differences between groups (P ? 0.28). Dietary goals were not fully met; self-reported contrasts were closer to 2% protein, 8% fat, and 14% carbohydrate at 6 mo and 1%, 7%, and 10%, respectively, at 2 y. Conclusion: Participants lost more fat than lean mass after consumption of all diets, with no differences in changes in body composition, abdominal fat, or hepatic fat between assigned macronutrient amounts. This trial was registered at clinicaltrials.gov as NCT00072995. PMID:22258266

  16. Proteins Wriggle

    Microsoft Academic Search

    Michael Cahill; Sean Cahill; Kevin Cahill

    2002-01-01

    We propose an algorithmic strategy for improving the efficiency of Monte Carlo searches for the low-energy states of proteins. Our strategy is motivated by a model of how proteins alter their shapes. In our model, when proteins fold under physiological conditions, their backbone dihedral angles change synchronously in groups of four or more to avoid steric clashes and respect the

  17. Protein-protein interactions: methods for detection and analysis.

    PubMed Central

    Phizicky, E M; Fields, S

    1995-01-01

    The function and activity of a protein are often modulated by other proteins with which it interacts. This review is intended as a practical guide to the analysis of such protein-protein interactions. We discuss biochemical methods such as protein affinity chromatography, affinity blotting, coimmunoprecipitation, and cross-linking; molecular biological methods such as protein probing, the two-hybrid system, and phage display: and genetic methods such as the isolation of extragenic suppressors, synthetic mutants, and unlinked noncomplementing mutants. We next describe how binding affinities can be evaluated by techniques including protein affinity chromatography, sedimentation, gel filtration, fluorescence methods, solid-phase sampling of equilibrium solutions, and surface plasmon resonance. Finally, three examples of well-characterized domains involved in multiple protein-protein interactions are examined. The emphasis of the discussion is on variations in the approaches, concerns in evaluating the results, and advantages and disadvantages of the techniques. PMID:7708014

  18. Diets containing soy or rice protein isolate increase insulin sensitivity and improve lipid homeostasis in weanling rats fed high fat, high cholesterol Western diets as a result of activation of PPAR and LXR-mediated pathways

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The current study examined the effects of feeding soy protein isolate (SPI) and rice protein isolate (RPI) on insulin sensitivity and fat breakdown in weanling rats consuming high fat/high cholesterol diets. Male Sprague-Dawley rats were placed on semi-purified diets containing the milk protein case...

  19. Coverage of protein domain families with structural protein-protein interactions: current progress and future trends.

    PubMed

    Goncearenco, Alexander; Shoemaker, Benjamin A; Zhang, Dachuan; Sarychev, Alexey; Panchenko, Anna R

    2014-01-01

    Protein interactions have evolved into highly precise and regulated networks adding an immense layer of complexity to cellular systems. The most accurate atomistic description of protein binding sites can be obtained directly from structures of protein complexes. The availability of structurally characterized protein interfaces significantly improves our understanding of interactomes, and the progress in structural characterization of protein-protein interactions (PPIs) can be measured by calculating the structural coverage of protein domain families. We analyze the coverage of protein domain families (defined according to CDD and Pfam databases) by structures, structural protein-protein complexes and unique protein binding sites. Structural PPI coverage of currently available protein families is about 30% without any signs of saturation in coverage growth dynamics. Given the current growth rates of domain databases and structural PPI deposition, complete domain coverage with PPIs is not expected in the near future. As a result of this study we identify families without any protein-protein interaction evidence (listed on a supporting website http://www.ncbi.nlm.nih.gov/Structure/ibis/coverage/) and propose them as potential targets for structural studies with a focus on protein interactions. PMID:24931138

  20. Ubiquitin Domain Proteins in Disease

    Microsoft Academic Search

    Louise Madsen; Andrea Schulze; Michael Seeger; Rasmus Hartmann-Petersen

    2007-01-01

    The human genome encodes several ubiquitin-like (UBL) domain proteins (UDPs). Members of this protein family are involved in a variety of cellular functions and many are connected to the ubiquitin proteasome system, an essential pathway for protein degradation in eukaryotic cells. Despite their structural similarity, the UBL domains appear to have a range of different targets, resulting in a considerable

  1. Ubiquitin domain proteins in disease

    Microsoft Academic Search

    Louise Madsen; Andrea Schulze; Michael Seeger; Rasmus Hartmann-Petersen

    2007-01-01

    The human genome encodes several ubiquitin-like (UBL) domain proteins (UDPs). Members of this protein family are involved in a variety of cellular functions and many are connected to the ubiquitin proteasome system, an essential pathway for protein degradation in eukaryotic cells. Despite their structural similarity, the UBL domains appear to have a range of different targets, resulting in a considerable

  2. PRISM: Protein-protein Interaction Prediction by Structural Matching

    PubMed Central

    Keskin, Ozlem; Nussinov, Ruth; Gursoy, Attila

    2009-01-01

    Prism (Protein Interactions by Structural Matching) is a system which employs a novel prediction algorithm for protein-protein interactions. It adopts a bottom-up approach that combines structure and sequence conservation in protein interfaces. The algorithm seeks possible binary interactions between proteins through the structure similarity and evolutionary conservation of known interfaces. It is composed of a database holding protein interface structures derived from the Protein Data Bank (PDB) and predicted protein-protein interactions. It also provides related information about the proteins, and an interactive protein interface viewer. In the current version, 3799 structurally non-redundant interfaces are used to predict the interactions among 6170 proteins. A substantial number of interactions are verified in two publicly available interaction databases (DIP and BIND). As the verified interactions demonstrate the suitability of our approach, unverified ones may point to undiscovered interactions. Prism can be accessed through a user friendly web site (http://prism.ccbb.ku.edu.tr) and it is planned to be updated regularly as new protein structures become available in the PDB. Users may browse through the non-redundant dataset of representative interfaces which the prediction algorithm depends on, retrieve the list of similar structures to these interfaces, or see the results of interaction predictions for a particular protein. Another service provided is the interactive prediction. This is done by running the algorithm for the user input structures. PMID:18592198

  3. A framework for protein structure classification and identification of novel protein structures

    Microsoft Academic Search

    You Jung Kim; Jignesh M. Patel

    2006-01-01

    Background: Protein structure classification plays a central role in understanding the function of a protein molecule with respect to all known proteins in a structure database. With the rapid increase in the number of new protein structures, the need for automated and accurate methods for protein classification is increasingly important. Results: In this paper we present a unified framework for

  4. A novel functional module detection algorithm for protein-protein interaction networks

    Microsoft Academic Search

    Woochang Hwang; Young-rae Cho; Aidong Zhang; Murali Ramanathan

    2006-01-01

    BACKGROUND: The sparse connectivity of protein-protein interaction data sets makes identification of functional modules challenging. The purpose of this study is to critically evaluate a novel clustering technique for clustering and detecting functional modules in protein-protein interaction networks, termed STM. RESULTS: STM selects representative proteins for each cluster and iteratively refines clusters based on a combination of the signal transduced

  5. Activation of human lymphocytes by concanavalin A or purified protein derivative results in no alteration of fluorescence polarization of lipid probes although the electrophoretic mobility of the cells is changed.

    PubMed

    Parola, A H; Kaplan, J H; Lockwood, S H; Uzgiris, E E

    1981-12-21

    Upon stimulation with either concanavalin A or the tuberculin antigen, purified protein derivative, human peripheral blood lymphocytes, purified on Ficoll-Hypaque, did not exhibit a concomitant lipid fluidity alteration as measured by fluorescence polarization (P) of the lipid probe, 1,6-diphenyl-1,3-5-hexatriene (DPH). This result was independent of the incubation period, ranging from 10 min to 72 h. However, a general reduction in polarization value, from P = 0.287 (maintained for up to 2 h of incubation) to P = 0.225 after 20 h was observed for both experimental and control samples. Moreover, fluorescence polarization studies of the nonpenetrating modified DPH cationic lipid probe, 1-[4'-trimethylaminophenyl]-6-phenyl-1,3-5-hexatriene (TMA-DPH), also failed to show any change in lipid fluidity subsequent to a 1-3 h incubation of lymphocytes with concanavalin A. Cell electrophoretic mobility, however, was altered (mean cell mobility increased by 10-15%) in a fast response to stimulation and was observed within several hours of in vitro application of concanavalin A and purified protein derivative. This initial response disappeared with further incubation at 37 degrees C (greater than 3 h) and was followed by a decline of cellular mobility of the concanavalin A-exposed cells after 48 and 72 h of incubation. The unstimulated control cells did not change in mobility as a function of incubation time. The slow decline in mean cell mobility of the experimental cells is believed to be associated with blastogenesis. It is concluded that neither blastogenic transformation nor short term membrane alterations associated with human lymphocyte activation lead to lipid fluidity changes as measured in steady state by the fluorescence polarization of both DPH and TMA-DPH. PMID:7317420

  6. Transport Proteins

    NSDL National Science Digital Library

    Jack D. Thatcher (Lewisburg; West Virginia School of Osteopathic Medicine REV)

    2013-04-16

    This Teaching Resource provides and describes two animated lessons that illustrate general properties of transport proteins. The lesson called “transport protein classes” depicts major classes and subclasses of transport proteins. The “transporters, mechanism of action” lesson explains how transporters and P class ATPase (adenosine triphosphatase) pumps function. These animations serve as valuable resources for any collegiate-level course that describes these important factors. Courses that might use them include introductory biology, biochemistry, cell biology, physiology, and biophysics.

  7. Balanced Protein-Water Interactions Improve Properties of Disordered Proteins and Non-Specific Protein Association.

    PubMed

    Best, Robert B; Zheng, Wenwei; Mittal, Jeetain

    2014-11-11

    Some frequently encountered deficiencies in all-atom molecular simulations, such as nonspecific protein-protein interactions being too strong, and unfolded or disordered states being too collapsed, suggest that proteins are insufficiently well solvated in simulations using current state-of-the-art force fields. To address these issues, we make the simplest possible change, by modifying the short-range protein-water pair interactions, and leaving all the water-water and protein-protein parameters unchanged. We find that a modest strengthening of protein-water interactions is sufficient to recover the correct dimensions of intrinsically disordered or unfolded proteins, as determined by direct comparison with small-angle X-ray scattering (SAXS) and Förster resonance energy transfer (FRET) data. The modification also results in more realistic protein-protein affinities, and average solvation free energies of model compounds which are more consistent with experiment. Most importantly, we show that this scaling is small enough not to affect adversely the stability of the folded state, with only a modest effect on the stability of model peptides forming ?-helix and ?-sheet structures. The proposed adjustment opens the way to more accurate atomistic simulations of proteins, particularly for intrinsically disordered proteins, protein-protein association, and crowded cellular environments. PMID:25400522

  8. Protein coadaptation and the design of novel approaches to identify protein-protein interactions.

    PubMed

    Fares, Mario A; Ruiz-González, Mario X; Labrador, Juan Pablo

    2011-04-01

    Proteins rarely function in isolation but they form part of complex networks of interactions with other proteins within or among cells. The importance of a particular protein for cell viability is directly dependent upon the number of interactions where it participates and the function it performs: the larger the number of interactions of a protein the greater its functional importance is for the cell. With the advent of genome sequencing and "omics" technologies it became feasible conducting large-scale searches for protein interacting partners. Unfortunately, the accuracy of such analyses has been underwhelming owing to methodological limitations and to the inherent complexity of protein interactions. In addition to these experimental approaches, many computational methods have been developed to identify protein-protein interactions by assuming that interacting proteins coevolve resulting from the coadaptation dynamics between the amino acids of their interacting faces. We review the main technological advances made in the field of interactomics and discuss the feasibility of computational methods to identify protein-protein interactions based on the estimation of coevolution. As proof-of-concept, we present a classical case study: the interactions of cell surface proteins (receptors) and their ligands. Finally, we take this discussion one step forward to include interactions between organisms and species to understand the generation of biological complexity. Development of technologies for accurate detection of protein-protein interactions may shed light on processes that go from the fine-tuning of pathways and metabolic networks to the emergence of biological complexity. PMID:21488148

  9. Protein Attachment on Nanodiamonds.

    PubMed

    Lin, Chung-Lun; Lin, Cheng-Huang; Chang, Huan-Cheng; Su, Meng-Chih

    2015-07-16

    A recent advance in nanotechnology is the scale-up production of small and nonaggregated diamond nanoparticles suitable for biological applications. Using detonation nanodiamonds (NDs) with an average diameter of ?4 nm as the adsorbents, we have studied the static attachment of three proteins (myoglobin, bovine serum albumin, and insulin) onto the nanoparticles by optical spectroscopy, mass spectrometry, and dynamic light scattering, and electrophoretic zeta potential measurements. Results show that the protein surface coverage is predominantly determined by the competition between protein-protein and protein-ND interactions, giving each protein a unique and characteristic structural configuration in its own complex. Specifically, both myoglobin and bovine serum albumin show a Langmuir-type adsorption behavior, forming 1:1 complexes at saturation, whereas insulin folds into a tightly bound multimer before adsorption. The markedly different adsorption patterns appear to be independent of the protein concentration and are closely related to the affinity of the individual proteins for the NDs. The present study provides a fundamental understanding for the use of NDs as a platform for nanomedical drug delivery. PMID:25815400

  10. Palindromes in proteins.

    PubMed

    Giel-Pietraszuk, Malgorzata; Hoffmann, Marcin; Dolecka, Sylwia; Rychlewski, Jacek; Barciszewski, Jan

    2003-02-01

    Palindromes in DNA consist of nucleotides sequences that read the same from the 5'-end to the 3'-end, and its double helix is related by twofold axis. They occur in genomes of all organisms and have various functions. For example, restriction enzymes often recognize palindromic sequences of DNA. Palindromes in telomeres are crucial for initiation of replication. One can ask the questions, Do palindromes occur in protein, and if so, what function they play? We have searched the protein SWISSPROT database for palindromic sequences. A great number (26%) of different protein palindromes were found. One example of such protein is systemin, an 18-amino-acid-long peptide. It contains palindrome in its beta-sheet domain that interacts with palindromic fragment of DNA. The other palindrome containing protein is cellular human tumor suppressor p53. Oligonucleotide LTI-ITL has been observed in the crystal structure and is located close to a DNA recognizing domain. As the number of possible palindromic sequences of a given length is far much greater for proteins (20N) than for nucleic acids (4N), the study on their role seems to be an exciting challenge. Our results have clearly showed that palindromes are frequently occurring motives in proteins. Moreover, even very few examples that we have examined so far indicate the importance of further studies on protein palindromes. PMID:12760415

  11. Mining Dense Overlapping Subgraphs in weighted protein-protein interaction networks.

    PubMed

    Lee, Anthony J T; Lin, Ming-Chih; Hsu, Chia-Ming

    2011-03-01

    Many methods have been proposed for mining protein complexes from a protein-protein interaction network; however, most of them focus on unweighted networks and cannot find overlapping protein complexes. Since one protein may serve different roles within different functional groups, mining overlapping protein complexes in a weighted protein-protein interaction network has attracted more and more attention recently. In this paper, we propose an effective method, called MDOS (Mining Dense Overlapping Subgraphs), for mining dense overlapping protein complexes (subgraphs) in a weighted protein-protein interaction network. The proposed method can integrate the information about known complexes into a weighted protein-protein interaction network to improve the mining results. The experiment results show that our method mines more known complexes and has higher sensitivity and accuracy than the CODENSE and MCL methods. PMID:21095218

  12. Protein Peeling 2: a web server to convert protein structures into series of protein units.

    PubMed

    Gelly, J-C; Etchebest, C; Hazout, S; de Brevern, A G

    2006-07-01

    Protein Peeling 2 (PP2) is a web server for the automatic identification of protein units (PUs) given the 3D coordinates of a protein. PUs are an intermediate level of protein structure description between protein domains and secondary structures. It is a new tool to better understand and analyze the organization of protein structures. PP2 uses only the matrices of protein contact probabilities and cuts the protein structures optimally using Matthews' coefficient correlation. An index assesses the compactness quality of each PU. Results are given both textually and graphically using JMol and PyMol softwares. The server can be accessed from http://www.ebgm.jussieu.fr/~gelly/index.html. PMID:16845113

  13. How a Cell Deals with Abnormal Proteins

    Microsoft Academic Search

    A. Aigelsreiter; E. Janig; C. Stumptner; A. Fuchsbichler; K. Zatloukal; H. Denk

    2007-01-01

    Defective protein folding is responsible for many diseases. Although these diseases seem to be quite diverse at the first glance, there is evidence for common pathogenetic principles. The basis of the pathological changes is the cell’s inability to prevent protein misfolding, to revert misfolded proteins to normal or to eliminate misfolded proteins by degradation. This could result in deposition of

  14. EDITORIAL: Precision proteins Precision proteins

    NASA Astrophysics Data System (ADS)

    Demming, Anna

    2010-06-01

    Since the birth of modern day medicine, during the times of Hippocrates in ancient Greece, the profession has developed from the rudimentary classification of disease into a rigorous science with an inspiring capability to treat and cure. Scientific methodology has distilled clinical diagnostic tools from the early arts of prognosis, which used to rely as much on revelation and prophecy, as intuition and judgement [1]. Over the past decade, research into the interactions between proteins and nanosystems has provided some ingenious and apt techniques for delving into the intricacies of anatomical systems. In vivo biosensing has emerged as a vibrant field of research, as much of medical diagnosis relies on the detection of substances or an imbalance in the chemicals in the body. The inherent properties of nanoscale structures, such as cantilevers, make them well suited to biosensing applications that demand the detection of molecules at very low concentrations. Measurable deflections in cantilevers functionalised with antibodies provide quantitative indicators of the presence of specific antigens when the two react. Such developments have roused mounting interest in the interactions of proteins with nanostructures, such as carbon nanotubes [3], which have demonstrated great potential as generic biomarkers. Plasmonic properties are also being exploited in sensing applications, such as the molecular sentinel recently devised by researchers in the US. The device uses the plasmonic properties of a silver nanoparticle linked to a Raman labelled hairpin DNA probe to signal changes in the probe geometry resulting from interactions with substances in the environment. Success stories so far include the detection of two specific genes associated with breast cancer [4]. A greater understanding of how RNA interference regulates gene expression has highlighted the potential of using this natural process as another agent for combating disease in personalized medicine. However, the large molecular weight, net negative charge and hydrophilicity of synthetic small interfering RNAs makes it hard for the molecules to cross the plasma membrane and enter the cell cytoplasm. Immune responses can also diminish the effectiveness of this approach. In this issue, Shiri Weinstein and Dan Peer from Tel Aviv University provide an overview of the challenges and recent progress in the use of nanocarriers for delivering RNAi effector molecules into target tissues and cells more effectively [5]. Also in this issue, researchers in Korea report new results that demonstrate the potential of nanostructures in neural network engineering [6]. Min Jee Jang et al report directional growth of neurites along linear carbon nanotube patterns, demonstrating great progress in neural engineering and the scope for using nanotechnology to treat neural diseases. Modern medicine cannot claim to have abolished the pain and suffering that accompany disease. But a comparison between the ghastly and often ineffective iron implements of early medicine and the smart gadgets and treatments used in hospitals today speaks volumes for the extraordinary progress that has been made, and the motivation behind this research. References [1] Wallis F 2000 Signs and senses: diagnosis and prognosis in early medieval pulse and urine texts Soc. Hist. Med. 13 265-78 [2] Arntz Y, Seelig J D, Lang H P, Zhang J, Hunziker P, Ramseyer J P, Meyer E, Hegner M and Gerber Ch 2003 Label-free protein assay based on a nanomechanical cantiliever array Nanotechnology 14 86-90 [3] Gowtham S, Scheicher R H, Pandey R, Karna S P and Ahuja R 2008 First-principles study of physisorption of nucleic acid bases on small-diameter carbon nanotubes Nanotechnology 19 125701 [4] Wang H-N and Vo-Dinh T 2009 Multiplex detection of breast cancer biomarkers using plasmonic molecular sentinel nanoprobes Nanotechnology 20 065101 [5] Weinstein S and Peer D 2010 RNAi nanomedicines: challenges and opportunities within the immune system Nanotechnology 21 232001 [6] Jang M J, Namgung S, Hong S, and Nam Y 2010 Directional neurite gro

  15. Protein Phosphatases

    NSDL National Science Digital Library

    Stephen R. Salton (Mount Sinai School of Medicine; Department of Neuroscience REV)

    2005-03-01

    This Teaching Resource provides lecture notes and slides for a class covering the structure and function of protein phosphatases and is part of the course "Cell Signaling Systems: A Course for Graduate Students." The lecture begins with a discussion of the importance of phosphatases in physiology, recognized by the award of a Nobel Prize in 1992, and then proceeds to describe the two types of protein phosphatases: serine/threonine and tyrosine phosphatases. The information covered includes the structure, regulation, and substrate specificity of protein phosphatases, with an emphasis on their importance in disease and clinical settings.

  16. Evolution of Chloroplast J Proteins

    PubMed Central

    Chiu, Chi-Chou; Chen, Lih-Jen; Su, Pai-Hsiang; Li, Hsou-min

    2013-01-01

    Hsp70 chaperones are involved in multiple biological processes and are recruited to specific processes by designated J domain-containing cochaperones, or J proteins. To understand the evolution and functions of chloroplast Hsp70s and J proteins, we identified the Arabidopsis chloroplast J protein constituency using a combination of genomic and proteomic database searches and individual protein import assays. We show that Arabidopsis chloroplasts have at least 19 J proteins, the highest number of confirmed J proteins for any organelle. These 19 J proteins are classified into 11 clades, for which cyanobacteria and glaucophytes only have homologs for one clade, green algae have an additional three clades, and all the other 7 clades are specific to land plants. Each clade also possesses a clade-specific novel motif that is likely used to interact with different client proteins. Gene expression analyses indicate that most land plant-specific J proteins show highly variable expression in different tissues and are down regulated by low temperatures. These results show that duplication of chloroplast Hsp70 in land plants is accompanied by more than doubling of the number of its J protein cochaperones through adding new J proteins with novel motifs, not through duplications within existing families. These new J proteins likely recruit chloroplast Hsp70 to perform tissue specific functions related to biosynthesis rather than to stress resistance. PMID:23894646

  17. Protein Purification

    NSDL National Science Digital Library

    2011-12-19

    This animation produced by WGBH and Digizyme, Inc. demonstrates how a protein of interest is isolated from other contents in a transformed bacterial cell—a process called purification—using a lab technique called hydrophobic interaction chromatography.

  18. Altering Protein Structure & Function Through Protein Engineering

    E-print Network

    Taralp, Alpay

    Altering Protein Structure & Function Through Protein Engineering Alpay Taralp, Sabanci University, Istanbul taralp@sabanciuniv.edu © 2006, Alpay Taralp, Sabanci University #12;Protein engineering has been reaction parameters, etc. Q: What can we accomplish by using protein engineering? improve existing

  19. The comparative genomics of protein interactions.

    PubMed

    Peregrín-Alvarez, José M; Ouzounis, Christos A

    2007-01-01

    The detection of gene fusion events across genomes can be used for the prediction of functional associations of proteins, including physical interactions or complex formation. These predictions are obtained by the detection of similarity for pairs of 'component' proteins to 'composite' proteins. Since the amount of composite proteins is limited in nature, we augment this set by creating artificial fusion proteins from experimentally determined protein interacting pairs. The goal is to study the extent of protein interaction partners with increasing phylogenetic distance, using an automated method. We have thus detected component pairs within seven entire genome sequences of similar size, using artificially generated composite proteins that have been shown to interact experimentally. Our results indicate that protein interactions are not conserved over large phylogenetic distances. In addition, we provide a set of predictions for functionally associated proteins across seven species using experimental information and demonstrate the applicability of fusion analysis for the comparative genomics of protein interactions. PMID:18546511

  20. PROTEIN DESIGN: Proteins from Scratch

    NSDL National Science Digital Library

    William F. DeGrado (University of Pennsylvania School of Medicine; Department of Biochemistry and Biophysics)

    1997-10-03

    Access to the article is free, however registration and sign-in are required. It is not easy to predict a protein's structure from its linear amino acid sequence, and the reverse problem--designing a protein that will assume a particular folded shape--is even harder. But on page 82 in this issue, Dahiyat and Mayo have succeeded in creating a zinc finger fold de novo, which forms the same shaped protein as the natural one but without the usual stabilizing zinc ion. In his Perspective, DeGrado explains why this problem has been so intractable and where we can go now that Dahiyat and Mayo have paved the way.

  1. RESEARCH ARTICLE Proteolipid Protein Cannot Replace P0

    E-print Network

    Lawrence Wrabetz,2 and Bruce D. Trapp1 The central nervous system (CNS) of terrestrial vertebrates membrane that surrounds many axons in the central (CNS) and peripheral nervous systems (PNS). The major of Peripheral Nervous System Myelin Xinghua Yin,1 Sumiko Kiryu-Seo,1 Grahame J. Kidd,1 M. Laura Feltri,2

  2. Protein constituents of the eggshell: eggshell-specific matrix proteins

    Microsoft Academic Search

    Megan L. H. Rose; Maxwell T. Hincke

    2009-01-01

    In this article, we review the results of recent proteomic and genomic analyses of eggshell matrix proteins and draw attention\\u000a to the impact of these data on current understanding of eggshell formation and function. Eggshell-specific matrix proteins\\u000a from avian (ovocleidins and ovocalyxins) and non-avian (paleovaterin) shells are discussed. Two possible roles for eggshell-specific\\u000a matrix proteins have been proposed; both reflect

  3. Envelope gene of the Friend spleen focus-forming virus: deletion and insertions in 3' gp70/p15E-encoding region have resulted in unique features in the primary structure of its protein product.

    PubMed

    Wolff, L; Scolnick, E; Ruscetti, S

    1983-08-01

    A nucleotide sequence was determined for the envelope (env) gene of the polycythemia-inducing strain of the acute leukemia-inducing Friend spleen focus-forming virus (SFFV) and from this the amino acid sequence of its gene product, gp52, was deduced. All major elements of the gene were found to be related to genes of other retroviruses that code for functional glycoproteins. Although the carboxyl terminus of gp52 is encoded by sequences highly related to sequences in its putative parent, ecotropic Friend murine leukemia virus, the majority of the protein (69%), including the amino terminus, is encoded by dualtropic virus-like sequences. Nucleotide sequence comparisons suggest that the nonecotropic region may be more closely related to the 5' substitution in dualtropic mink cell focus-inducing viruses that it is to the 5' end of xenotropic virus env genes. A large deletion and two unique insertions have been located in the env gene of polycythemia-inducing SFFV and may account for some of the unusual structural characteristics, aberrant processing, and pathogenic properties of gp52. As a consequence of the deletion, amino-terminal gp70 and carboxyl-terminal p15E-encoding sequences are juxtaposed and it appears that translation from the p15E region, 3' to the deletion, continues in the standard reading frame used by other retroviruses. Insertions of six base pairs and one base pair at the very 3' end of the gp52-encoding region results in a SFFV-unique amino acid sequence and a premature termination codon. PMID:6308646

  4. Circulating concentrations of insulin-like growth factor-I, insulin-like growth factor binding protein-3, genetic polymorphisms and mammographic density in premenopausal Mexican women: results from the ESMaestras cohort

    PubMed Central

    Rinaldi, S.; Biessy, C.; Hernandez, M; Lesueur, F.; dos-Santos-Silva, I.; Rice, M. S.; Lajous, M.; Lopez-Ridaura, R.; Torres-Mejía, G.; Romieu, I.

    2015-01-01

    The insulin-like growth factor (IGF) axis plays an essential role in the development of the mammary gland. High circulating levels of IGF-I and of its major binding protein IGFBP3 have been related with increased mammographic density in Caucasian premenopausal women. Some common single nucleotide polymorphisms (SNPs) in genes of the IGF pathway have also been suggested to play a role in mammographic density. We conducted a cross-sectional study nested within the large Mexican ESMaestras cohort, to investigate the relation between circulating levels of IGF-I, IGFBP-3, the IGF-I/IGFBP-3 ratio, five common SNPs in the IGF-1, IGFBP-3 and IGF-1R genes, and mammographic density in 593 premenopausal Mexican women. Mean age at mammogram was 43.1 (standard deviation–SD=3.7) years, and average body mass index (BMI) at recruitment was 28.5 kg/m2. Mean percent mammographic density was 36.5% (SD: 17.1), with mean dense tissue area of 48.3 (SD: 33.3) cm2. Mean IGF-I and IGFBP-3 concentrations were 15.33 (SD: 5.52) nmol/l and 114.96 (SD: 21.34) nmol/l, respectively. No significant associations were seen between percent density and biomarker concentrations but women with higher IGF-I and IGF-I/IGFBP-3 concentrations had lower absolute dense (ptrend =0.03 and 0.09, respectively) and non-dense tissue areas (ptrend <0.001 for both parameters). However, these associations were null after adjustment by BMI. SNPs in specific genes were associated with circulating levels of growth factors, but not with mammographic density features. These results do not support the hypothesis of a strong association between circulating levels of growth hormones and mammographic density in Mexican premenopausal women. PMID:24037648

  5. Young proteins experience more variable selection pressures than old proteins

    PubMed Central

    Vishnoi, Anchal; Kryazhimskiy, Sergey; Bazykin, Georgii A.; Hannenhalli, Sridhar; Plotkin, Joshua B.

    2010-01-01

    It is well known that young proteins tend to experience weaker purifying selection and evolve more quickly than old proteins. Here, we show that, in addition, young proteins tend to experience more variable selection pressures over time than old proteins. We demonstrate this pattern in three independent taxonomic groups: yeast, Drosophila, and mammals. The increased variability of selection pressures on young proteins is highly significant even after controlling for the fact that young proteins are typically shorter and experience weaker purifying selection than old proteins. The majority of our results are consistent with the hypothesis that the function of a young gene tends to change over time more readily than that of an old gene. At the same time, our results may be caused in part by young genes that serve constant functions over time, but nevertheless appear to evolve under changing selection pressures due to depletion of adaptive mutations. In either case, our results imply that the evolution of a protein-coding sequence is partly determined by its age and origin, and not only by the phenotypic properties of the encoded protein. We discuss, via specific examples, the consequences of these findings for understanding of the sources of evolutionary novelty. PMID:20921233

  6. Lmo7 is an emerin-binding protein that regulates the transcription of emerin and many other muscle-relevant genes.

    PubMed

    Holaska, James M; Rais-Bahrami, Soroush; Wilson, Katherine L

    2006-12-01

    X-linked Emery-Dreifuss muscular dystrophy (X-EDMD) is inherited through mutations in emerin, a nuclear membrane protein. Emerin has proposed roles in nuclear architecture and gene regulation, but direct molecular links to disease were unknown. We report that Lim-domain only 7 (Lmo7) binds emerin directly with 125 nM affinity; the C-terminal half of human Lmo7 (hLmo7C) was sufficient to bind emerin in vitro. Lmo7 appeared relevant to EDMD because a deletion that removes Lmo7 (plus eight exons of a neighboring gene) in mice causes dystrophic muscles [Semenova, E., Wang, X., Jablonski, M.M., Levorse, J. and Tilghman, S.M. (2003) An engineered 800 kilobase deletion of Uchl3 and Lmo7 on mouse chromosome 14 causes defects in viability, postnatal growth and degeneration of muscle and retina. Hum. Mol. Genet., 12, 1301-1312]. Lmo7 localizes in the nucleus, cytoplasm and cell surface, particularly adhesion junctions [Ooshio, T., Irie, K., Morimoto, K., Fukuhara, A., Imai, T. and Takai, Y. (2004) Involvement of LMO7 in the association of two cell-cell adhesion molecules, nectin and E-cadherin, through afadin and alpha-actinin in epithelial cells. J. Biol. Chem., 279, 31365-31373]. Our data suggest endogenous Lmo7 is a nucleocytoplasmic shuttling protein, and might also localize at focal adhesions in HeLa cells. Two key results show that Lmo7 regulates emerin gene expression: rat Lmo7 isoforms directly activated a luciferase reporter gene in vivo, and emerin mRNA expression decreased 93% in Lmo7-downregulated HeLa cells. Thus, Lmo7 not only binds emerin protein but is also required for emerin gene transcription. Microarray analysis of Lmo7-downregulated HeLa cells identified over 4200 misregulated genes, including 46 genes important for muscle or heart. Misregulation of 11 genes, including four (CREBBP, NAP1L1, LAP2, RBL2) known to be misregulated in X-EDMD patients and emerin-null mice [Bakay, M., Wang, Z., Melcon, G., Schiltz, L., Xuan, J., Zhao, P., Sartorelli, V., Seo, J., Pegoraro, E., Angelini, C. et al. (2006) Nuclear envelope dystrophies show a transcriptional fingerprint suggesting disruption of Rb-MyoD pathways in muscle regeneration. Brain, 129, 996-1013; Melcon, G., Kozlov, S., Cutler, D.A., Sullivan, T., Hernandez, L., Zhao, P., Mitchell, S., Nader, G., Bakay, M., Rottman, J.N. et al. (2006) Loss of emerin at the nuclear envelope disrupts the Rb1/E2F and MyoD pathways during muscle regeneration. Hum. Mol. Genet., 15, 637-651] was confirmed by real-time PCR. Overexpression of wild-type emerin, but not emerin mutant P183H (which causes EDMD and selectively disrupts binding to Lmo7), decreased the expression of CREBBP, NAP1L1 and LAP2, suggesting Lmo7 activity is both EDMD-relevant and inhibited by direct binding to emerin. We conclude that Lmo7 positively regulates many EDMD-relevant genes (including emerin), and is feedback-regulated by binding to emerin. PMID:17067998

  7. Cotton and Protein Interactions

    SciTech Connect

    Goheen, Steven C.; Edwards, J. V.; Rayburn, Alfred R.; Gaither, Kari A.; Castro, Nathan J.

    2006-06-30

    The adsorbent properties of important wound fluid proteins and cotton cellulose are reviewed. This review focuses on the adsorption of albumin to cotton-based wound dressings and some chemically modified derivatives targeted for chronic wounds. Adsorption of elastase in the presence of albumin was examined as a model to understand the interactive properties of these wound fluid components with cotton fibers. In the chronic non-healing wound, elastase appears to be over-expressed, and it digests tissue and growth factors, interfering with the normal healing process. Albumin is the most prevalent protein in wound fluid, and in highly to moderately exudative wounds, it may bind significantly to the fibers of wound dressings. Thus, the relative binding properties of both elastase and albumin to wound dressing fibers are of interest in the design of more effective wound dressings. The present work examines the binding of albumin to two different derivatives of cotton, and quantifies the elastase binding to the same derivatives following exposure of albumin to the fiber surface. An HPLC adsorption technique was employed coupled with a colorimetric enzyme assay to quantify the relative binding properties of albumin and elastase to cotton. The results of wound protein binding are discussed in relation to the porosity and surface chemistry interactions of cotton and wound proteins. Studies are directed to understanding the implications of protein adsorption phenomena in terms of fiber-protein models that have implications for rationally designing dressings for chronic wounds.

  8. Coexpression of the maize delta-zein and beta-zein genes results in stable accumulation of delta-zein in endoplasmic reticulum-derived protein bodies formed by beta-zein.

    PubMed Central

    Bagga, S; Adams, H P; Rodriguez, F D; Kemp, J D; Sengupta-Gopalan, C

    1997-01-01

    Zeins, the major seed storage proteins of maize, are of four distinct types: alpha, beta, delta, and gamma. They are synthesized on the rough endoplasmic reticulum (ER) in a sequential manner and deposited in ER-derived protein bodies. We investigated the potential for producing sulfur-rich beta-zein and delta-zein proteins in leaf and seed tissues by expressing the corresponding genes in a constitutive manner in transgenic tobacco. The delta-zein and beta-zein, when synthesized individually, were stable in the vegetative tissues and were deposited in unique, zein-specific ER-derived protein bodies. Coexpression of delta-zein and beta-zein genes, however, showed that delta-zein was colocalized in beta-zein-containing protein bodies and that the level of delta-zein was fivefold higher in delta-/beta-zein plants than in delta-zein plants. We conclude that delta-zein interacts with beta-zein and that the interaction has a stabilizing effect on delta-zein. PMID:9338969

  9. Integrating experimental and literature protein-protein interaction data for protein complex prediction

    PubMed Central

    2015-01-01

    Background Accurate determination of protein complexes is crucial for understanding cellular organization and function. High-throughput experimental techniques have generated a large amount of protein-protein interaction (PPI) data, allowing prediction of protein complexes from PPI networks. However, the high-throughput data often includes false positives and false negatives, making accurate prediction of protein complexes difficult. Method The biomedical literature contains large quantities of PPI data that, along with high-throughput experimental PPI data, are valuable for protein complex prediction. In this study, we employ a natural language processing technique to extract PPI data from the biomedical literature. This data is subsequently integrated with high-throughput PPI and gene ontology data by constructing attributed PPI networks, and a novel method for predicting protein complexes from the attributed PPI networks is proposed. This method allows calculation of the relative contribution of high-throughput and biomedical literature PPI data. Results Many well-characterized protein complexes are accurately predicted by this method when apply to two different yeast PPI datasets. The results show that (i) biomedical literature PPI data can effectively improve the performance of protein complex prediction; (ii) our method makes good use of high-throughput and biomedical literature PPI data along with gene ontology data to achieve state-of-the-art protein complex prediction capabilities. PMID:25708571

  10. Identification of hot spot residues at protein-protein interface.

    PubMed

    Li, Lei; Zhao, Bing; Cui, Zhanhua; Gan, Jacob; Sakharkar, Meena Kishore; Kangueane, Pandjassarame

    2006-01-01

    It is known that binding free energy of protein-protein interaction is mainly contributed by hot spot (high energy) interface residues. Here, we investigate the characteristics of hot spots by examining inter-atomic sidechain-sidechain interactions using a dataset of 296 alanine-mutated interface residues. Results show that hot spots participate in strong and energetically favorable sidechain-sidechain interactions. Subsequently, we describe a novel, yet simple 'hot spot' prediction model with an accuracy that is similar to many available approaches. The model is also shown to efficiently distinguish specific protein-protein interactions from non-specific interactions. PMID:17597870

  11. Identification of hot spot residues at protein-protein interface

    PubMed Central

    Li, Lei; Zhao, Bing; Cui, Zhanhua; Gan, Jacob; Sakharkar, Meena Kishore; Kangueane, Pandjassarame

    2006-01-01

    It is known that binding free energy of protein-protein interaction is mainly contributed by hot spot (high energy) interface residues. Here, we investigate the characteristics of hot spots by examining inter-atomic sidechain-sidechain interactions using a dataset of 296 alanine-mutated interface residues. Results show that hot spots participate in strong and energetically favorable sidechain-sidechain interactions. Subsequently, we describe a novel, yet simple ‘hot spot’ prediction model with an accuracy that is similar to many available approaches. The model is also shown to efficiently distinguish specific protein-protein interactions from non-specific interactions. PMID:17597870

  12. Conserved network motifs allow protein-protein interaction prediction

    Microsoft Academic Search

    Istvan Albert; Reka Albert

    2004-01-01

    Motivation: High-throughput protein interaction detection methods are strongly affected by false positive and false neg- ative results. Focused experiments are needed to complement the large-scale methods by validating previously detected interactions but it is often difficult to decide which proteins to probe as interaction partners. Developing reliable computa- tional methods assisting this decision process is a pressing need in bioinformatics.

  13. Impact of protein supplementation and care and support on body composition and CD4 count among HIV-infected women living in rural India: results from a randomized pilot clinical trial.

    PubMed

    Nyamathi, Adeline; Sinha, Sanjeev; Ganguly, Kalyan K; Ramakrishna, Padma; Suresh, P; Carpenter, Catherine L

    2013-07-01

    Body composition in HIV-infected individuals is subject to many influences. We conducted a pilot 6-month randomized trial of 68 women living with AIDS (WLA) from rural India. High protein intervention combined with education and supportive care delivered by HIV-trained village women (activated social health activist [Asha] life [AL]) was compared to standard protein with usual care delivered by village community assistants (usual care [UC]). Measurements included CD4 counts, ART adherence, socio-demographics, disease characteristics (questionnaires); and anthropometry (bioimpedance analyzer). Repeated measures analysis of variance modeled associations. AL significantly gained in BMI, muscle mass, fat mass, ART adherence, and CD4 counts compared to UC, with higher weight and muscle mass gains among ART adherent (?66%) participants who had healthier immunity (CD4 ?450). BMI of WLA improved through high protein supplementation combined with education and supportive care. Future research is needed to determine which intervention aspect was most responsible. PMID:23370835

  14. Non-linear fluorimetry of fluorescent proteins

    Microsoft Academic Search

    A. A. Banishev; E. A. Shirshin; V. V. Fadeev

    2007-01-01

    The results of development of the method of fluorescent proteins investigation are discussed. Photophysical parameters and concentrations of the chromophores of protein mRFP1 have been determined with use of non-linear fluorimetry.

  15. Ubiquitin domain proteins in disease

    PubMed Central

    Madsen, Louise; Schulze, Andrea; Seeger, Michael; Hartmann-Petersen, Rasmus

    2007-01-01

    The human genome encodes several ubiquitin-like (UBL) domain proteins (UDPs). Members of this protein family are involved in a variety of cellular functions and many are connected to the ubiquitin proteasome system, an essential pathway for protein degradation in eukaryotic cells. Despite their structural similarity, the UBL domains appear to have a range of different targets, resulting in a considerable diversity with respect to UDP function. Here, we give a short summary of the biochemical and physiological roles of the UDPs, which have been linked to human diseases including neurodegeneration and cancer. Publication history: Republished from Current BioData's Targeted Proteins database (TPdb; ). PMID:18047733

  16. Protein crystals and their growth

    Microsoft Academic Search

    Alexander A. Chernov

    2003-01-01

    Recent results on the associations between protein molecules in crystal lattices, crystal–solution surface energy, elastic properties, strength, and spontaneous crystal cracking are reviewed and discussed. In addition, some basic approaches to understanding the solubility of proteins are followed by an overview of crystal nucleation and growth. It is argued that variability of mixing in batch crystallization may be a source

  17. Increasing Stability Reduces Conformational Heterogeneity in a Protein Folding

    E-print Network

    Increasing Stability Reduces Conformational Heterogeneity in a Protein Folding Intermediate, the results show that protein folding intermediates are ensembles of different structural forms direct experi- mental evidence in support of a basic tenet of energy landscape theory for protein folding

  18. Is secondary wavelength always necessary in turbidimetric urine protein measurements?

    Microsoft Academic Search

    Fatma Meriç Y?lmaz; Gülsen Y?lmaz; Do?an Yücel

    2008-01-01

    ObjectivesTurbidimetric urine protein methods like Benzethonium Chloride (BTC) and Benzalkonium Chloride (BC) are previously reported to give falsely low protein results in very high protein concentrations (>2000 mg\\/L).

  19. Phylogenomics of Prokaryotic Ribosomal Proteins

    PubMed Central

    Yutin, Natalya; Puigbň, Pere; Koonin, Eugene V.; Wolf, Yuri I.

    2012-01-01

    Archaeal and bacterial ribosomes contain more than 50 proteins, including 34 that are universally conserved in the three domains of cellular life (bacteria, archaea, and eukaryotes). Despite the high sequence conservation, annotation of ribosomal (r-) protein genes is often difficult because of their short lengths and biased sequence composition. We developed an automated computational pipeline for identification of r-protein genes and applied it to 995 completely sequenced bacterial and 87 archaeal genomes available in the RefSeq database. The pipeline employs curated seed alignments of r-proteins to run position-specific scoring matrix (PSSM)-based BLAST searches against six-frame genome translations, mitigating possible gene annotation errors. As a result of this analysis, we performed a census of prokaryotic r-protein complements, enumerated missing and paralogous r-proteins, and analyzed the distributions of ribosomal protein genes among chromosomal partitions. Phyletic patterns of bacterial and archaeal r-protein genes were mapped to phylogenetic trees reconstructed from concatenated alignments of r-proteins to reveal the history of likely multiple independent gains and losses. These alignments, available for download, can be used as search profiles to improve genome annotation of r-proteins and for further comparative genomics studies. PMID:22615861

  20. Non-Arrhenius protein aggregation.

    PubMed

    Wang, Wei; Roberts, Christopher J

    2013-07-01

    Protein aggregation presents one of the key challenges in the development of protein biotherapeutics. It affects not only product quality but also potentially impacts safety, as protein aggregates have been shown to be linked with cytotoxicity and patient immunogenicity. Therefore, investigations of protein aggregation remain a major focus in pharmaceutical companies and academic institutions. Due to the complexity of the aggregation process and temperature-dependent conformational stability, temperature-induced protein aggregation is often non-Arrhenius over even relatively small temperature windows relevant for product development, and this makes low-temperature extrapolation difficult based simply on accelerated stability studies at high temperatures. This review discusses the non-Arrhenius nature of the temperature dependence of protein aggregation, explores possible causes, and considers inherent hurdles for accurately extrapolating aggregation rates from conventional industrial approaches for selecting accelerated conditions and from conventional or more advanced methods of analyzing the resulting rate data. PMID:23615748

  1. PROTEIN ENGINEERING KBK050 Protein Engineering

    E-print Network

    PROTEIN ENGINEERING KBK050 Protein Engineering Poäng: 5.0 Betygskala: TH Kursansvarig: Leif Bülow proteiners struktur och funktion. Innehĺll: I kursen behandlas hur proteiner kan muteras slumpmässigt och modifierat protein. Litteratur: Brown, T.A.: Gene Cloning, Chapman and Hall, 3rd ed. 1995; Brändén, C

  2. Construction of a cancer-perturbed protein-protein interaction network for discovery of apoptosis drug targets

    Microsoft Academic Search

    Liang-Hui Chu; Bor-Sen Chen

    2008-01-01

    BACKGROUND: Cancer is caused by genetic abnormalities, such as mutations of oncogenes or tumor suppressor genes, which alter downstream signal transduction pathways and protein-protein interactions. Comparisons of the interactions of proteins in cancerous and normal cells can shed light on the mechanisms of carcinogenesis. RESULTS: We constructed initial networks of protein-protein interactions involved in the apoptosis of cancerous and normal

  3. Outer membrane proteins of Escherichia coli. V. Evidence that protein 1 and bacteriophage-directed protein 2 are different polypeptides.

    PubMed Central

    Diedrich, D L; Summers, A O; Schnaitman, C A

    1977-01-01

    Protein 1 from the outer membrane of Escherichia coli K-12 and protein 2 from a phage PA-2 lysogen of the same strain were isolated by differential sodium dodecyl sulfate extraction and purified by ion-exchange and gel filtration chromatography. Rabbit antisera were prepared against these proteins and showed no cross-reaction between proteins 1 and 2. The proteins have the same N-terminal amino acid but show small yet significant differences in amino acid composition. The proteins were cleaved with cyanogenbromide in solvents containing both formic acid and trifluoroacetic acid. By comparing the cleavage in these solvents, it was established that protein 1 yielded 5 cyanogen bromide peptides, and the sum of the molecular weights of these was equivalent to the molecular weight of the uncleaved protein. Protein 2 yielded 4 cyanogen bromide peptides, none of which was identical to those of protein 1, and the sum of these peptides was also equivalent to the apparent molecular weight of the uncleaved protein. Significant differences were also observed when tryptic peptides from the two proteins were compared. These results indicate that protein 1 and the phage-directed protein 2 are distinct, different, and apparently homogeneous proteins. PMID:328488

  4. Water-protein interactions from high-resolution protein crystallography.

    PubMed Central

    Nakasako, Masayoshi

    2004-01-01

    To understand the role of water in life at molecular and atomic levels, structures and interactions at the protein-water interface have been investigated by cryogenic X-ray crystallography. The method enabled a much clearer visualization of definite hydration sites on the protein surface than at ambient temperature. Using the structural models of proteins, including several hydration water molecules, the characteristics in hydration structures were systematically analysed for the amount, the interaction geometries between water molecules and proteins, and the local and global distribution of water molecules on the surface of proteins. The tetrahedral hydrogen-bond geometry of water molecules in bulk solvent was retained at the interface and enabled the extension of a three-dimensional chain connection of a hydrogen-bond network among hydration water molecules and polar protein atoms over the entire surface of proteins. Networks of hydrogen bonds were quite flexible to accommodate and/or to regulate the conformational changes of proteins such as domain motions. The present experimental results may have profound implications in the understanding of the physico-chemical principles governing the dynamics of proteins in an aqueous environment and a discussion of why water is essential to life at a molecular level. PMID:15306376

  5. Novel computational methods to design protein-protein interactions

    NASA Astrophysics Data System (ADS)

    Zhou, Alice Qinhua; O'Hern, Corey; Regan, Lynne

    2014-03-01

    Despite the abundance of structural data, we still cannot accurately predict the structural and energetic changes resulting from mutations at protein interfaces. The inadequacy of current computational approaches to the analysis and design of protein-protein interactions has hampered the development of novel therapeutic and diagnostic agents. In this work, we apply a simple physical model that includes only a minimal set of geometrical constraints, excluded volume, and attractive van der Waals interactions to 1) rank the binding affinity of mutants of tetratricopeptide repeat proteins with their cognate peptides, 2) rank the energetics of binding of small designed proteins to the hydrophobic stem region of the influenza hemagglutinin protein, and 3) predict the stability of T4 lysozyme and staphylococcal nuclease mutants. This work will not only lead to a fundamental understanding of protein-protein interactions, but also to the development of efficient computational methods to rationally design protein interfaces with tunable specificity and affinity, and numerous applications in biomedicine. NSF DMR-1006537, PHY-1019147, Raymond and Beverly Sackler Institute for Biological, Physical and Engineering Sciences, and Howard Hughes Medical Institute.

  6. Functional centrality: detecting lethality of proteins in protein interaction networks.

    PubMed

    Tew, Kar Leong; Li, Xiao-Li; Tan, Soon-Heng

    2007-01-01

    Identifying lethal proteins is important for understanding the intricate mechanism governing life. Researchers have shown that the lethality of a protein can be computed based on its topological position in the protein-protein interaction (PPI) network. Performance of current approaches has been less than satisfactory as the lethality of a protein is a functional characteristic that cannot be determined solely by network topology. Furthermore, a significant number of lethal proteins have low connectivity in the interaction networks but are overlooked by most current methods. Our work reveals that a protein's lethality correlates more strongly with its "functional centrality" than pure topological centrality. We define functional centrality as the topological centrality within a subnetwork of proteins with similar functions. Evaluation experiments on four Saccharomyces cerevisiae PPI datasets showed that NFC performed significantly better than all the other existing computational techniques. Our method was able to detect low connectivity lethal proteins that were previously undetected by conventional methods. The results and an online version of NFC is available at http://lethalproteins.i2r.a-star.edu.sg. PMID:18546514

  7. Expression of foreign proteins in Escherichia coli by fusing with an archaeal FK506 binding protein

    Microsoft Academic Search

    A. Ideno; M. Furutani; T. Iwabuchi; T. Iida; Y. Iba; Y. Kurosawa; H. Sakuraba; T. Ohshima; Y. Kawarabayashi; T. Maruyama

    2004-01-01

    Improper protein-folding often results in inclusion-body formation in a protein expression system using Escherichia coli. To express such proteins in the soluble fraction of E. coli cytoplasm, we developed an expression system by fusing the target protein with an archaeal FK506 binding protein (FKBP). It has been reported that an archaeal FKBP from a hyperthermophilic archaeon, Thermococcus sp. KS-1 (TcFKBP18),

  8. Cutting Edge: Codeletion of the Ras GTPase-Activating Proteins (RasGAPs) Neurofibromin 1 and p120 RasGAP in T Cells Results in the Development of T Cell Acute Lymphoblastic Leukemia.

    PubMed

    Lubeck, Beth A; Lapinski, Philip E; Oliver, Jennifer A; Ksionda, Olga; Parada, Luis F; Zhu, Yuan; Maillard, Ivan; Chiang, Mark; Roose, Jeroen; King, Philip D

    2015-07-01

    Ras GTPase-activating proteins (RasGAPs) inhibit signal transduction initiated through the Ras small GTP-binding protein. However, which members of the RasGAP family act as negative regulators of T cell responses is not completely understood. In this study, we investigated potential roles for the RasGAPs RASA1 and neurofibromin 1 (NF1) in T cells through the generation and analysis of T cell-specific RASA1 and NF1 double-deficient mice. In contrast to mice lacking either RasGAP alone in T cells, double-deficient mice developed T cell acute lymphoblastic leukemia/lymphoma, which originated at an early point in T cell development and was dependent on activating mutations in the Notch1 gene. These findings highlight RASA1 and NF1 as cotumor suppressors in the T cell lineage. PMID:26002977

  9. Analysis of 3-nitrotyrosine in biological fluids and protein hydrolyzates by high-performance liquid chromatography using a postseparation, on-line reduction column and electrochemical detection: results with various nitrating agents.

    PubMed

    Ohshima, H; Celan, I; Chazotte, L; Pignatelli, B; Mower, H F

    1999-01-01

    Nitric oxide reacts rapidly with superoxide to form the strong nitrating agent peroxynitrite, which is responsible for much of the tissue damage associated with diverse pathophysiological conditions such as inflammation. The occurrence of free or protein-bound nitrotyrosine (NTYR) has been considered as evidence for in vivo formation of peroxynitrite. However, various agents can nitrate tyrosine, and their relative significance in vivo has not been determined due to lack of a sensitive method to analyze NTYR in tissue proteins and biological fluids. We have developed a new HPLC-electrochemical detection method to analyze NTYR in protein hydrolyzates or biological fluids. The sample is injected directly into a reversed-phase HPLC column and NTYR is subsequently reduced by a platinum column to 3-aminotyrosine, which is quantified with an electrochemical detector. The method is simple, selective, and sensitive (detection limit, 0.1 pmol per 20-microl injection). We have applied this method to compare in vitro the ability of various nitrating agents to form NTYR in bovine serum albumin and human plasma. Yields of NTYR formed in human plasma proteins incubated with 1 or 10 mM nitrating agent decreased in the following order: synthetic peroxynitrite > 3-morpholinosydonimine, a generator of both NO and superoxide > Angeli's salt, which forms nitroxyl anion (NO-) > spermine-NONOate, which releases NO > sodium nitrite plus hypochlorite, which forms the nitrating agent nitryl chloride (NO2Cl). A simple purification method using a C18 Sep-Pak cartridge is also described for analysis of free NTYR in human plasma. PMID:10369183

  10. Protein-Polymer Functionalized Nanopatterned Surfaces

    NASA Astrophysics Data System (ADS)

    Wang, Haoyu; Akcora, Pinar

    2015-03-01

    Understanding and controlling the protein interactions with surfaces for biosensors and biomedical implants is a fundamental problem for biocompatible nanomaterial design. Proteins attached in ordered nanopores can exhibit superior biological activities compared to smooth microstructured surfaces. We developed heterogeneous and nanopatterned surfaces decorated with polymer brushes and proteins to control protein fates through elasticity. The heterogeneity of surfaces is controlled with well-defined chemistry, pattern size and geometry, stiffness of polymers and protein types. We will present our recent nanoindentation results on nanopatterned and biofunctionalized flat surfaces and discuss the pattern size effect on protein activity, hence conformation.

  11. Engineering protein farnesyltransferase for enzymatic protein labeling applications.

    PubMed

    Dozier, Jonathan K; Khatwani, Santoshkumar L; Wollack, James W; Wang, Yen-Chih; Schmidt-Dannert, Claudia; Distefano, Mark D

    2014-07-16

    Creating covalent protein conjugates is an active area of research due to the wide range of uses for protein conjugates spanning everything from biological studies to protein therapeutics. Protein Farnesyltransferase (PFTase) has been used for the creation of site-specific protein conjugates, and a number of PFTase substrates have been developed to facilitate that work. PFTase is an effective catalyst for protein modification because it transfers Farnesyl diphosphate (FPP) analogues to protein substrates on a cysteine four residues from the C-terminus. While much work has been done to synthesize various FPP analogues, there are few reports investigating how mutations in PFTase alter the kinetics with these unnatural analogues. Herein we examined how different mutations within the PFTase active site alter the kinetics of the PFTase reaction with a series of large FPP analogues. We found that mutating either a single tryptophan or tyrosine residue to alanine results in greatly improved catalytic parameters, particularly in kcat. Mutation of tryptophan 102? to alanine caused a 4-fold increase in kcat and a 10-fold decrease in KM for a benzaldehyde-containing FPP analogue resulting in an overall 40-fold increase in catalytic efficiency. Similarly, mutation of tyrosine 205? to alanine caused a 25-fold increase in kcat and a 10-fold decrease in KM for a coumarin-containing analogue leading to a 300-fold increase in catalytic efficiency. Smaller but significant changes in catalytic parameters were also obtained for cyclo-octene- and NBD-containing FPP analogues. The latter compound was used to create a fluorescently labeled form of Ciliary Neurotrophic Factor (CNTF), a protein of therapeutic importance. Additionally, computational modeling was performed to study how the large non-natural isoprenoid analogues can fit into the active sites enlarged via mutagenesis. Overall, these results demonstrate that PFTase can be improved via mutagenesis in ways that will be useful for protein engineering and the creation of site-specific protein conjugates. PMID:24946229

  12. TGF-beta signaling proteins and the Protein Ontology

    PubMed Central

    Arighi, Cecilia N; Liu, Hongfang; Natale, Darren A; Barker, Winona C; Drabkin, Harold; Blake, Judith A; Smith, Barry; Wu, Cathy H

    2009-01-01

    Background The Protein Ontology (PRO) is designed as a formal and principled Open Biomedical Ontologies (OBO) Foundry ontology for proteins. The components of PRO extend from a classification of proteins on the basis of evolutionary relationships at the homeomorphic level to the representation of the multiple protein forms of a gene, including those resulting from alternative splicing, cleavage and/or post-translational modifications. Focusing specifically on the TGF-beta signaling proteins, we describe the building, curation, usage and dissemination of PRO. Results PRO is manually curated on the basis of PrePRO, an automatically generated file with content derived from standard protein data sources. Manual curation ensures that the treatment of the protein classes and the internal and external relationships conform to the PRO framework. The current release of PRO is based upon experimental data from mouse and human proteins wherein equivalent protein forms are represented by single terms. In addition to the PRO ontology, the annotation of PRO terms is released as a separate PRO association file, which contains, for each given PRO term, an annotation from the experimentally characterized sub-types as well as the corresponding database identifiers and sequence coordinates. The annotations are added in the form of relationship to other ontologies. Whenever possible, equivalent forms in other species are listed to facilitate cross-species comparison. Splice and allelic variants, gene fusion products and modified protein forms are all represented as entities in the ontology. Therefore, PRO provides for the representation of protein entities and a resource for describing the associated data. This makes PRO useful both for proteomics studies where isoforms and modified forms must be differentiated, and for studies of biological pathways, where representations need to take account of the different ways in which the cascade of events may depend on specific protein modifications. Conclusion PRO provides a framework for the formal representation of protein classes and protein forms in the OBO Foundry. It is designed to enable data retrieval and integration and machine reasoning at the molecular level of proteins, thereby facilitating cross-species comparisons, pathway analysis, disease modeling and the generation of new hypotheses. PMID:19426460

  13. A surprising simplicity to protein folding

    Microsoft Academic Search

    David Baker

    2000-01-01

    The polypeptide chains that make up proteins have thousands of atoms and hence millions of possible inter-atomic interactions. It might be supposed that the resulting complexity would make prediction of protein structure and protein-folding mechanisms nearly impossible. But the fundamental physics underlying folding may be much simpler than this complexity would lead us to expect: folding rates and mechanisms appear

  14. Kinetics versus Thermodynamics in Protein Folding

    Microsoft Academic Search

    David Baker; David A. Agard

    1994-01-01

    Until quite recently it has been generally believed that the observed tertiary structure of a protein is controlled by thermodynamic and not kinetic proceses. In this essay we review several recent results which call into question the universality of the thermodynamic hypothesis and discuss their implications for the understanding of protein folding. The thermodynamic hypothesis of protein folding has had

  15. SAMPI: Protein Identification with Mass Spectra Alignments

    Microsoft Academic Search

    Hans-michael Kaltenbach; Andreas Wilke; Sebastian Böcker

    2007-01-01

    Background: Mass spectrometry based peptide mass fingerprints (PMFs) offer a fast, efficient, and robust method for protein identification. A protein is digested (usually by trypsin) and its mass spectrum is compared to simulated spectra for protein sequences in a database. However, existing tools for analyzing PMFs often suffer from missing or heuristic analysis of the significance of search results and

  16. Investigating possible changes in protein structure during dendrimer-protein binding.

    PubMed

    Chiba, F; Mann, G; Twyman, L J

    2010-11-21

    Building on our previous results that revealed a sized based mechanism for dendrimer/protein binding, the mechanism of complexation is further probed using CD spectroscopy; the results demonstrate that dendrimer/protein binding is not accompanied by changes in the protein's structure and that binding takes place on the interfacial area/active site entrance. PMID:20862438

  17. Genome-wide Protein-protein Interaction Screening by Protein-fragment Complementation Assay (PCA) in Living Cells

    PubMed Central

    Filteau, Marie; Leducq, Jean-Baptiste; Dubé, Alexandre K.; Landry, Christian R.

    2015-01-01

    Proteins are the building blocks, effectors and signal mediators of cellular processes. A protein’s function, regulation and localization often depend on its interactions with other proteins. Here, we describe a protocol for the yeast protein-fragment complementation assay (PCA), a powerful method to detect direct and proximal associations between proteins in living cells. The interaction between two proteins, each fused to a dihydrofolate reductase (DHFR) protein fragment, translates into growth of yeast strains in presence of the drug methotrexate (MTX). Differential fitness, resulting from different amounts of reconstituted DHFR enzyme, can be quantified on high-density colony arrays, allowing to differentiate interacting from non-interacting bait-prey pairs. The high-throughput protocol presented here is performed using a robotic platform that parallelizes mating of bait and prey strains carrying complementary DHFR-fragment fusion proteins and the survival assay on MTX. This protocol allows to systematically test for thousands of protein-protein interactions (PPIs) involving bait proteins of interest and offers several advantages over other PPI detection assays, including the study of proteins expressed from their endogenous promoters without the need for modifying protein localization and for the assembly of complex reporter constructs. PMID:25867901

  18. Protein Design Zhilei Chen

    E-print Network

    Zhao, Huimin

    Protein Design Zhilei Chen Center for Biophysics and Computational Biology, University of Illinois of Illinois, Urbana, Illinois, U.S.A. INTRODUCTION Protein design refers to the ability to alter protein structure to achieve the desired protein function. Of the two classes of proteins (binding proteins

  19. Protein-protein interaction networks (PPI) and complex diseases.

    PubMed

    Safari-Alighiarloo, Nahid; Taghizadeh, Mohammad; Rezaei-Tavirani, Mostafa; Goliaei, Bahram; Peyvandi, Ali Asghar

    2014-01-01

    The physical interaction of proteins which lead to compiling them into large densely connected networks is a noticeable subject to investigation. Protein interaction networks are useful because of making basic scientific abstraction and improving biological and biomedical applications. Based on principle roles of proteins in biological function, their interactions determine molecular and cellular mechanisms, which control healthy and diseased states in organisms. Therefore, such networks facilitate the understanding of pathogenic (and physiologic) mechanisms that trigger the onset and progression of diseases. Consequently, this knowledge can be translated into effective diagnostic and therapeutic strategies. Furthermore, the results of several studies have proved that the structure and dynamics of protein networks are disturbed in complex diseases such as cancer and autoimmune disorders. Based on such relationship, a novel paradigm is suggested in order to confirm that the protein interaction networks can be the target of therapy for treatment of complex multi-genic diseases rather than individual molecules with disrespect the network. PMID:25436094

  20. Comparative effects of selenite and selenate on nitrate assimilation in barley seedlings

    NASA Technical Reports Server (NTRS)

    Aslam, M.; Harbit, K. B.; Huffaker, R. C.

    1990-01-01

    The effect of SeO3= and SeO4= on NO3- assimilation in 8-d-old barley (Hordeum vulgare L.) seedlings was studied over a 24-h period. Selenite at 0.1 mol m-3 in the uptake solutions severely inhibited the induction of NO3- uptake and active nitrate reductases. Selenate, at 1.0 mol m-3 in the nutrient solution, had little effect on induction of activities of these systems until after 12 h; however, when the seedlings were pretreated with 1.0 mol m-3 SeO4= for 24 h, subsequent NO3- uptake from SeO4(=) -free solutions was inhibited about 60%. Sulphate partially alleviated the inhibitory effect of SeO3= when supplied together in the ambient solutions, but had no effect in seedlings pretreated with SeO3=. By contrast, SO4= partially alleviated the inhibitory effect of SeO4= even in seedlings pretreated with SeO4=. Since uptake of NO3- by intact seedlings was also inhibited by SO3=, the percentage of the absorbed NO3- that was reduced was not affected. By contrast, SeO4=, which affected NO3- uptake much less, inhibited the percentage reduced of that absorbed. However, when supplied to detached leaves, both SeO3= and SeO4= inhibited the in vivo reduction of NO3- as well as induction of nitrate reductase and nitrite reductase activities. Selenite was more inhibitory than SeO4= ; approximately a five to 10 times higher concentration of SeO4= than SeO3= was required to achieve similar inhibition. In detached leaves, the inhibitory effect of both SeO3= and SeO4= on in vivo NO3- reduction as well as on the induction of nitrate reductase activity was partially alleviated by SO4=. The inhibitory effects of Se salts on the induction of the nitrite reductase were, however, completely alleviated by SO4=. The results show that in barley seedlings SeO3= is more toxic than SeO4=. The reduction of SeO4= to SeO3= may be a rate limiting step in causing Se toxicity.

  1. The Two-Hybrid System: A Method to Identify and Clone Genes for Proteins that Interact with a Protein of Interest

    Microsoft Academic Search

    Cheng-Ting Chien; Paul L. Bartel; Rolf Sternglanz

    1991-01-01

    We describe a method that detects proteins capable of interacting with a known protein and that results in the immediate availability of the cloned genes for these interacting proteins. Plasmids are constructed to encode two hybrid proteins. One hybrid consists of the DNA-binding domain of the yeast transcriptional activator protein GAL4 fused to the known protein; the other hybrid consists

  2. Protein Quality Control in the Nucleus

    PubMed Central

    Nielsen, Sofie V.; Poulsen, Esben G.; Rebula, Caio A.; Hartmann-Petersen, Rasmus

    2014-01-01

    In their natural environment, cells are regularly exposed to various stress conditions that may lead to protein misfolding, but also in the absence of stress, misfolded proteins occur as the result of mutations or failures during protein synthesis. Since such partially denatured proteins are prone to aggregate, cells have evolved several elaborate quality control systems to deal with these potentially toxic proteins. First, various molecular chaperones will seize the misfolded protein and either attempt to refold the protein or target it for degradation via the ubiquitin-proteasome system. The degradation of misfolded proteins is clearly compartmentalized, so unique degradation pathways exist for misfolded proteins depending on whether their subcellular localization is ER/secretory, mitochondrial, cytosolic or nuclear. Recent studies, mainly in yeast, have shown that the nucleus appears to be particularly active in protein quality control. Thus, specific ubiquitin-protein ligases located in the nucleus, target not only misfolded nuclear proteins, but also various misfolded cytosolic proteins which are transported to the nucleus prior to their degradation. In comparison, much less is known about these mechanisms in mammalian cells. Here we highlight recent advances in our understanding of nuclear protein quality control, in particular regarding substrate recognition and proteasomal degradation. PMID:25010148

  3. Exogenous amyloidogenic proteins function as seeds in amyloid ?-protein aggregation.

    PubMed

    Ono, Kenjiro; Takahashi, Ryoichi; Ikeda, Tokuhei; Mizuguchi, Mineyuki; Hamaguchi, Tsuyoshi; Yamada, Masahito

    2014-04-01

    Amyloid ?-protein (A?) aggregation is considered to be a critical step in the neurodegeneration of Alzheimer's disease (AD). In addition to A?, many proteins aggregate into the amyloid state, in which they form elongated fibers with spines comprising stranded ?-sheets. However, the cross-seeding effects of other protein aggregates on A? aggregation pathways are not completely clear. To investigate the cross-seeding effects of exogenous and human non-CNS amyloidogenic proteins on A? aggregation pathways, we examined whether and how sonicated fibrils of casein, fibroin, sericin, actin, and islet amyloid polypeptide affected A?40 and A?42 aggregation pathways using the thioflavin T assay and electron microscopy. Interestingly, the fibrillar seeds of all amyloidogenic proteins functioned as seeds. The cross-seeding effect of actin was stronger but that of fibroin was weaker than that of other proteins. Furthermore, our nuclear magnetic resonance spectroscopic studies identified the binding sites of A? with the amyloidogenic proteins. Our results indicate that the amyloidogenic proteins, including those contained in foods and cosmetics, contribute to A? aggregation by binding to A?, suggesting their possible roles in the propagation of A? amyloidosis. PMID:24440525

  4. Correlation of C-reactive protein haplotypes with serum C-reactive protein level and response to anti-tumor necrosis factor therapy in UK rheumatoid arthritis patients: results from the Biologics in Rheumatoid Arthritis Genetics and Genomics Study Syndicate cohort

    PubMed Central

    2012-01-01

    Introduction In many European countries, restrictions exist around the prescription of anti-tumor necrosis factor (anti-TNF) treatments for rheumatoid arthritis (RA). Eligibility and response to treatment is assessed by using the disease activity score 28 (DAS28) algorithm, which incorporates one of two inflammatory markers, erythrocyte sedimentation rate (ESR) or C-reactive protein (CRP). Although DAS28-CRP provides a more reliable measure of disease activity, functional variants exist within the CRP gene that affect basal CRP production. Therefore, we aimed to determine the relation between functional genetic variants at the CRP gene locus and levels of serum CRP in RA patients, and whether these variants, alone or in combination, are correlated with DAS28-CRP and change in DAS28-CRP after anti-TNF treatment. Methods DNA samples from the Biologics in Rheumatoid Arthritis Genetics and Genomics Study Syndicate (BRAGGSS) were genotyped for rs1205, rs1800947, and rs3091244 by using either TaqMan or the Sequenom MassARRAY iPLEX system. Estimated haplotypes were constructed for each sample by using the expectation maximization algorithm implemented in the haplo.stats package within the R statistical program. CRP values were log transformed, and the association between single nucleotide polymorphisms (SNPs), haplotypes of SNPs and baseline CRP, baseline DAS28-CRP, and change in DAS28-CRP were evaluated by using linear regression in STATA v.10. Results Baseline CRP measurements were available for 599 samples with 442 also having data 6 months after treatment with an anti-TNF. For these 442 samples, the study had > 80% power to detect a clinically meaningful difference of 0.6 DAS28 Units for an allele frequency of 5%. Estimated haplotype frequencies corresponded with previous frequencies reported in the literature. Overall, no significant association was observed between any of the markers investigated and baseline CRP levels. Further, CRP haplotypes did not correlate with baseline CRP (P = 0.593), baseline DAS28-CRP (P = 0.540), or change in DAS28-CRP after treatment with an anti-TNF over a 6-month period (P = 0.302). Conclusions Although CRP genotype may influence baseline CRP levels, in patients with very active disease, no such association was found. This suggests that genetic variation at the CRP locus does not influence DAS28-CRP, which may continue to be used in determining eligibility for and response to anti-TNF treatment, without adjusting for CRP genotype. PMID:23039402

  5. Computational approaches for detecting protein complexes from protein interaction networks: a survey

    PubMed Central

    2010-01-01

    Background Most proteins form macromolecular complexes to perform their biological functions. However, experimentally determined protein complex data, especially of those involving more than two protein partners, are relatively limited in the current state-of-the-art high-throughput experimental techniques. Nevertheless, many techniques (such as yeast-two-hybrid) have enabled systematic screening of pairwise protein-protein interactions en masse. Thus computational approaches for detecting protein complexes from protein interaction data are useful complements to the limited experimental methods. They can be used together with the experimental methods for mapping the interactions of proteins to understand how different proteins are organized into higher-level substructures to perform various cellular functions. Results Given the abundance of pairwise protein interaction data from high-throughput genome-wide experimental screenings, a protein interaction network can be constructed from protein interaction data by considering individual proteins as the nodes, and the existence of a physical interaction between a pair of proteins as a link. This binary protein interaction graph can then be used for detecting protein complexes using graph clustering techniques. In this paper, we review and evaluate the state-of-the-art techniques for computational detection of protein complexes, and discuss some promising research directions in this field. Conclusions Experimental results with yeast protein interaction data show that the interaction subgraphs discovered by various computational methods matched well with actual protein complexes. In addition, the computational approaches have also improved in performance over the years. Further improvements could be achieved if the quality of the underlying protein interaction data can be considered adequately to minimize the undesirable effects from the irrelevant and noisy sources, and the various biological evidences can be better incorporated into the detection process to maximize the exploitation of the increasing wealth of biological knowledge available. PMID:20158874

  6. Protein mediated membrane adhesion

    NASA Astrophysics Data System (ADS)

    Carlson, Andreas; Mahadevan, L.

    2015-05-01

    Adhesion in the context of mechanical attachment, signaling, and movement in cellular dynamics is mediated by the kinetic interactions between membrane-embedded proteins in an aqueous environment. Here, we present a minimal theoretical framework for the dynamics of membrane adhesion that accounts for the kinetics of protein binding, the elastic deformation of the membrane, and the hydrodynamics of squeeze flow in the membrane gap. We analyze the resulting equations using scaling estimates to characterize the spatiotemporal features of the adhesive patterning and corroborate them using numerical simulations. In addition to characterizing aspects of cellular dynamics, our results might also be applicable to a range of phenomena in physical chemistry and materials science where flow, deformation, and kinetics are coupled to each other in slender geometries.

  7. [ALR, the multifunctional protein].

    PubMed

    Balogh, Tibor; Szarka, András

    2015-03-29

    ALR is a mystic protein. It has a so called "long" 22 kDa and a "short" 15 kDa forms. It has been described after partial hepatectomy and it has just been considered as a key protein of liver regeneration. At the beginning of the 21st century it has been revealed that the "long" form is localized in the mitochondrial intermembrane space and it is an element of the mitochondrial protein import and disulphide relay system. Several proteins of the substrates of the mitochondrial disulphide relay system are necessary for the proper function of the mitochondria, thus any mutation of the ALR gene leads to mitochondrial diseases. The "short" form of ALR functions as a secreted extracellular growth factor and it promotes the protection, regeneration and proliferation of hepatocytes. The results gained on the recently generated conditional ALR mutant mice suggest that ALR can play an important role in the pathogenesis of alcoholic and non-alcoholic steatosis. Since the serum level of ALR is modified in several liver diseases it can be a promising marker molecule in laboratory diagnostics. PMID:25796277

  8. Water at interface with proteins

    E-print Network

    Giancarlo Franzese; Valentino Bianco; Svilen Iskrov

    2010-12-07

    Water is essential for the activity of proteins. However, the effect of the properties of water on the behavior of proteins is only partially understood. Recently, several experiments have investigated the relation between the dynamics of the hydration water and the dynamics of protein. These works have generated a large amount of data whose interpretation is debated. New experiments measure the dynamics of water at low temperature on the surface of proteins, finding a qualitative change (crossover) that might be related to the slowing down and stop of the protein's activity (protein glass transition), possibly relevant for the safe preservation of organic material at low temperature. To better understand the experimental data several scenarios have been discussed. Here, we review these experiments and discuss their interpretations in relation with the anomalous properties of water. We summarize the results for the thermodynamics and dynamics of supercooled water at an interface. We consider also the effect of water on protein stability, making a step in the direction of understanding, by means of Monte Carlo simulations and theoretical calculations, how the interplay of water cooperativity and hydrogen bonds interfacial strengthening affects the protein cold denaturation.

  9. Dual targeting of peroxisomal proteins

    PubMed Central

    Ast, Julia; Stiebler, Alina C.; Freitag, Johannes; Bölker, Michael

    2013-01-01

    Cellular compartmentalization into organelles serves to separate biological processes within the environment of a single cell. While some metabolic reactions are specific to a single organelle, others occur in more than one cellular compartment. Specific targeting of proteins to compartments inside of eukaryotic cells is mediated by defined sequence motifs. To achieve multiple targeting to different compartments cells use a variety of strategies. Here, we focus on mechanisms leading to dual targeting of peroxisomal proteins. In many instances, isoforms of peroxisomal proteins with distinct intracellular localization are encoded by separate genes. But also single genes can give rise to differentially localized proteins. Different isoforms can be generated by use of alternative transcriptional start sites, by differential splicing or ribosomal read-through of stop codons. In all these cases different peptide variants are produced, of which only one carries a peroxisomal targeting signal. Alternatively, peroxisomal proteins contain additional signals that compete for intracellular targeting. Dual localization of proteins residing in both the cytoplasm and in peroxisomes may also result from use of inefficient targeting signals. The recent observation that some bona fide cytoplasmic enzymes were also found in peroxisomes indicates that dual targeting of proteins to both the cytoplasm and the peroxisome might be more widespread. Although current knowledge of proteins exhibiting only partial peroxisomal targeting is far from being complete, we speculate that the metabolic capacity of peroxisomes might be larger than previously assumed. PMID:24151469

  10. Prediction of Heterodimeric Protein Complexes from Weighted Protein-Protein Interaction Networks Using Novel Features and Kernel Functions

    PubMed Central

    Ruan, Peiying; Hayashida, Morihiro; Maruyama, Osamu; Akutsu, Tatsuya

    2013-01-01

    Since many proteins express their functional activity by interacting with other proteins and forming protein complexes, it is very useful to identify sets of proteins that form complexes. For that purpose, many prediction methods for protein complexes from protein-protein interactions have been developed such as MCL, MCODE, RNSC, PCP, RRW, and NWE. These methods have dealt with only complexes with size of more than three because the methods often are based on some density of subgraphs. However, heterodimeric protein complexes that consist of two distinct proteins occupy a large part according to several comprehensive databases of known complexes. In this paper, we propose several feature space mappings from protein-protein interaction data, in which each interaction is weighted based on reliability. Furthermore, we make use of prior knowledge on protein domains to develop feature space mappings, domain composition kernel and its combination kernel with our proposed features. We perform ten-fold cross-validation computational experiments. These results suggest that our proposed kernel considerably outperforms the naive Bayes-based method, which is the best existing method for predicting heterodimeric protein complexes. PMID:23776458

  11. Discover protein sequence signatures from protein-protein interaction data

    E-print Network

    Fang, Jianwen; Haasl, R. J.; Dong, Yinghua; Lushington, Gerald H.

    2005-11-23

    Background: The development of high-throughput technologies such as yeast two- hybrid systems and mass spectrometry technologies has made it possible to generate large protein-protein interaction ( PPI) datasets. Mining ...

  12. Protein (western) blotting

    Microsoft Academic Search

    Denise Egger; Kurt Bienz

    1994-01-01

    The different steps involved in protein (Western) blotting and subsequent analysis of the proteins are reviewed. Electrophoretic\\u000a separation of proteins, procedures of transfer to membranes, immunological and nonimmunological protein detection systems,\\u000a and characterization of protein-nucleic acid and protein-protein interactions are described. Emphasis is on the sensitivity\\u000a of the methods described and on possible variations that allow the individual steps of

  13. Cobalt proteins.

    PubMed

    Kobayashi, M; Shimizu, S

    1999-04-01

    In the form of vitamin B12, cobalt plays a number of crucial roles in many biological functions. However, recent studies have provided information on the biochemistry and bioinorganic chemistry of several proteins containing cobalt in a form other than that in the corrin ring of vitamin B12. To date, eight noncorrin-cobalt-containing enzymes (methionine aminopeptidase, prolidase, nitrile hydratase, glucose isomerase, methylmalonyl-CoA carboxytransferase, aldehyde decarbonylase, lysine-2,3-aminomutase, and bromoperoxidase) have been isolated and characterized. A cobalt transporter is involved in the metallocenter biosynthesis of the host cobalt-containing enzyme, nitrile hydratase. Understanding the differences between cobalt and nickel transporters might lead to drug development for gastritis and peptic ulceration. PMID:10103026

  14. Main-chain complementarity in protein-protein recognition

    E-print Network

    Vakser, Ilya A.

    1996-09-01

    at the molecular surface. The structures were taken from known co-crystallized complexes. The intermolecular energy calculation was per- formed by a systematic 6-D search on a grid. The results revealed that in all cases tested (except antigen... of the general fold are important components in protein-protein recognition. In that study we demonstrated that the systematic grid search for possible binding modes of molecules, deprived of any structural details below the 7 A level, still retrieves most...

  15. Evolutionary optimization of protein folding.

    PubMed

    Debčs, Cédric; Wang, Minglei; Caetano-Anollés, Gustavo; Gräter, Frauke

    2013-01-01

    Nature has shaped the make up of proteins since their appearance, [Formula: see text]3.8 billion years ago. However, the fundamental drivers of structural change responsible for the extraordinary diversity of proteins have yet to be elucidated. Here we explore if protein evolution affects folding speed. We estimated folding times for the present-day catalog of protein domains directly from their size-modified contact order. These values were mapped onto an evolutionary timeline of domain appearance derived from a phylogenomic analysis of protein domains in 989 fully-sequenced genomes. Our results show a clear overall increase of folding speed during evolution, with known ultra-fast downhill folders appearing rather late in the timeline. Remarkably, folding optimization depends on secondary structure. While alpha-folds showed a tendency to fold faster throughout evolution, beta-folds exhibited a trend of folding time increase during the last [Formula: see text]1.5 billion years that began during the "big bang" of domain combinations. As a consequence, these domain structures are on average slow folders today. Our results suggest that fast and efficient folding of domains shaped the universe of protein structure. This finding supports the hypothesis that optimization of the kinetic and thermodynamic accessibility of the native fold reduces protein aggregation propensities that hamper cellular functions. PMID:23341762

  16. Intramesoporous silica structure differentiating protein loading density

    PubMed Central

    Qi, Wen; Li, Xiaolin; Chen, Baowei; Yao, Pei; Lei, Chenghong; Liu, Jun

    2012-01-01

    We report that hydrothermal aging temperature had a critical effect on intramesoporous structure of mesoporous silica and thus the intramesoporous structure affected protein loading in the mesoporous silica significantly. For a neutral protein Immunoglobulin G with a Y-like molecular shape, the larger desorption pore size allowed the larger protein loading. For a charged protein glucose oxidase with an elliptical molecular shape, the larger surface area resulted in the larger protein loading. Fluorescence emission spectra from tyrosinyl and tryptophanyl residues of the proteins in mesoporous silicas indicated that the charged protein was electrostatically attached inside the mesopores in a way of monolayer, while the neutral protein IgG could continue to aggregate after the monolayer occupancy. PMID:22745517

  17. Solubility of commercial milk protein concentrates and milk protein isolates.

    PubMed

    Sikand, V; Tong, P S; Roy, S; Rodriguez-Saona, L E; Murray, B A

    2011-12-01

    High-protein milk protein concentrate (MPC) and milk protein isolate (MPI) powders may have lower solubility than low-protein MPC powders, but information is limited on MPC solubility. Our objectives in this study were to (1) characterize the solubility of commercially available powder types with differing protein contents such as MPC40, MPC80, and MPI obtained from various manufacturers (sources), and (2) determine if such differences could be associated with differences in mineral, protein composition, and conformational changes of the powders. To examine possible predictors of solubility as measured by percent suspension stability (%SS), mineral analysis, Fourier transform infrared (FTIR) spectroscopy, and quantitative protein analysis by HPLC was performed. After accounting for overall differences between powder types, %SS was found to be strongly associated with the calcium, magnesium, phosphorus, and sodium content of the powders. The FTIR score plots were in agreement with %SS results. A principal component analysis of FTIR spectra clustered the highly soluble MPC40 separately from the rest of samples. Furthermore, 2 highly soluble MPI samples were clustered separately from the rest of the MPC80 and MPI samples. We found that the 900 to 1,200 cm?ą region exhibited the highest discriminating power, with dominant bands at 1,173 and 968 cm?ą, associated with phosphate vibrations. The 2 highly soluble MPI powders were observed to have lower ?-casein and ?-(S1)-casein contents and slightly higher whey protein contents than the other powders. The differences in the solubility of MPC and MPI were associated with a difference in mineral composition, which may be attributed to differences in processing conditions. Additional studies on the role of minerals composition on MPC80 solubility are warranted. Such a study would provide a greater understanding of factors associated with differences in solubility and can provide insight on methods to improve solubility of high-protein milk protein concentrates. PMID:22118108

  18. Graph pyramids for protein function prediction

    PubMed Central

    2015-01-01

    Background Uncovering the hidden organizational characteristics and regularities among biological sequences is the key issue for detailed understanding of an underlying biological phenomenon. Thus pattern recognition from nucleic acid sequences is an important affair for protein function prediction. As proteins from the same family exhibit similar characteristics, homology based approaches predict protein functions via protein classification. But conventional classification approaches mostly rely on the global features by considering only strong protein similarity matches. This leads to significant loss of prediction accuracy. Methods Here we construct the Protein-Protein Similarity (PPS) network, which captures the subtle properties of protein families. The proposed method considers the local as well as the global features, by examining the interactions among 'weakly interacting proteins' in the PPS network and by using hierarchical graph analysis via the graph pyramid. Different underlying properties of the protein families are uncovered by operating the proposed graph based features at various pyramid levels. Results Experimental results on benchmark data sets show that the proposed hierarchical voting algorithm using graph pyramid helps to improve computational efficiency as well the protein classification accuracy. Quantitatively, among 14,086 test sequences, on an average the proposed method misclassified only 21.1 sequences whereas baseline BLAST score based global feature matching method misclassified 362.9 sequences. With each correctly classified test sequence, the fast incremental learning ability of the proposed method further enhances the training model. Thus it has achieved more than 96% protein classification accuracy using only 20% per class training data. PMID:26044522

  19. An ontology-based search engine for protein-protein interactions

    PubMed Central

    2010-01-01

    Background Keyword matching or ID matching is the most common searching method in a large database of protein-protein interactions. They are purely syntactic methods, and retrieve the records in the database that contain a keyword or ID specified in a query. Such syntactic search methods often retrieve too few search results or no results despite many potential matches present in the database. Results We have developed a new method for representing protein-protein interactions and the Gene Ontology (GO) using modified Gödel numbers. This representation is hidden from users but enables a search engine using the representation to efficiently search protein-protein interactions in a biologically meaningful way. Given a query protein with optional search conditions expressed in one or more GO terms, the search engine finds all the interaction partners of the query protein by unique prime factorization of the modified Gödel numbers representing the query protein and the search conditions. Conclusion Representing the biological relations of proteins and their GO annotations by modified Gödel numbers makes a search engine efficiently find all protein-protein interactions by prime factorization of the numbers. Keyword matching or ID matching search methods often miss the interactions involving a protein that has no explicit annotations matching the search condition, but our search engine retrieves such interactions as well if they satisfy the search condition with a more specific term in the ontology. PMID:20122195

  20. G protein-coupled receptor accessory proteins and signaling: pharmacogenomic insights.

    PubMed

    Thompson, Miles D; Cole, David E C; Jose, Pedro A; Chidiac, Peter

    2014-01-01

    The identification and characterization of the genes encoding G protein-coupled receptors (GPCRs) and the proteins necessary for the processes of ligand binding, GPCR activation, inactivation, and receptor trafficking to the membrane are discussed in the context of human genetic disease. In addition to functional GPCR variants, the identification of genetic disruptions affecting proteins necessary to GPCR functions have provided insights into the function of these pathways. Gs? and G? subunit polymorphisms have been found to result in complex phenotypes. Disruptions in accessory proteins that normally modify or organize heterotrimeric G-protein coupling may also result in disease states. These include the contribution of variants of the regulator of G protein signaling (RGS) protein to hypertension; the role variants of the activator of G protein signaling (AGS) proteins to phenotypes (such as the type III AGS8 variant to hypoxia); the contribution of G protein-coupled receptor kinase (GRK) proteins, such as GRK4, in disorders such as hypertension. The role of accessory proteins in GPCR structure and function is discussed in the context of genetic disorders associated with disruption of the genes that encode them. An understanding of the pharmacogenomics of GPCR and accessory protein signaling provides the basis for examining both GPCR pharmacogenetics and the genetics of monogenic disorders that result from disruption of given receptor systems. PMID:25150869

  1. Protein Crystal Based Nanomaterials

    NASA Technical Reports Server (NTRS)

    Bell, Jeffrey A.; VanRoey, Patrick

    2001-01-01

    This is the final report on a NASA Grant. It concerns a description of work done, which includes: (1) Protein crystals cross-linked to form fibers; (2) Engineering of protein to favor crystallization; (3) Better knowledge-based potentials for protein-protein contacts; (4) Simulation of protein crystallization.

  2. Non-classical protein secretion in bacteria

    PubMed Central

    Bendtsen, Jannick D; Kiemer, Lars; Fausbřll, Anders; Brunak, Sřren

    2005-01-01

    Background We present an overview of bacterial non-classical secretion and a prediction method for identification of proteins following signal peptide independent secretion pathways. We have compiled a list of proteins found extracellularly despite the absence of a signal peptide. Some of these proteins also have known roles in the cytoplasm, which means they could be so-called "moon-lightning" proteins having more than one function. Results A thorough literature search was conducted to compile a list of currently known bacterial non-classically secreted proteins. Pattern finding methods were applied to the sequences in order to identify putative signal sequences or motifs responsible for their secretion. We have found no signal or motif characteristic to any majority of the proteins in the compiled list of non-classically secreted proteins, and conclude that these proteins, indeed, seem to be secreted in a novel fashion. However, we also show that the apparently non-classically secreted proteins are still distinguished from cellular proteins by properties such as amino acid composition, secondary structure and disordered regions. Specifically, prediction of disorder reveals that bacterial secretory proteins are more structurally disordered than their cytoplasmic counterparts. Finally, artificial neural networks were used to construct protein feature based methods for identification of non-classically secreted proteins in both Gram-positive and Gram-negative bacteria. Conclusion We present a publicly available prediction method capable of discriminating between this group of proteins and other proteins, thus allowing for the identification of novel non-classically secreted proteins. We suggest candidates for non-classically secreted proteins in Escherichia coli and Bacillus subtilis. The prediction method is available online. PMID:16212653

  3. Energetics-based Protein Profiling on a Proteomic Scale: Identification of Proteins Resistant to Proteolysis

    Microsoft Academic Search

    Sharleen Zhou; Jacqueline Gilmore; Susan Marqusee

    2007-01-01

    Native states of proteins are flexible, populating more than just the unique native conformation. The energetics and dynamics resulting from this conformational ensemble are inherently linked to protein function and regulation. Proteolytic susceptibility is one feature determined by this conformational energy landscape. As an attempt to investigate energetics of proteins on a proteomic scale, we challenged the Escherichia coli proteome

  4. Modulation of mitogen-activated protein kinase-activated protein kinase 3 by hepatitis C virus core protein.

    PubMed

    Ngo, Huong T T; Pham, Long V; Kim, Jong-Wook; Lim, Yun-Sook; Hwang, Soon B

    2013-05-01

    Hepatitis C virus (HCV) is highly dependent on cellular proteins for its own propagation. In order to identify the cellular factors involved in HCV propagation, we performed protein microarray assays using the HCV core protein as a probe. Of ~9,000 host proteins immobilized in a microarray, approximately 100 cellular proteins were identified as HCV core-interacting partners. Of these candidates, mitogen-activated protein kinase-activated protein kinase 3 (MAPKAPK3) was selected for further characterization. MAPKAPK3 is a serine/threonine protein kinase that is activated by stress and growth inducers. Binding of HCV core to MAPKAPK3 was confirmed by in vitro pulldown assay and further verified by coimmunoprecipitation assay. HCV core protein interacted with MAPKAPK3 through amino acid residues 41 to 75 of core and the N-terminal half of kinase domain of MAPKAPK3. In addition, both RNA and protein levels of MAPKAPK3 were elevated in both HCV subgenomic replicon cells and cell culture-derived HCV (HCVcc)-infected cells. Silencing of MAPKAPK3 expression resulted in decreases in both protein and HCV infectivity levels but not in the intracellular HCV RNA level. We showed that MAPKAPK3 increased HCV IRES-mediated translation and MAPKAPK3-dependent HCV IRES activity was further increased by core protein. These data suggest that HCV core may modulate MAPKAPK3 to facilitate its own propagation. PMID:23487458

  5. Discover Protein Complexes in Protein-Protein Interaction Networks Using Parametric Local Modularity

    Microsoft Academic Search

    Jongkwang Kim; Kai Tan

    2010-01-01

    BACKGROUND: Recent advances in proteomic technologies have enabled us to create detailed protein-protein interaction maps in multiple species and in both normal and diseased cells. As the size of the interaction dataset increases, powerful computational methods are required in order to effectively distil network models from large-scale interactome data. RESULTS: We present an algorithm, miPALM (Module Inference by Parametric Local

  6. Conformational search of Proteins and Protein Loops

    E-print Network

    Venkataramani, Ranjitha

    2008-02-26

    This thesis focuses on understanding the structure and dynamics of proteins using simulations. The main system of interest in our case was cytochrome b5, an electron transport protein, which exists in both Microsomal and Outer Mitochondrial isoforms...

  7. Evolutionary Monte Carlo for protein folding simulations Faming Lianga)

    E-print Network

    Liang, Faming

    Evolutionary Monte Carlo for protein folding simulations Faming Lianga) Department of Statistics to simulations of protein folding on simple lattice models, and to finding the ground state of a protein. In all structures in protein folding. The numerical results show that it is drastically superior to other methods

  8. CATH – a hierarchic classification of protein domain structures

    Microsoft Academic Search

    CA Orengo; AD Michie; S Jones; DT Jones; MB Swindells; JM Thornton

    1997-01-01

    Background: Protein evolution gives rise to families of structurally related proteins, within which sequence identities can be extremely low. As a result, structure-based classifications can be effective at identifying unanticipated relationships in known structures and in optimal cases function can also be assigned. The ever increasing number of known protein structures is too large to classify all proteins manually, therefore,

  9. Flow behavior of mixed-protein incipient gels

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Strong protein gel networks may result from synergistic interactions with other proteins or food materials above that achievable with a single protein alone. We determined varying flow and viscoelastic behavior of calcium caseinate (CC) or whey protein isolate (WPI) mixed with egg albumin (EA), fish...

  10. A fast indexing approach for protein structure comparison

    Microsoft Academic Search

    Lei Zhang; James Bailey; Arun S. Konagurthu; Kotagiri Ramamohanarao

    2010-01-01

    BACKGROUND: Protein structure comparison is a fundamental task in structural biology. While the number of known protein structures has grown rapidly over the last decade, searching a large database of protein structures is still relatively slow using existing methods. There is a need for new techniques which can rapidly compare protein structures, whilst maintaining high matching accuracy. RESULTS: We have

  11. Protein folding, protein homeostasis, and cancer

    PubMed Central

    Van Drie, John H.

    2011-01-01

    Proteins fold into their functional 3-dimensional structures from a linear amino acid sequence. In vitro this process is spontaneous; while in vivo it is orchestrated by a specialized set of proteins, called chaperones. Protein folding is an ongoing cellular process, as cellular proteins constantly undergo synthesis and degradation. Here emerging links between this process and cancer are reviewed. This perspective both yields insights into the current struggle to develop novel cancer chemotherapeutics and has implications for future chemotherapy discovery. PMID:21272445

  12. Encounter complexes and dimensionality reduction in protein–protein association

    PubMed Central

    Kozakov, Dima; Li, Keyong; Hall, David R; Beglov, Dmitri; Zheng, Jiefu; Vakili, Pirooz; Schueler-Furman, Ora; Paschalidis, Ioannis Ch; Clore, G Marius; Vajda, Sandor

    2014-01-01

    An outstanding challenge has been to understand the mechanism whereby proteins associate. We report here the results of exhaustively sampling the conformational space in protein–protein association using a physics-based energy function. The agreement between experimental intermolecular paramagnetic relaxation enhancement (PRE) data and the PRE profiles calculated from the docked structures shows that the method captures both specific and non-specific encounter complexes. To explore the energy landscape in the vicinity of the native structure, the nonlinear manifold describing the relative orientation of two solid bodies is projected onto a Euclidean space in which the shape of low energy regions is studied by principal component analysis. Results show that the energy surface is canyon-like, with a smooth funnel within a two dimensional subspace capturing over 75% of the total motion. Thus, proteins tend to associate along preferred pathways, similar to sliding of a protein along DNA in the process of protein-DNA recognition. DOI: http://dx.doi.org/10.7554/eLife.01370.001 PMID:24714491

  13. The t(10;11)(p13;q14) in the U937 cell line results in the fusion of the AF10 gene and CALM, encoding a new member of the AP-3 clathrin assembly protein family.

    PubMed Central

    Dreyling, M H; Martinez-Climent, J A; Zheng, M; Mao, J; Rowley, J D; Bohlander, S K

    1996-01-01

    The translocation t(10;11)(p13;q14) is a recurring chromosomal abnormality that has been observed in patients with acute lymphoblastic leukemia as well as acute myeloid leukemia. We have recently reported that the monocytic cell line U937 has a t(10;11)(p13;q14) translocation. Using a combination of positional cloning and candidate gene approach, we cloned the breakpoint and were able to show that AF10 is fused to a novel gene that we named CALM (Clathrin Assembly Lymphoid Myeloid leukemia gene) located at 11q14. AF10, a putative transcription factor, had recently been cloned as one of the fusion partners of MLL. CALM has a very high homology in its N-terminal third to the murine ap-3 gene which is one of the clathrin assembly proteins. The N-terminal region of ap-3 has been shown to bind to clathrin and to have a high-affinity binding site for phosphoinositols. The identification of the CALM/AF10 fusion gene in the widely used U937 cell line will contribute to our understanding of the malignant phenotype of this line. Images Fig. 1 Fig. 3 PMID:8643484

  14. Theoretical Perspectives on Protein Folding

    E-print Network

    D. Thirumalai; Edward P. O'Brien; Greg Morrison; Changbong Hyeon

    2010-07-18

    Understanding how monomeric proteins fold under in vitro conditions is crucial to describing their functions in the cellular context. Significant advances both in theory and experiments have resulted in a conceptual framework for describing the folding mechanisms of globular proteins. The experimental data and theoretical methods have revealed the multifaceted character of proteins. Proteins exhibit universal features that can be determined using only the number of amino acid residues (N) and polymer concepts. The sizes of proteins in the denatured and folded states, cooperativity of the folding transition, dispersions in the melting temperatures at the residue level, and time scales of folding are to a large extent determined by N. The consequences of finite N especially on how individual residues order upon folding depends on the topology of the folded states. Such intricate details can be predicted using the Molecular Transfer Model that combines simulations with measured transfer free energies of protein building blocks from water to the desired concentration of the denaturant. By watching one molecule fold at a time, using single molecule methods, the validity of the theoretically anticipated heterogeneity in the folding routes, and the N-dependent time scales for the three stages in the approach to the native state have been established. Despite the successes of theory, of which only a few examples are documented here, we conclude that much remains to be done to solve the "protein folding problem" in the broadest sense.

  15. Atomic force microscopy reveals the mechanical design of a modular protein

    NASA Astrophysics Data System (ADS)

    Li, Hongbin; Oberhauser, Andres F.; Fowler, Susan B.; Clarke, Jane; Fernandez, Julio M.

    2000-06-01

    Tandem modular proteins underlie the elasticity of natural adhesives, cell adhesion proteins, and muscle proteins. The fundamental unit of elastic proteins is their individually folded modules. Here, we use protein engineering to construct multimodular proteins composed of Ig modules of different mechanical strength. We examine the mechanical properties of the resulting tandem modular proteins by using single protein atomic force microscopy. We show that by combining modules of known mechanical strength, we can generate proteins with novel elastic properties. Our experiments reveal the simple mechanical design of modular proteins and open the way for the engineering of elastic proteins with defined mechanical properties, which can be used in tissue and fiber engineering.

  16. Querying Protein-Protein Interaction Networks

    Microsoft Academic Search

    Guillaume Blin; Florian Sikora; Stéphane Vialette

    2009-01-01

    Recent techniques increase the amount of our knowledge of interactions between proteins. To lter, interpret and organize this data, many authors have provided tools for querying patterns in the shape of paths or trees in Protein-Protein Interaction networks. In this paper, we propose an exact algorithm for querying graphs pattern based on dynamic programming and color-coding. We provide an implementation

  17. Amyloid Precursor Protein Binding Protein1 Modulates Cell Cycle Progression in Fetal Neural Stem Cells

    Microsoft Academic Search

    Yuyoung Joo; Sungji Ha; Bo-Hyun Hong; Jeong A. Kim; Keun-A. Chang; Hyunjeong Liew; Seonghan Kim; Woong Sun; Joung-Hun Kim; Young Hae Chong; Yoo-Hun Suh; Hye-Sun Kim; Brian Christie

    2010-01-01

    Amyloid precursor protein binding protein-1 (APP-BP1) binds to the carboxyl terminus of the amyloid precursor protein (APP) and serves as the bipartite activation enzyme for the ubiquitin-like protein, NEDD8. In the present study, we explored the physiological role of APP-BP1 in the cell cycle progression of fetal neural stem cells. Our results show that cell cycle progression of the cells

  18. Protein-Protein Interactions among Helicobacter pylori Cag Proteins

    PubMed Central

    Busler, Valerie J.; Torres, Victor J.; McClain, Mark S.; Tirado, Oscar; Friedman, David B.; Cover, Timothy L.

    2006-01-01

    Many Helicobacter pylori isolates contain a 40-kb region of chromosomal DNA known as the cag pathogenicity island (PAI). The risk for development of gastric cancer or peptic ulcer disease is higher among humans infected with cag PAI-positive H. pylori strains than among those infected with cag PAI-negative strains. The cag PAI encodes a type IV secretion system that translocates CagA into gastric epithelial cells. To identify Cag proteins that are expressed by H. pylori during growth in vitro, we compared the proteomes of a wild-type H. pylori strain and an isogenic cag PAI deletion mutant using two-dimensional difference gel electrophoresis (2D-DIGE) in multiple pH ranges. Seven Cag proteins were identified by this approach. We then used a yeast two-hybrid system to detect potential protein-protein interactions among 14 Cag proteins. One heterotypic interaction (CagY/7 with CagX/8) and two homotypic interactions (involving H. pylori VirB11/ATPase and Cag5) were similar to interactions previously reported to occur among homologous components of the Agrobacterium tumefaciens type IV secretion system. Other interactions involved Cag proteins that do not have known homologues in other bacterial species. Biochemical analysis confirmed selected interactions involving five of the proteins that were identified by 2D-DIGE. Protein-protein interactions among Cag proteins are likely to have an important role in the assembly of the H. pylori type IV secretion apparatus. PMID:16788188

  19. PDZ Domain Proteins: Plug and Play!

    NSDL National Science Digital Library

    Claire Nourry (Marseille; Laboratory of Molecular Pharmacology REV)

    2003-04-22

    PDZ domains (an acronym from PSD-95, Dlg, and ZO-1) are protein modules found in many cytoplasmic proteins (more than 400 in humans); they are discussed in this STKE Review. They are associated with a wide range of other protein-protein interaction domains (for example, WW, PTB, LRR, SH3) or with domains that exhibit particular enzymatic activities (such as guanosine triphosphatases, serine-threonine kinases, phosphatases), and they participate in various intracellular protein networks. PDZ domains bind to very diverse carboxyl-termini of protein partners in a specific (and sometimes reversible) manner, which enables the formation of supramolecular networks. Scaffolding of proteins by PDZ domain proteins usually occurs at specific sites within the cell (such as the plasma membrane or the Golgi apparatus) and is frequently involved in localizing proteins to specialized subcellular compartments of polarized cells, such as the presynaptic terminals and postsynaptic densities of neurons and the basolateral or apical membranes of epithelial cells. Genetic models in invertebrates and vertebrates that are now available for some PDZ proteins illuminate the large set of biological processes in which this protein family is involved, from the establishment and maintenance of the cytoarchitecture to signaling events. Accordingly, defects of PDZ proteins that play a central role in tissue homeostasis result in pathological conditions including cancer and developmental abnormalities. The simplicity of PDZ domain interactions has enabled the design of pharmacological inhibitors of potential therapeutic interest.

  20. Contribution of hydrogen bonds to protein stability

    PubMed Central

    Pace, C Nick; Fu, Hailong; Fryar, Katrina Lee; Landua, John; Trevino, Saul R; Schell, David; Thurlkill, Richard L; Imura, Satoshi; Scholtz, J Martin; Gajiwala, Ketan; Sevcik, Jozef; Urbanikova, Lubica; Myers, Jeffery K; Takano, Kazufumi; Hebert, Eric J; Shirley, Bret A; Grimsley, Gerald R

    2014-01-01

    Our goal was to gain a better understanding of the contribution of the burial of polar groups and their hydrogen bonds to the conformational stability of proteins. We measured the change in stability, ?(?G), for a series of hydrogen bonding mutants in four proteins: villin headpiece subdomain (VHP) containing 36 residues, a surface protein from Borrelia burgdorferi (VlsE) containing 341 residues, and two proteins previously studied in our laboratory, ribonucleases Sa (RNase Sa) and T1 (RNase T1). Crystal structures were determined for three of the hydrogen bonding mutants of RNase Sa: S24A, Y51F, and T95A. The structures are very similar to wild type RNase Sa and the hydrogen bonding partners form intermolecular hydrogen bonds to water in all three mutants. We compare our results with previous studies of similar mutants in other proteins and reach the following conclusions. (1) Hydrogen bonds contribute favorably to protein stability. (2) The contribution of hydrogen bonds to protein stability is strongly context dependent. (3) Hydrogen bonds by side chains and peptide groups make similar contributions to protein stability. (4) Polar group burial can make a favorable contribution to protein stability even if the polar groups are not hydrogen bonded. (5) The contribution of hydrogen bonds to protein stability is similar for VHP, a small protein, and VlsE, a large protein. PMID:24591301

  1. Structure-function relationship of viral coat proteins. A site directed spectroscopic study of M13 coat protein

    Microsoft Academic Search

    D. Stopar

    1997-01-01

    This thesis describes the results of a spectroscopic study of the major coat protein of bacteriophage M13. During the infection process this protein is incorporated into the cytoplasmic membrane of Escherichia coli host cells. To specifically monitor the local structural changes and changes in the environment of the protein upon membrane insertion, a set of cysteine site-specific mutants of protein

  2. Hexosamine pathway and (ER) protein quality control.

    PubMed

    Denzel, Martin S; Antebi, Adam

    2015-04-01

    Aminosugars produced in the hexosamine pathway (HP) are utilized in protein glycosylation reactions involved in protein maturation and cellular signaling. Recent evidence revealed a role of the HP in protein quality control and ageing. Elevation of the HP product UDP-N-acetylglucosamine in the nematode Caenorhabditis elegans results in resistance towards toxic aggregation-prone proteins, and extended lifespan. Glutamine-fructose 6 phosphate aminotransferase (GFAT-1), the HP's key enzyme, is a target of the unfolded protein response (UPR). Thus, cardiac stress in mice results in GFAT-1 activation that triggers a cytoprotective response. Feeding of glucosamine to aged mice increases their life expectancy. Here we discuss HP activation and cellular protein quality control mechanisms that result in stress resistance and suppression of age-related proteotoxicity. PMID:25463841

  3. Identification of Essential Proteins Based on a New Combination of Local Interaction Density and Protein Complexes

    PubMed Central

    Luo, Jiawei; Qi, Yi

    2015-01-01

    Background Computational approaches aided by computer science have been used to predict essential proteins and are faster than expensive, time-consuming, laborious experimental approaches. However, the performance of such approaches is still poor, making practical applications of computational approaches difficult in some fields. Hence, the development of more suitable and efficient computing methods is necessary for identification of essential proteins. Method In this paper, we propose a new method for predicting essential proteins in a protein interaction network, local interaction density combined with protein complexes (LIDC), based on statistical analyses of essential proteins and protein complexes. First, we introduce a new local topological centrality, local interaction density (LID), of the yeast PPI network; second, we discuss a new integration strategy for multiple bioinformatics. The LIDC method was then developed through a combination of LID and protein complex information based on our new integration strategy. The purpose of LIDC is discovery of important features of essential proteins with their neighbors in real protein complexes, thereby improving the efficiency of identification. Results Experimental results based on three different PPI(protein-protein interaction) networks of Saccharomyces cerevisiae and Escherichia coli showed that LIDC outperformed classical topological centrality measures and some recent combinational methods. Moreover, when predicting MIPS datasets, the better improvement of performance obtained by LIDC is over all nine reference methods (i.e., DC, BC, NC, LID, PeC, CoEWC, WDC, ION, and UC). Conclusions LIDC is more effective for the prediction of essential proteins than other recently developed methods. PMID:26125187

  4. PPLook: an automated data mining tool for protein-protein interaction

    PubMed Central

    2010-01-01

    Background Extracting and visualizing of protein-protein interaction (PPI) from text literatures are a meaningful topic in protein science. It assists the identification of interactions among proteins. There is a lack of tools to extract PPI, visualize and classify the results. Results We developed a PPI search system, termed PPLook, which automatically extracts and visualizes protein-protein interaction (PPI) from text. Given a query protein name, PPLook can search a dataset for other proteins interacting with it by using a keywords dictionary pattern-matching algorithm, and display the topological parameters, such as the number of nodes, edges, and connected components. The visualization component of PPLook enables us to view the interaction relationship among the proteins in a three-dimensional space based on the OpenGL graphics interface technology. PPLook can also provide the functions of selecting protein semantic class, counting the number of semantic class proteins which interact with query protein, counting the literature number of articles appearing the interaction relationship about the query protein. Moreover, PPLook provides heterogeneous search and a user-friendly graphical interface. Conclusions PPLook is an effective tool for biologists and biosystem developers who need to access PPI information from the literature. PPLook is freely available for non-commercial users at http://meta.usc.edu/softs/PPLook. PMID:20550717

  5. Spectral affinity in protein networks

    PubMed Central

    2009-01-01

    Background Protein-protein interaction (PPI) networks enable us to better understand the functional organization of the proteome. We can learn a lot about a particular protein by querying its neighborhood in a PPI network to find proteins with similar function. A spectral approach that considers random walks between nodes of interest is particularly useful in evaluating closeness in PPI networks. Spectral measures of closeness are more robust to noise in the data and are more precise than simpler methods based on edge density and shortest path length. Results We develop a novel affinity measure for pairs of proteins in PPI networks, which uses personalized PageRank, a random walk based method used in context-sensitive search on the Web. Our measure of closeness, which we call PageRank Affinity, is proportional to the number of times the smaller-degree protein is visited in a random walk that restarts at the larger-degree protein. PageRank considers paths of all lengths in a network, therefore PageRank Affinity is a precise measure that is robust to noise in the data. PageRank Affinity is also provably related to cluster co-membership, making it a meaningful measure. In our experiments on protein networks we find that our measure is better at predicting co-complex membership and finding functionally related proteins than other commonly used measures of closeness. Moreover, our experiments indicate that PageRank Affinity is very resilient to noise in the network. In addition, based on our method we build a tool that quickly finds nodes closest to a queried protein in any protein network, and easily scales to much larger biological networks. Conclusion We define a meaningful way to assess the closeness of two proteins in a PPI network, and show that our closeness measure is more biologically significant than other commonly used methods. We also develop a tool, accessible at http://xialab.bu.edu/resources/pnns, that allows the user to quickly find nodes closest to a queried vertex in any protein network available from BioGRID or specified by the user. PMID:19943959

  6. Protein surface-distribution and protein-protein interactions in the binding of peripheral proteins to charged lipid membranes.

    PubMed Central

    Heimburg, T; Marsh, D

    1995-01-01

    The binding of native cytochrome c to negatively charged lipid dispersions of dioleoyl phosphatidylglycerol has been studied over a wide range of ionic strengths. Not only is the strength of protein binding found to decrease rapidly with increasing ionic strength, but also the binding curves reach an apparent saturation level that decreases rapidly with increasing ionic strength. Analysis of the binding isotherms with a general statistical thermodynamic model that takes into account not only the free energy of the electrostatic double layer, but also the free energy of the surface distribution of the protein, demonstrates that the apparent saturation effects could arise from a competition between the out-of-plane binding reaction and the lateral in-plane interactions between proteins at the surface. It is found that association with nonlocalized sites results in binding isotherms that display the apparent saturation effect to a much more pronounced extent than does the Langmuir adsorption isotherm for binding to localized sites. With the model for nonlocalized sites, the binding isotherms of native cytochrome c can be described adequately by taking into account only the entropy of the surface distribution of the protein, without appreciable enthalpic interactions between the bound proteins. The binding of cytochrome c to dioleoyl phosphatidylglycerol dispersions at a temperature at which the bound protein is denatured on the lipid surface, but is nondenatured when free in solution, has also been studied. The binding curves for the surface-denatured protein differ from those for the native protein in that the apparent saturation at high ionic strength is less pronounced. This indicates the tendency of the denatured protein to aggregate on the lipid surface, and can be described by the binding isotherms for nonlocalized sites only if attractive interactions between the surface-bound proteins are included in addition to the distributional entropic terms. Additionally, it is found that the binding capacity for the native protein is increased at low ionic strength to a value that is greater than that for complete surface coverage, and that corresponds more closely to neutralization of the effective charge (determined from the ionic strength dependence), rather than of the total net charge, on the protein. Electron spin resonance experiments with spin-labeled lipids indicate that this different mode of binding arises from a penetration or disturbance of the bilayer surface by the protein that may alleviate the effects of in-plane interactions under conditions of strong binding. PMID:7696507

  7. A method for investigating protein-protein interactions related to Salmonella typhimurium pathogenesis

    SciTech Connect

    Chowdhury, Saiful M.; Shi, Liang; Yoon, Hyunjin; Ansong, Charles; Rommereim, Leah M.; Norbeck, Angela D.; Auberry, Kenneth J.; Moore, R. J.; Adkins, Joshua N.; Heffron, Fred; Smith, Richard D.

    2009-03-01

    We successfully modified an existing method to investigate protein-protein interactions in the pathogenic bacterium Salmonella typhimurium (STM). This method includes i) addition of a histidine-biotin-histidine tag to the bait proteins via recombinant DNA techniques; ii) in vivo cross-linking with formaldehyde; iii) tandem affinity purification of bait proteins under fully denaturing conditions; and iv) identification of the proteins cross-linked to the bait proteins by liquid-chromatography in conjunction with tandem mass-spectrometry. In vivo cross-linking stabilized protein interactions permitted the subsequent two-step purification step conducted under denaturing conditions. The two-step purification greatly reduced nonspecific binding of non-cross-linked proteins to bait proteins. Two different negative controls were employed to reduce false-positive identification. In an initial demonstration of this approach, we tagged three selected STM proteins? HimD, PduB and PhoP? with known binding partners that ranged from stable (e.g., HimD) to transient (i.e., PhoP). Distinct sets of interacting proteins were identified with each bait protein, including the known binding partners such as HimA for HimD, as well as anticipated and unexpected binding partners. Our results suggest that novel protein-protein interactions may be critical to pathogenesis by Salmonella typhimurium. .

  8. Towards the development of Bacillus subtilis as a cell factory for membrane proteins and protein complexes

    PubMed Central

    Zweers, Jessica C; Barák, Imrich; Becher, Dörte; Driessen, Arnold JM; Hecker, Michael; Kontinen, Vesa P; Saller, Manfred J; Vavrová, L'udmila; van Dijl, Jan Maarten

    2008-01-01

    Background The Gram-positive bacterium Bacillus subtilis is an important producer of high quality industrial enzymes and a few eukaryotic proteins. Most of these proteins are secreted into the growth medium, but successful examples of cytoplasmic protein production are also known. Therefore, one may anticipate that the high protein production potential of B. subtilis can be exploited for protein complexes and membrane proteins to facilitate their functional and structural analysis. The high quality of proteins produced with B. subtilis results from the action of cellular quality control systems that efficiently remove misfolded or incompletely synthesized proteins. Paradoxically, cellular quality control systems also represent bottlenecks for the production of various heterologous proteins at significant concentrations. Conclusion While inactivation of quality control systems has the potential to improve protein production yields, this could be achieved at the expense of product quality. Mechanisms underlying degradation of secretory proteins are nowadays well understood and often controllable. It will therefore be a major challenge for future research to identify and modulate quality control systems of B. subtilis that limit the production of high quality protein complexes and membrane proteins, and to enhance those systems that facilitate assembly of these proteins. PMID:18394159

  9. 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

  10. Origins of Protein Functions in Cells

    NASA Technical Reports Server (NTRS)

    Seelig, Burchard; Pohorille, Andrzej

    2011-01-01

    In modern organisms proteins perform a majority of cellular functions, such as chemical catalysis, energy transduction and transport of material across cell walls. Although great strides have been made towards understanding protein evolution, a meaningful extrapolation from contemporary proteins to their earliest ancestors is virtually impossible. In an alternative approach, the origin of water-soluble proteins was probed through the synthesis and in vitro evolution of very large libraries of random amino acid sequences. In combination with computer modeling and simulations, these experiments allow us to address a number of fundamental questions about the origins of proteins. Can functionality emerge from random sequences of proteins? How did the initial repertoire of functional proteins diversify to facilitate new functions? Did this diversification proceed primarily through drawing novel functionalities from random sequences or through evolution of already existing proto-enzymes? Did protein evolution start from a pool of proteins defined by a frozen accident and other collections of proteins could start a different evolutionary pathway? Although we do not have definitive answers to these questions yet, important clues have been uncovered. In one example (Keefe and Szostak, 2001), novel ATP binding proteins were identified that appear to be unrelated in both sequence and structure to any known ATP binding proteins. One of these proteins was subsequently redesigned computationally to bind GTP through introducing several mutations that introduce targeted structural changes to the protein, improve its binding to guanine and prevent water from accessing the active center. This study facilitates further investigations of individual evolutionary steps that lead to a change of function in primordial proteins. In a second study (Seelig and Szostak, 2007), novel enzymes were generated that can join two pieces of RNA in a reaction for which no natural enzymes are known. Recently it was found that, as in the previous case, the proteins have a structure unknown among modern enzymes. In this case, in vitro evolution started from a small, non-enzymatic protein. A similar selection process initiated from a library of random polypeptides is in progress. These results not only allow for estimating the occurrence of function in random protein assemblies but also provide evidence for the possibility of alternative protein worlds. Extant proteins might simply represent a frozen accident in the world of possible proteins. Alternative collections of proteins, even with similar functions, could originate alternative evolutionary paths.

  11. Macromolecular crowding and protein stability.

    PubMed

    Wang, Yaqiang; Sarkar, Mohona; Smith, Austin E; Krois, Alexander S; Pielak, Gary J

    2012-10-10

    An understanding of cellular chemistry requires knowledge of how crowded environments affect proteins. The influence of crowding on protein stability arises from two phenomena, hard-core repulsions and soft (i.e., chemical) interactions. Most efforts to understand crowding effects on protein stability, however, focus on hard-core repulsions, which are inherently entropic and stabilizing. We assessed these phenomena by measuring the temperature dependence of NMR-detected amide proton exchange and used these data to extract the entropic and enthalpic contributions of crowding to the stability of ubiquitin. Contrary to expectations, the contribution of chemical interactions is large and in many cases dominates the contribution from hardcore repulsions. Our results show that both chemical interactions and hard-core repulsions must be considered when assessing the effects of crowding and help explain previous observations about protein stability and dynamics in cells. PMID:22954326

  12. Protein Surface Characterization Using an Invariant Descriptor

    PubMed Central

    Abu Deeb, Zainab; Adjeroh, Donald A.; Jiang, Bing-Hua

    2011-01-01

    Aim. To develop a new invariant descriptor for the characterization of protein surfaces, suitable for various analysis tasks, such as protein functional classification, and search and retrieval of protein surfaces over a large database. Methods. We start with a local descriptor of selected circular patches on the protein surface. The descriptor records the distance distribution between the central residue and the residues within the patch, keeping track of the number of particular pairwise residue cooccurrences in the patch. A global descriptor for the entire protein surface is then constructed by combining information from the local descriptors. Our method is novel in its focus on residue-specific distance distributions, and the use of residue-distance co-occurrences as the basis for the proposed protein surface descriptors. Results. Results are presented for protein classification and for retrieval for three protein families. For the three families, we obtained an area under the curve for precision and recall ranging from 0.6494 (without residue co-occurrences) to 0.6683 (with residue co-occurrences). Large-scale screening using two other protein families placed related family members at the top of the rank, with a number of uncharacterized proteins also retrieved. Comparative results with other proposed methods are included. PMID:22144981

  13. Infrared Protein Crystallography

    SciTech Connect

    J Sage; Y Zhang; J McGeehan; R Ravelli; M Weik; J van Thor

    2011-12-31

    We consider the application of infrared spectroscopy to protein crystals, with particular emphasis on exploiting molecular orientation through polarization measurements on oriented single crystals. Infrared microscopes enable transmission measurements on individual crystals using either thermal or nonthermal sources, and can accommodate flow cells, used to measure spectral changes induced by exposure to soluble ligands, and cryostreams, used for measurements of flash-cooled crystals. Comparison of unpolarized infrared measurements on crystals and solutions probes the effects of crystallization and can enhance the value of the structural models refined from X-ray diffraction data by establishing solution conditions under which they are most relevant. Results on several proteins are consistent with similar equilibrium conformational distributions in crystal and solutions. However, the rates of conformational change are often perturbed. Infrared measurements also detect products generated by X-ray exposure, including CO{sub 2}. Crystals with favorable symmetry exhibit infrared dichroism that enhances the synergy with X-ray crystallography. Polarized infrared measurements on crystals can distinguish spectral contributions from chemically similar sites, identify hydrogen bonding partners, and, in opportune situations, determine three-dimensional orientations of molecular groups. This article is part of a Special Issue entitled: Protein Structure and Function in the Crystalline State.

  14. Protein antioxidants in thalassemia.

    PubMed

    Awadallah, Samir

    2013-01-01

    It is common knowledge that thalassemic patients are under significant oxidative stress. Chronic hemolysis, frequent blood transfusion, and increased intestinal absorption of iron are the main factors that result in iron overload with its subsequent pathophysiologic complications. Iron overload frequently associates with the generation of redox-reactive labile iron, which in turn promotes the production of other reactive oxygen species (ROS). If not neutralized, uncontrolled production of ROS often leads to damage of various intra- and extracellular components such as DNA, proteins, lipids, and small antioxidant molecules among others. A number of endogenous and exogenous defense mechanisms can neutralize and counteract the damaging effects of labile iron and the reactive substances associated with it. Endogenous antioxidant enzymes, such as superoxide dismutase, catalase, glutathione peroxidase, and ferroxidase, may directly or sequentially terminate the activities of ROS. Nonenzymatic endogenous defense mechanisms include metal binding proteins (ceruloplasmin, haptoglobin, albumin, and others) and endogenously produced free radical scavengers (glutathione (GSH), ubiquinols, and uric acid). Exogenous agents that are known to function as antioxidants (vitamins C and E, selenium, and zinc) are mostly diet-derived. In this review, we explore recent findings related to various antioxidative mechanisms operative in thalassemic patients with special emphasis on protein antioxidants. PMID:23724742

  15. Characterization of membrane association of Rinderpest virus matrix protein

    SciTech Connect

    Subhashri, R. [Department of Microbiology and Cell biology, Indian Institute of Science, Bangalore, Karnataka 560012 (India); Shaila, M.S. [Department of Microbiology and Cell biology, Indian Institute of Science, Bangalore, Karnataka 560012 (India)]. E-mail: shaila@mcbl.iisc.ernet.in

    2007-04-20

    Paramyxovirus matrix protein is believed to play a crucial role in the assembly and maturation of the virus particle by bringing the major viral components together at the budding site in the host cell. The membrane association capability of many enveloped virus matrix proteins has been characterized to be their intrinsic property. In this work, we have characterized the membrane association of Rinderpest virus matrix (M) protein. The M protein of Rinderpest virus when expressed in the absence of other viral proteins is present both in the cytoplasm and plasma membrane. When expressed as GFP fusion protein, the M protein gets localized into plasma membrane protrusions. High salt and alkaline conditions resulted in partial dissociation of M protein from cell membrane. Thus, M protein behaves like an integral membrane protein although its primary structure suggests it to be a peripheral membrane protein.

  16. Enriched Protein-Protein Interactions from Biomedical Text

    E-print Network

    Edinburgh, University of

    Enriched Protein-Protein Interactions from Biomedical Text Barry Haddow, Michael Matthews University of Edinburgh 13th March 2007 Barry Haddow, Michael Matthews Enriched Protein-Protein Interactions from Biomedical Text #12;Overview The TXM Project Protein-Protein Interactions Enriched Protein-Protein

  17. Prion protein and aging

    PubMed Central

    Gasperini, Lisa; Legname, Giuseppe

    2014-01-01

    The cellular prion protein (PrPC) has been widely investigated ever since its conformational isoform, the prion (or PrPSc), was identified as the etiological agent of prion disorders. The high homology shared by the PrPC-encoding gene among mammals, its high turnover rate and expression in every tissue strongly suggest that PrPC may possess key physiological functions. Therefore, defining PrPC roles, properties and fate in the physiology of mammalian cells would be fundamental to understand its pathological involvement in prion diseases. Since the incidence of these neurodegenerative disorders is enhanced in aging, understanding PrPC functions in this life phase may be of crucial importance. Indeed, a large body of evidence suggests that PrPC plays a neuroprotective and antioxidant role. Moreover, it has been suggested that PrPC is involved in Alzheimer disease, another neurodegenerative pathology that develops predominantly in the aging population. In prion diseases, PrPC function is likely lost upon protein aggregation occurring in the course of the disease. Additionally, the aging process may alter PrPC biochemical properties, thus influencing its propensity to convert into PrPSc. Both phenomena may contribute to the disease development and progression. In Alzheimer disease, PrPC has a controversial role because its presence seems to mediate ?-amyloid toxicity, while its down-regulation correlates with neuronal death. The role of PrPC in aging has been investigated from different perspectives, often leading to contrasting results. The putative protein functions in aging have been studied in relation to memory, behavior and myelin maintenance. In aging mice, PrPC changes in subcellular localization and post-translational modifications have been explored in an attempt to relate them to different protein roles and propensity to convert into PrPSc. Here we provide an overview of the most relevant studies attempting to delineate PrPC functions and fate in aging. PMID:25364751

  18. Protein derivitization-expressed protein ligation.

    PubMed

    Mitchell, Sarah F; Lorsch, Jon R

    2014-01-01

    Expressed protein ligation (EPL) combines two methods to ligate a synthetic peptide to a recombinant protein. Native chemical ligation (NCL) is a process in which two synthesized peptides are ligated by reaction of a C-terminal thioester on one peptide with an N-terminal cysteine residue of another protein. The chemistry of inteins, self-excising protein fragments that ligate the surrounding protein back together, creates isolatable intermediates with the two chemical groups necessary for NCL, a C-terminal thioester and an N-terminal cysteine residue. This technique allows for the incorporation of synthetic amino acids, radiolabeled amino acids, and fluorescent moieties at specific locations in a protein. It has the advantage of allowing attachment of such synthetic peptides to the termini of a recombinant protein, allowing for the synthesis of large proteins with modified amino acids. This technique utilizes the IMPACT(TM)-System created by New England Biolabs, who provide a variety of vectors in which the multicloning site is directly upstream of an intein sequence fused to a chitin-binding domain (CBD). The CBD binds tightly and specifically to chitin beads, allowing for an efficient one-step purification. This step can be used to obtain highly purified proteins (see Protein Affinity Purification using Intein/Chitin Binding Protein Tags). After purification of the recombinant protein, cleavage from the intein is achieved through the addition of a reactive thiol compound, usually sodium 2-mercaptoethanesulfonate (MESNA) (see also Proteolytic affinity tag cleavage). This reaction creates a protein with a C-terminal thioester that can then react with a peptide containing an N-terminal cysteine residue, ligating the two proteins via a peptide bond. PMID:24423270

  19. My 65 years in protein chemistry.

    PubMed

    Scheraga, Harold A

    2015-05-01

    This is a tour of a physical chemist through 65 years of protein chemistry from the time when emphasis was placed on the determination of the size and shape of the protein molecule as a colloidal particle, with an early breakthrough by James Sumner, followed by Linus Pauling and Fred Sanger, that a protein was a real molecule, albeit a macromolecule. It deals with the recognition of the nature and importance of hydrogen bonds and hydrophobic interactions in determining the structure, properties, and biological function of proteins until the present acquisition of an understanding of the structure, thermodynamics, and folding pathways from a linear array of amino acids to a biological entity. Along the way, with a combination of experiment and theoretical interpretation, a mechanism was elucidated for the thrombin-induced conversion of fibrinogen to a fibrin blood clot and for the oxidative-folding pathways of ribonuclease A. Before the atomic structure of a protein molecule was determined by x-ray diffraction or nuclear magnetic resonance spectroscopy, experimental studies of the fundamental interactions underlying protein structure led to several distance constraints which motivated the theoretical approach to determine protein structure, and culminated in the Empirical Conformational Energy Program for Peptides (ECEPP), an all-atom force field, with which the structures of fibrous collagen-like proteins and the 46-residue globular staphylococcal protein A were determined. To undertake the study of larger globular proteins, a physics-based coarse-grained UNited-RESidue (UNRES) force field was developed, and applied to the protein-folding problem in terms of structure, thermodynamics, dynamics, and folding pathways. Initially, single-chain and, ultimately, multiple-chain proteins were examined, and the methodology was extended to protein-protein interactions and to nucleic acids and to protein-nucleic acid interactions. The ultimate results led to an understanding of a variety of biological processes underlying natural and disease phenomena. PMID:25850343

  20. Protein N-Myristoylation Plays a Critical Role in the Endoplasmic Reticulum Morphological Change Induced by Overexpression of Protein Lunapark, an Integral Membrane Protein of the Endoplasmic Reticulum

    PubMed Central

    Moriya, Koko; Nagatoshi, Kei; Noriyasu, Yoshimi; Okamura, Tsuyoshi; Takamitsu, Emi; Suzuki, Takashi; Utsumi, Toshihiko

    2013-01-01

    N-myristoylation of eukaryotic cellular proteins has been recognized as a modification that occurs mainly on cytoplasmic proteins. In this study, we examined the membrane localization, membrane integration, and intracellular localization of four recently identified human N-myristoylated proteins with predicted transmembrane domains. As a result, it was found that protein Lunapark, the human ortholog of yeast protein Lnp1p that has recently been found to be involved in network formation of the endoplasmic reticulum (ER), is an N-myristoylated polytopic integral membrane protein. Analysis of tumor necrosis factor-fusion proteins with each of the two putative transmembrane domains and their flanking regions of protein Lunapark revealed that transmembrane domain 1 and 2 functioned as type II signal anchor sequence and stop transfer sequence, respectively, and together generated a double-spanning integral membrane protein with an N-/C-terminal cytoplasmic orientation. Immunofluorescence staining of HEK293T cells transfected with a cDNA encoding protein Lunapark tagged with FLAG-tag at its C-terminus revealed that overexpressed protein Lunapark localized mainly to the peripheral ER and induced the formation of large polygonal tubular structures. Morphological changes in the ER induced by overexpressed protein Lunapark were significantly inhibited by the inhibition of protein N-myristoylation by means of replacing Gly2 with Ala. These results indicated that protein N-myristoylation plays a critical role in the ER morphological change induced by overexpression of protein Lunapark. PMID:24223779

  1. Network Analysis of Circular Permutations in Multidomain Proteins Reveals Functional Linkages for Uncharacterized Proteins

    PubMed Central

    Adjeroh, Donald; Jiang, Yue; Jiang, Bing-Hua; Lin, Jie

    2014-01-01

    Various studies have implicated different multidomain proteins in cancer. However, there has been little or no detailed study on the role of circular multidomain proteins in the general problem of cancer or on specific cancer types. This work represents an initial attempt at investigating the potential for predicting linkages between known cancer-associated proteins with uncharacterized or hypothetical multidomain proteins, based primarily on circular permutation (CP) relationships. First, we propose an efficient algorithm for rapid identification of both exact and approximate CPs in multidomain proteins. Using the circular relations identified, we construct networks between multidomain proteins, based on which we perform functional annotation of multidomain proteins. We then extend the method to construct subnetworks for selected cancer subtypes, and performed prediction of potential link-ages between uncharacterized multidomain proteins and the selected cancer types. We include practical results showing the performance of the proposed methods. PMID:25741177

  2. Seed proteins of the wild and the cultivated Amaranthus species

    Microsoft Academic Search

    A. V. Zheleznov; L. P. Solonenko; N. B. Zheleznova

    1997-01-01

    The 13–21% variation in seed protein content was observed in wild and cultivated forms of amaranth. Seed proteins of amaranth\\u000a are highly nutritive and composed presumably of easily digestable albumins and globulins (over 50% of total protein); of 20.8%\\u000a alkali-soluble proteins-glutelins with similar nutritive value and only of 12% alcohol-soluble proteins-prolamins which are\\u000a lacking in essential amino acids. The results

  3. Theory of protein folding

    Microsoft Academic Search

    José Nelson Onuchic; Peter G. Wolynes

    2004-01-01

    Protein folding should be complex. Proteins organize themselves into specific three-dimensional structures, through a myriad of conformational changes. The classical view of protein folding describes this process as a nearly sequential series of discrete intermediates. In contrast, the energy landscape theory of folding considers folding as the progressive organization of an ensemble of partially folded structures through which the protein

  4. PROTEINS, HARDNESS AND ALLERGENS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteins are key constituents of wheat grain and flour. A number of specific proteins/classes of proteins are critical to the processing and end-use quality, and hence value and utility of wheat. The commercially important proteins of wheat may be broadly classified as glutenins and gliadins of va...

  5. Protein folding by interaction.

    PubMed

    Buchner, Johannes; Kessler, Horst

    2014-07-01

    Repeat proteins consisting of helical segments seem to fold by a matrix-assisted mechanism in which folded segments induce structure in intrinsically disordered parts of the protein, as shown by Watson and colleagues in this issue of Structure for an Armadillo repeat protein and previously by the Balbach group for an Ankyrin repeat protein. PMID:25007223

  6. Heterotrimeric G Protein Cycle

    NSDL National Science Digital Library

    Anita Preininger (Vanderbilt University Medical Center; Department of Pharmacology)

    2004-02-03

    This animation shows the basic heterotrimeric G protein cycle and allows the user to then add three different regulators of the cycle, an RGS (regulator of G protein signaling) protein, a GDI (guanine nucleotide dissociation inhibitor) protein, or a guanine nucleotide exchange factor (GEF).

  7. Recombinant Protein Purification Handbook

    E-print Network

    Lebendiker, Mario

    Recombinant Protein Purification Handbook Principles and Methods GE Healthcare #12;GST Gene Fusion-6429-60 Microcarrier Cell Culture Principles and Methods 18-1140-62 Challenging Protein Purification Handbook 28-9095-31 Recombinant Protein Purification Handbook Principles and Methods 18-1142-75 Protein Purification Handbook 18

  8. Dynamic personalities of proteins

    Microsoft Academic Search

    Katherine Henzler-Wildman; Dorothee Kern

    2007-01-01

    Because proteins are central to cellular function, researchers have sought to uncover the secrets of how these complex macromolecules execute such a fascinating variety of functions. Although static structures are known for many proteins, the functions of proteins are governed ultimately by their dynamic character (or 'personality'). The dream is to 'watch' proteins in action in real time at atomic

  9. PREFACE: Protein protein interactions: principles and predictions

    NASA Astrophysics Data System (ADS)

    Nussinov, Ruth; Tsai, Chung-Jung

    2005-06-01

    Proteins are the `workhorses' of the cell. Their roles span functions as diverse as being molecular machines and signalling. They carry out catalytic reactions, transport, form viral capsids, traverse membranes and form regulated channels, transmit information from DNA to RNA, making possible the synthesis of new proteins, and they are responsible for the degradation of unnecessary proteins and nucleic acids. They are the vehicles of the immune response and are responsible for viral entry into the cell. Given their importance, considerable effort has been centered on the prediction of protein function. A prime way to do this is through identification of binding partners. If the function of at least one of the components with which the protein interacts is known, that should let us assign its function(s) and the pathway(s) in which it plays a role. This holds since the vast majority of their chores in the living cell involve protein-protein interactions. Hence, through the intricate network of these interactions we can map cellular pathways, their interconnectivities and their dynamic regulation. Their identification is at the heart of functional genomics; their prediction is crucial for drug discovery. Knowledge of the pathway, its topology, length, and dynamics may provide useful information for forecasting side effects. The goal of predicting protein-protein interactions is daunting. Some associations are obligatory, others are continuously forming and dissociating. In principle, from the physical standpoint, any two proteins can interact, but under what conditions and at which strength? The principles of protein-protein interactions are general: the non-covalent interactions of two proteins are largely the outcome of the hydrophobic effect, which drives the interactions. In addition, hydrogen bonds and electrostatic interactions play important roles. Thus, many of the interactions observed in vitro are the outcome of experimental overexpression. Protein disorder is important in protein-protein association. It has been estimated that a large fraction of cellular proteins are `natively disordered', i.e., unstable in solution. The disordered state has a significant residual structure. In this state, a protein exists in an ensemble of rapidly interconverting conformers. They play roles in cell-cycle control, signal transduction, transcriptional and translational regulation, and in large macromolecular complexes. It has been suggested that natively disordered proteins are more `adaptive', and thus advantageous in regulation and in binding diverse ligands. Alternatively, since the native conformation is still likely to be the most abundant within the ensemble, disordered proteins, which typically have larger interface to size ratios, lead to smaller protein, genome and cell sizes, and thus are functionally advantageous. To be able to predict protein-protein interactions, we need to discern various aspects of their associations: from their shape complementarity to the organization and relative contributions of the different physical components to their stability. They involve the static and the dynamic. Proteins interact through their surfaces. Thus, to analyze their interactions, we typically study residues (or atoms) which are in contact across the two-chain interface. In addition, we often inspect the residues in their vicinity, exploring their supporting matrix. The hope is that through the understanding of the principles and mechanisms of the interactions, we shall eventually be able to solve the protein-protein interaction puzzle.

  10. Therapeutic Protein Aggregation: Mechanisms, Design, and Control

    PubMed Central

    Roberts, Christopher J.

    2014-01-01

    While it is well known that proteins are only marginally stable in their folded states, it is often less well appreciated that most proteins are inherently aggregation-prone in their unfolded or partially unfolded states, and the resulting aggregates can be extremely stable and long-lived. For therapeutic proteins, aggregates are a significant risk factor for deleterious immune responses in patients, and can form via a variety of mechanisms. Controlling aggregation using a mechanistic approach may allow improved design of therapeutic protein stability, as a complement to existing design strategies that target desired protein structures and function. Recent results highlight the importance of balancing protein environment with the inherent aggregation propensities of polypeptide chains. PMID:24908382

  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. A nutrigenomics view of protein intake: macronutrient, bioactive peptides, and protein turnover.

    PubMed

    Chou, Chieh Jason; Affolter, Michael; Kussmann, Martin

    2012-01-01

    Proteins are needed for the development and sustainability of life. They are the molecular machines and building blocks in the human body that drive or exert most biological functions and confer structure and function to cell and tissue architecture. Dietary proteins provide essential amino acids and complement lipid and carbohydrate as a major source of energy. Therefore, humans must consume a sufficient amount and quality of proteins to stay healthy and avoid deficiencies. Even with a reasonable amount of intake, variability in protein consumption can result in measurable health consequences in specific conditions. This said, dietary protein delivers more than energy and building blocks to the human body: the pools of body, tissue, and cell proteins, peptides, and amino acids are under complex metabolic control, resulting in a highly dynamic protein turnover, that is, the interplay between synthesis and degradation. Proteins also contain peptide sequences that can be interpreted as bioactive precursors which can be liberated upon digestion to exert biological functions locally (e.g., in the gut) or systemically (i.e., via the bloodstream). In this chapter, we will first review holistic readouts of protein intake assessed by omics technologies such as gene expression, proteomics, and metabolite profiling. Second, we will look at protein benefits beyond macronutrient supply and describe how to generate, analyze, and leverage bioactive peptides. In the third part, we will discuss protein turnover as tackled by proteomics tools that allow single-protein resolution at proteome-wide scale. PMID:22656373

  13. Genome-Wide Protein Interaction Screens Reveal Functional Networks Involving Sm-Like Proteins

    PubMed Central

    Fromont-Racine, Micheline; Mayes, Andrew E.; Brunet-Simon, Adeline; Rain, Jean-Christophe; Colley, Alan; Dix, Ian; Decourty, Laurence; Joly, Nicolas; Ricard, Florence; Beggs, Jean D.

    2000-01-01

    A set of seven structurally related Sm proteins forms the core of the snRNP particles containing the spliceosomal U1, U2, U4 and U5 snRNAs. A search of the genomic sequence of Saccharomyces cerevisiae has identified a number of open reading frames that potentially encode structurally similar proteins termed Lsm (Like Sm) proteins. With the aim of analysing all possible interactions between the Lsm proteins and any protein encoded in the yeast genome, we performed exhaustive and iterative genomic two-hybrid screens, starting with the Lsm proteins as baits. Indeed, extensive interactions amongst eight Lsm proteins were found that suggest the existence of a Lsm complex or complexes. These Lsm interactions apparently involve the conserved Sm domain that also mediates interactions between the Sm proteins. The screens also reveal functionally significant interactions with splicing factors, in particular with Prp4 and Prp24, compatible with genetic studies and with the reported association of Lsm proteins with spliceosomal U6 and U4/U6 particles. In addition, interactions with proteins involved in mRNA turnover, such as Mrt1, Dcp1, Dcp2 and Xrn1, point to roles for Lsm complexes in distinct RNA metabolic processes, that are confirmed in independent functional studies. These results provide compelling evidence that two-hybrid screens yield functionally meaningful information about protein–protein interactions and can suggest functions for uncharacterized proteins, especially when they are performed on a genome-wide scale. PMID:10900456

  14. Defective Proteasome Delivery of Polyubiquitinated Proteins by Ubiquilin-2 Proteins Containing ALS Mutations

    PubMed Central

    Chang, Lydia; Monteiro, Mervyn J.

    2015-01-01

    Ubiquilin proteins facilitate delivery of ubiquitinated proteins to the proteasome for degradation. Interest in the proteins has been heightened by the discovery that gene mutations in UBQLN2 cause dominant inheritance of amyotrophic lateral sclerosis (ALS). However, the mechanisms by which the mutations cause ALS are not known. Here we report on the underlying defect of ubiquilin-2 proteins containing ALS-linked mutations in affecting proteasome-mediated degradation. We found that overexpression of ubiquilin-2 proteins containing any one of five different ALS mutations slow degradation of Myc, a prototypic proteasome substrate. Examination of coprecipitating proteins indicated that the mutant proteins are generally capable of binding polyubiquitinated proteins, but defective in binding the proteasome. GST-pulldown studies revealed that many of the mutants bind weaker to the S5a subunit of the proteasome, compared with wild type (WT) ubiquilin-2 protein. The results suggest the mutant proteins are unable to deliver their captured cargo to the proteasome for degradation, which presumably leads to toxicity. Quantification of cell death is consistent with this idea. Measurement of protein turnover further indicated the mutant proteins have longer half-lives than WT ubiquilin-2. Our studies provide novel insight into the mechanism by which ALS-linked mutations in UBQLN2 interfere with protein degradation. PMID:26075709

  15. Protein Sequence, Structure, Stability and Functionality

    E-print Network

    J. C. Phillips

    2008-02-25

    Protein-protein interactions (protein functionalities) are mediated by water, which compacts individual proteins and promotes close and temporarily stable large-area protein-protein interfaces. Proteins are peptide chains decorated by amino acids, and protein scientists have long described protein-water interactions in terms of qualitative amino acid hydrophobicity scales. Here we examine several recent scales and argue plausibly (in terms of self-organized criticality) that one of them should be regarded as an absolute scale (within the protein universe), analogous to the dielectric scale of bond ionicity in inorganic octet compounds. Applications to repeat proteins (containing upwards of 900 amino acids) are successful, far beyond reasonable expectations, in all cases studied so far. While some of the results are obvious and can be obtained from the ex vitro spatial structures alone, many are hidden from plain view, and can be called phantom relations. As a byproduct, the network theory explains the exceptional functionality of leucine in zippers, heptads, and repeat consensus sites.

  16. Building blocks for protein interaction devices

    PubMed Central

    Grünberg, Raik; Ferrar, Tony S.; van der Sloot, Almer M.; Constante, Marco; Serrano, Luis

    2010-01-01

    Here, we propose a framework for the design of synthetic protein networks from modular protein–protein or protein–peptide interactions and provide a starter toolkit of protein building blocks. Our proof of concept experiments outline a general work flow for part–based protein systems engineering. We streamlined the iterative BioBrick cloning protocol and assembled 25 synthetic multidomain proteins each from seven standardized DNA fragments. A systematic screen revealed two main factors controlling protein expression in Escherichia coli: obstruction of translation initiation by mRNA secondary structure or toxicity of individual domains. Eventually, 13 proteins were purified for further characterization. Starting from well-established biotechnological tools, two general–purpose interaction input and two readout devices were built and characterized in vitro. Constitutive interaction input was achieved with a pair of synthetic leucine zippers. The second interaction was drug-controlled utilizing the rapamycin-induced binding of FRB(T2098L) to FKBP12. The interaction kinetics of both devices were analyzed by surface plasmon resonance. Readout was based on Förster resonance energy transfer between fluorescent proteins and was quantified for various combinations of input and output devices. Our results demonstrate the feasibility of parts-based protein synthetic biology. Additionally, we identify future challenges and limitations of modular design along with approaches to address them. PMID:20215443

  17. Protein Folding and Misfolding on Surfaces

    PubMed Central

    Stefani, Massimo

    2008-01-01

    Protein folding, misfolding and aggregation, as well as the way misfolded and aggregated proteins affects cell viability are emerging as key themes in molecular and structural biology and in molecular medicine. Recent advances in the knowledge of the biophysical basis of protein folding have led to propose the energy landscape theory which provides a consistent framework to better understand how a protein folds rapidly and efficiently to the compact, biologically active structure. The increased knowledge on protein folding has highlighted its strict relation to protein misfolding and aggregation, either process being in close competition with the other, both relying on the same physicochemical basis. The theory has also provided information to better understand the structural and environmental factors affecting protein folding resulting in protein misfolding and aggregation into ordered or disordered polymeric assemblies. Among these, particular importance is given to the effects of surfaces. The latter, in some cases make possible rapid and efficient protein folding but most often recruit proteins/peptides increasing their local concentration thus favouring misfolding and accelerating the rate of nucleation. It is also emerging that surfaces can modify the path of protein misfolding and aggregation generating oligomers and polymers structurally different from those arising in the bulk solution and endowed with different physical properties and cytotoxicities. PMID:19330090

  18. Evolutionary aspect of protein self-organization

    NASA Astrophysics Data System (ADS)

    Rapis, E.

    2008-06-01

    Numerous experimental data (published in 1988 2006) show that, when an open protein-water system far from thermodynamic equilibrium is dehydrated (dried), abiogenic self-organization of the protein invariably takes place, which complicates the structure and also results in the formation of a 3D supramolecular architecture with synchronous replication of spiral vortices and domains (cells) with nuclei having spiral clockwise and counterclockwise symmetry typical of protein in the living organism. When a solvent evaporates, say, from a multicomponent solution, such as blood serum, a protein structure arises the morphology of which copies the morphology of the protein one-component system. Thus, the competing activity of protein is observed when it experiences phase transition in the course of self-organization. In light of a new evolutionary chemical theory based on the Rudenko concept, these data allow one to put forward a hypothesis that protein exhibits evolutionary properties under conditions far from thermodynamic equilibrium. This hypothesis relies on the assumption that the energetically active structure of protein self-organizing in the course of its phase transition may generate energy necessary for catalysis and autocatalysis when a one-component protein-water system dries out. An important piece of evidence in favor of this hypothesis is the presence of the basic type of symmetry (spiral mirror clockwise or counterclockwise symmetry) under the given nonequilibrium conditions in vitro, which is characteristic of animate nature, protein in the living organism in vivo, and abiogenic self-organization of protein in vitro.

  19. further results

    E-print Network

    SIAM (#1) 1035 1999 Jan 20 12:53:14

    2000-04-26

    placement or pure traction boundary conditions in two or three dimensions, and ... result assures optimal H1-like performance for standard finite element .... to absorbing Re into the pressure variable and source term, i.e., to replacing Re p by.

  20. Coevolution of gene expression among interacting proteins

    SciTech Connect

    Fraser, Hunter B.; Hirsh, Aaron E.; Wall, Dennis P.; Eisen,Michael B.

    2004-03-01

    Physically interacting proteins or parts of proteins are expected to evolve in a coordinated manner that preserves proper interactions. Such coevolution at the amino acid-sequence level is well documented and has been used to predict interacting proteins, domains, and amino acids. Interacting proteins are also often precisely coexpressed with one another, presumably to maintain proper stoichiometry among interacting components. Here, we show that the expression levels of physically interacting proteins coevolve. We estimate average expression levels of genes from four closely related fungi of the genus Saccharomyces using the codon adaptation index and show that expression levels of interacting proteins exhibit coordinated changes in these different species. We find that this coevolution of expression is a more powerful predictor of physical interaction than is coevolution of amino acid sequence. These results demonstrate previously uncharacterized coevolution of gene expression, adding a different dimension to the study of the coevolution of interacting proteins and underscoring the importance of maintaining coexpression of interacting proteins over evolutionary time. Our results also suggest that expression coevolution can be used for computational prediction of protein protein interactions.

  1. Estimating protein function using protein-protein relationships.

    PubMed

    Date, Shailesh V

    2007-01-01

    Many newly identified gene products from completely sequenced genomes are difficult to characterize in the absence of sequence homology to known proteins. In such a scenario, the context of the proteins' functional associations can be used for annotation; overrepresented functional linkages with a certain class of proteins or members of a pathway allow putative function assignments based on the "guilt-by-association" principle. Two computational functional genomics methods, phylogenetic profiling and identification of Rosetta stone linkages, are described in this chapter, which allow assessment of functional linkages between proteins, consequently facilitating annotation. Phylogenetic profiling involves measuring similarity between profiles that describe the presence or absence of a protein in a set of reference genomes, whereas Rosetta stone fusion sequences help link two or more independently transcribed and translated proteins. Both methods can be applied to investigate functional associations between individual proteins, and can also be extended to reconstruct the genome-wide network of functional linkages by querying the entire protein complement of an organism. PMID:18314580

  2. TULA proteins regulate activity of the protein tyrosine kinase Syk.

    PubMed

    Agrawal, Rachana; Carpino, Nick; Tsygankov, Alexander

    2008-06-01

    TULA belongs to a two-member family: TULA (STS-2) is a lymphoid protein, whereas STS-1/TULA-2 is expressed ubiquitously. TULA proteins were implicated in the regulation of signaling mediated by protein tyrosine kinases (PTKs). The initial experiments did not fully reveal the molecular mechanism of these effects, but suggested that both TULA proteins act in a similar fashion. It was shown recently that STS-1/TULA-2 dephosphorylates PTKs. In this study, we analyzed the effects of TULA proteins on Syk, a PTK playing an important role in lymphoid signaling. First, we have shown that TULA-2 decreases tyrosine phosphorylation of Syk in vivo and in vitro and that the intact phosphatase domain of TULA-2 is essential for this effect. We have also shown that TULA-2 exhibits a certain degree of substrate specificity. Our results also indicate that inactivated TULA-2 increases tyrosine phosphorylation of Syk in cells co-transfected to overexpress these proteins, thus acting as a dominant-negative form that suppresses dephosphorylation of Syk caused by endogenous TULA-2. Furthermore, we have demonstrated that phosphatase activity of TULA is negligible as compared to that of TULA-2 and that this finding correlates with an increase in Syk tyrosine phosphorylation in cells overexpressing TULA. This result is consistent with the dominant-negative effect of inactivated TULA-2, arguing that TULA acts in this system as a negative regulator of TULA-2-dependent dephosphorylation. To summarize, our findings indicate that TULA proteins may exert opposite effects on PTK-mediated signaling and suggest that a regulatory mechanism based on this feature may exist. PMID:18189269

  3. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2009 BINF 731 Protein Engineering Protein Engineering stability Increase proteins resistance to proteases Change codon composition Computational Mutagenesis Assumption: the structural differences between each mutant and the wild-type protein are usually minor and

  4. 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.

  5. Determination of Protein Carbonyl Groups by Immunoblotting

    Microsoft Academic Search

    C. E. Robinson; A. Keshavarzian; D. S. Pasco; T. O. Frommel; D. H. Winship; E. W. Holmes

    1999-01-01

    Free radical-mediated oxidation of proteins results in the formation of carbonyl groups in quantities that reflect the intensity of the oxidative stress. We have developed an immunochemical technique for the quantification of carbonyl groups in protein samples prepared from small tissue samples and cell cultures. Protein samples were slot-blotted onto a polyvinylidene difluoride membrane, which was sequentially treated with 2,4-dinitrophenylhydrazine

  6. Plastic deformation of protein monolayers.

    PubMed Central

    Singh-Zocchi, Mukta; Hanne, Jeungphill; Zocchi, Giovanni

    2002-01-01

    Globular proteins are peculiar solids that display both local stability of their conformation and the ability to undergo large cooperative changes of shape (conformational changes). If one forces a large deformation of the molecule, such that the structure is necessarily changed, it is not obvious whether the deformed globule can still remain a solid or whether it will melt. Is it possible to plastically deform a protein? Here we investigate this question with a micro-mechanical experiment on a small region (approximately 10 molecules) of a protein monolayer adsorbed on a rigid surface. For the two proteins studied, albumin and myoglobin, we observed that the molecules can be substantially deformed (approximately 1-2 nm deformation) by comparatively small stresses applied for sufficiently long times. The deformation is irreversible, and the protein remains in the solid state (i.e., displays a nonzero shear modulus). The dynamics of the deformation is approximately logarithmic in time, similar to creep in solids. These results show that globular proteins adsorbed on a surface can be plastically deformed. PMID:12324438

  7. Protein-protein docking on hardware accelerators: comparison of GPU and MIC architectures

    PubMed Central

    2015-01-01

    Background The hardware accelerators will provide solutions to computationally complex problems in bioinformatics fields. However, the effect of acceleration depends on the nature of the application, thus selection of an appropriate accelerator requires some consideration. Results In the present study, we compared the effects of acceleration using graphics processing unit (GPU) and many integrated core (MIC) on the speed of fast Fourier transform (FFT)-based protein-protein docking calculation. The GPU implementation performed the protein-protein docking calculations approximately five times faster than the MIC offload mode implementation. The MIC native mode implementation has the advantage in the implementation costs. However, the performance was worse with larger protein pairs because of memory limitations. Conclusion The results suggest that GPU is more suitable than MIC for accelerating FFT-based protein-protein docking applications. PMID:25707855

  8. 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 Ĺ 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

  9. Finding Occurrences of Protein Complexes in Protein-Protein Interaction Graphs ,

    E-print Network

    Paris-Sud XI, Université de

    Finding Occurrences of Protein Complexes in Protein-Protein Interaction Graphs , Guillaume Fertin analysis of protein-protein interaction graphs, we use a graph-based formalism to detect the preservation of a given protein complex G in the protein-protein interaction graph H of another species with respect to (w

  10. Electrostatic Properties of Protein-Protein Complexes

    Microsoft Academic Search

    Petras J. Kundrotas; Emil Alexov

    2006-01-01

    Statistical electrostatic analysis of 37 protein-protein complexes extracted from the previously developed database of protein complexes (ProtCom, http:\\/\\/www.ces.clemson.edu\\/compbio\\/protcom) is presented. It is shown that small interfaces have a higher content of charged and polar groups compared to large interfaces. In a vast majority of the cases the average pKa shifts for acidic residues induced by the complex formation are negative,

  11. Exploiting Protein Structures to Predict Protein Functions

    Microsoft Academic Search

    Alison Cuff; Oliver Redfern; Benoit Dessailly; Christine Orengo

    \\u000a The exponential growth of experimentally determined protein structures in the Protein Data Bank (PDB) has provided structural\\u000a data for an ever increasing proportion of genomic sequences. In combination with enhanced functional annotation from sequence,\\u000a it has become possible to predict protein function from structure. In this chapter we discuss a range of methods which aim\\u000a to recognise enzyme active sites

  12. Proteins as "dopable" bio-electronic materials

    NASA Astrophysics Data System (ADS)

    Cahen, David

    2013-02-01

    Proteins are surprisingly good solid-state electronic conductors. This holds also for proteins without any known biological electron transfer function. How do they do it? To answer this question we measure solid-state electron transport (ETp) across proteins that are "dry" (only tightly bound water, to retain the conformation, still present). We compare results for the electron transfer (ET) protein, Azurin (Az), the proton-pumping membrane protein Bacteriorhodopsin (bR), and for Human and Bovine Serum Albumin (HSA and BSA). Clear differences between these proteins are seen, which preserve their structure in the solid state measurement configuration. Importantly for future bioelectronics, the results are sensitive to protein modification, e.g., removing or disconnecting the retinal in bR and removing or replacing the Cu redox centre in Az. These cofactors can thus be viewed as natural dopants for proteins. Insight in the ETp mechanism comes from temperature-dependent studies. Az shows 40-360K temperature-independent ETp across its 3.5 nm long axis, until its denaturation temperature, indicative of tunneling. Cu removal, replacement (by Zn) or deuteration changes this to thermally activated ETp. This suggests hopping and involvement of the amide backbone in the ETp. The latter, which rhymes with indications from ETp experiments on oligopeptide and simulations of ET in proteins, opens the way for modeling what otherwise is an awfully complex system. Below 200K all proteins and their variants show temperature-independent ETp. We can furthermore make a totally electrically inactive protein, HSA, into an efficient ETp medium by doping it with natural poly-ene. Putting our data in perspective by comparing them to all known protein ETp data in the literature, we conclude that, in general, proteins are well described as dopable molecular wires.

  13. ProtPhylo: identification of protein–phenotype and protein–protein functional associations via phylogenetic profiling

    PubMed Central

    Cheng, Yiming; Perocchi, Fabiana

    2015-01-01

    ProtPhylo is a web-based tool to identify proteins that are functionally linked to either a phenotype or a protein of interest based on co-evolution. ProtPhylo infers functional associations by comparing protein phylogenetic profiles (co-occurrence patterns of orthology relationships) for more than 9.7 million non-redundant protein sequences from all three domains of life. Users can query any of 2048 fully sequenced organisms, including 1678 bacteria, 255 eukaryotes and 115 archaea. In addition, they can tailor ProtPhylo to a particular kind of biological question by choosing among four main orthology inference methods based either on pair-wise sequence comparisons (One-way Best Hits and Best Reciprocal Hits) or clustering of orthologous proteins across multiple species (OrthoMCL and eggNOG). Next, ProtPhylo ranks phylogenetic neighbors of query proteins or phenotypic properties using the Hamming distance as a measure of similarity between pairs of phylogenetic profiles. Candidate hits can be easily and flexibly prioritized by complementary clues on subcellular localization, known protein–protein interactions, membrane spanning regions and protein domains. The resulting protein list can be quickly exported into a csv text file for further analyses. ProtPhylo is freely available at http://www.protphylo.org. PMID:25956654

  14. ProtPhylo: identification of protein-phenotype and protein-protein functional associations via phylogenetic profiling.

    PubMed

    Cheng, Yiming; Perocchi, Fabiana

    2015-07-01

    ProtPhylo is a web-based tool to identify proteins that are functionally linked to either a phenotype or a protein of interest based on co-evolution. ProtPhylo infers functional associations by comparing protein phylogenetic profiles (co-occurrence patterns of orthology relationships) for more than 9.7 million non-redundant protein sequences from all three domains of life. Users can query any of 2048 fully sequenced organisms, including 1678 bacteria, 255 eukaryotes and 115 archaea. In addition, they can tailor ProtPhylo to a particular kind of biological question by choosing among four main orthology inference methods based either on pair-wise sequence comparisons (One-way Best Hits and Best Reciprocal Hits) or clustering of orthologous proteins across multiple species (OrthoMCL and eggNOG). Next, ProtPhylo ranks phylogenetic neighbors of query proteins or phenotypic properties using the Hamming distance as a measure of similarity between pairs of phylogenetic profiles. Candidate hits can be easily and flexibly prioritized by complementary clues on subcellular localization, known protein-protein interactions, membrane spanning regions and protein domains. The resulting protein list can be quickly exported into a csv text file for further analyses. ProtPhylo is freely available at http://www.protphylo.org. PMID:25956654

  15. Algorithmics group, MPI for molecular genetics Delineation of protein

    E-print Network

    Spang, Rainer

    Algorithmics group, MPI for molecular genetics Delineation of protein complexes Wasinee Rungsarityotin September 15, 2006 IMPRS Colloquium #12;Algorithmics group, MPI for molecular genetics Overview · Result · Conclusion and outlook #12;Algorithmics group, MPI for molecular genetics Protein complex

  16. Mitochondrial proteases and protein quality control in ageing and longevity.

    PubMed

    Hamon, Marie-Paule; Bulteau, Anne-Laure; Friguet, Bertrand

    2015-09-01

    Mitochondria have been implicated in the ageing process and the lifespan modulation of model organisms. Mitochondria are the main providers of energy in eukaryotic cells but also represent both a major source of reactive oxygen species and targets for protein oxidative damage. Since protein damage can impair mitochondrial function, mitochondrial proteases are critically important for protein maintenance and elimination of oxidized protein. In the mitochondrial matrix, protein quality control is mainly achieved by the Lon and Clp proteases which are also key players in damaged mitochondrial proteins degradation. Accumulation of damaged macromolecules resulting from oxidative stress and failure of protein maintenance constitutes a hallmark of cellular and organismal ageing and is believed to participate to the age-related decline of cellular function. Hence, age-related impairment of mitochondrial protein quality control may therefore contribute to the age-associated build-up of oxidized protein and alterations of mitochondrial redox and protein homeostasis. PMID:25578288

  17. Boomerang results

    NASA Astrophysics Data System (ADS)

    de Bernardis, P.

    The BOOMERanG experiment has mapped the mm/sub-mm sky during two long duration balloon flights (in 1998 and 2003). The first flight has produced maps of about 4% of the sky at 90, 150, 240 and 410 GHz with resolution of 10'. The faint structure of the Cosmic Microwave Background at horizon and sub-horizon scales is evident in these maps, and the wide frequency coverage allows for a careful estimate of the Galactic foreground. In the second flight a polarization-sensitive version of the instrument has been flown, to measure the linear polarization of the microwave sky at 150, 240 and 350 GHz. Preliminary results and plans for future developments are reported.

  18. Protein-protein interactions: principles, techniques, and their potential role in new drug development.

    PubMed

    Khan, Shagufta H; Ahmad, Faizan; Ahmad, Nihal; Flynn, Daniel C; Kumar, Raj

    2011-06-01

    A vast network of genes is inter-linked through protein-protein interactions and is critical component of almost every biological process under physiological conditions. Any disruption of the biologically essential network leads to pathological conditions resulting into related diseases. Therefore, proper understanding of biological functions warrants a comprehensive knowledge of protein-protein interactions and the molecular mechanisms that govern such processes. The importance of protein-protein interaction process is highlighted by the fact that a number of powerful techniques/methods have been developed to understand how such interactions take place under various physiological and pathological conditions. Many of the key protein-protein interactions are known to participate in disease-associated signaling pathways, and represent novel targets for therapeutic intervention. Thus, controlling protein-protein interactions offers a rich dividend for the discovery of new drug targets. Availability of various tools to study and the knowledge of human genome have put us in a unique position to understand highly complex biological network, and the mechanisms involved therein. In this review article, we have summarized protein-protein interaction networks, techniques/methods of their binding/kinetic parameters, and the role of these interactions in the development of potential tools for drug designing. PMID:21469753

  19. Assessing Energetic Contributions to Binding from a Disordered Region in a Protein-Protein Interaction

    SciTech Connect

    S Cho; C Swaminathan; D Bonsor; M Kerzic; R Guan; J Yang; C Kieke; P Anderson; D Kranz; et al.

    2011-12-31

    Many functional proteins are at least partially disordered prior to binding. Although the structural transitions upon binding of disordered protein regions can influence the affinity and specificity of protein complexes, their precise energetic contributions to binding are unknown. Here, we use a model protein-protein interaction system in which a locally disordered region has been modified by directed evolution to quantitatively assess the thermodynamic and structural contributions to binding of disorder-to-order transitions. Through X-ray structure determination of the protein binding partners before and after complex formation and isothermal titration calorimetry of the interactions, we observe a correlation between protein ordering and binding affinity for complexes along this affinity maturation pathway. Additionally, we show that discrepancies between observed and calculated heat capacities based on buried surface area changes in the protein complexes can be explained largely by heat capacity changes that would result solely from folding the locally disordered region. Previously developed algorithms for predicting binding energies of protein-protein interactions, however, are unable to correctly model the energetic contributions of the structural transitions in our model system. While this highlights the shortcomings of current computational methods in modeling conformational flexibility, it suggests that the experimental methods used here could provide training sets of molecular interactions for improving these algorithms and further rationalizing molecular recognition in protein-protein interactions.

  20. Wheat F-box protein recruits proteins and regulates their abundance during wheat spike development.

    PubMed

    Hong, Min Jeong; Kim, Dae Yeon; Kang, Si Yong; Kim, Dong Sub; Kim, Jin Baek; Seo, Yong Weon

    2012-10-01

    F-box proteins, components of the Skp1-Cullin1-F-box (SCF) protein E3 ubiquitin ligase complex, serve as the variable component responsible for substrate recognition and recruitment in SCF-mediated proteolysis. F-box proteins interact with Skp1 through the F-box motif and with ubiquitination substrates through C-terminal protein interaction domains. F-box proteins regulate plant development, various hormonal signal transduction processes, circadian rhythm, and cell cycle control. We isolated an F-box protein gene from wheat spikes at the onset of flowering. The Triticum aestivum cyclin F-box domain (TaCFBD) gene showed elevated expression levels during early inflorescence development and under cold stress treatment. TaCFBD green fluorescent protein signals were localized in the cytoplasm and plasma membrane. We used yeast two-hybrid screening to identify proteins that potentially interact with TaCFBD. Fructose bisphosphate aldolase, aspartic protease, VHS, glycine-rich RNA-binding protein, and the 26S proteasome non-ATPase regulatory subunit were positive candidate proteins. The bimolecular fluorescence complementation assay revealed the interaction of TaCFBD with partner proteins in the plasma membranes of tobacco cells. Our results suggest that the TaCFBD protein acts as an adaptor between target substrates and the SCF complex and provides substrate specificity to the SCF of ubiquitin ligase complexes. PMID:22729884

  1. SPPS: A Sequence-Based Method for Predicting Probability of Protein-Protein Interaction Partners

    PubMed Central

    Huang, Zhimin; Shi, Ting; Chen, Yingyi; Zhang, Jian

    2012-01-01

    Background The molecular network sustained by different types of interactions among proteins is widely manifested as the fundamental driving force of cellular operations. Many biological functions are determined by the crosstalk between proteins rather than by the characteristics of their individual components. Thus, the searches for protein partners in global networks are imperative when attempting to address the principles of biology. Results We have developed a web-based tool “Sequence-based Protein Partners Search” (SPPS) to explore interacting partners of proteins, by searching over a large repertoire of proteins across many species. SPPS provides a database containing more than 60,000 protein sequences with annotations and a protein-partner search engine in two modes (Single Query and Multiple Query). Two interacting proteins of human FBXO6 protein have been found using the service in the study. In addition, users can refine potential protein partner hits by using annotations and possible interactive network in the SPPS web server. Conclusions SPPS provides a new type of tool to facilitate the identification of direct or indirect protein partners which may guide scientists on the investigation of new signaling pathways. The SPPS server is available to the public at http://mdl.shsmu.edu.cn/SPPS/. PMID:22292078

  2. Molecular and ultrastructural analysis of forisome subunits reveals the principles of forisome assembly

    PubMed Central

    Müller, Boje; Groscurth, Sira; Menzel, Matthias; Rüping, Boris A.; Twyman, Richard M.; Prüfer, Dirk; Noll, Gundula A.

    2014-01-01

    Background and Aims Forisomes are specialized structural phloem proteins that mediate sieve element occlusion after wounding exclusively in papilionoid legumes, but most studies of forisome structure and function have focused on the Old World clade rather than the early lineages. A comprehensive phylogenetic, molecular, structural and functional analysis of forisomes from species covering a broad spectrum of the papilionoid legumes was therefore carried out, including the first analysis of Dipteryx panamensis forisomes, representing the earliest branch of the Papilionoideae lineage. The aim was to study the molecular, structural and functional conservation among forisomes from different tribes and to establish the roles of individual forisome subunits. Methods Sequence analysis and bioinformatics were combined with structural and functional analysis of native forisomes and artificial forisome-like protein bodies, the latter produced by expressing forisome genes from different legumes in a heterologous background. The structure of these bodies was analysed using a combination of confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and the function of individual subunits was examined by combinatorial expression, micromanipulation and light microscopy. Key Results Dipteryx panamensis native forisomes and homomeric protein bodies assembled from the single sieve element occlusion by forisome (SEO-F) subunit identified in this species were structurally and functionally similar to forisomes from the Old World clade. In contrast, homomeric protein bodies assembled from individual SEO-F subunits from Old World species yielded artificial forisomes differing in proportion to their native counterparts, suggesting that multiple SEO-F proteins are required for forisome assembly in these plants. Structural differences between Medicago truncatula native forisomes, homomeric protein bodies and heteromeric bodies containing all possible subunit combinations suggested that combinations of SEO-F proteins may fine-tune the geometric proportions and reactivity of forisomes. Conclusions It is concluded that forisome structure and function have been strongly conserved during evolution and that species-dependent subsets of SEO-F proteins may have evolved to fine-tune the structure of native forisomes. PMID:24694827

  3. Protein dynamical transition at 110 K

    PubMed Central

    Kim, Chae Un; Tate, Mark W.; Gruner, Sol M.

    2011-01-01

    Proteins are known to undergo a dynamical transition at around 200 K but the underlying mechanism, physical origin, and relationship to water are controversial. Here we report an observation of a protein dynamical transition as low as 110 K. This unexpected protein dynamical transition precisely correlated with the cryogenic phase transition of water from a high-density amorphous to a low-density amorphous state. The results suggest that the cryogenic protein dynamical transition might be directly related to the two liquid forms of water proposed at cryogenic temperatures. PMID:22167801

  4. 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.

  5. Engineering Cells to Improve Protein Expression

    PubMed Central

    Xiao, Su; Shiloach, Joseph; Betenbaugh, Michael J.

    2014-01-01

    Cellular engineering of bacteria, fungi, insect cells and mammalian cells is a promising methodology to improve recombinant protein production for structural, biochemical, and commercial applications. Increased understanding of the host organism biology has suggested engineering strategies targeting bottlenecks in transcription, translation, protein processing and secretory pathways, as well as cell growth and survival. A combination of metabolic engineering and synthetic biology has been used to improve the properties of cells for protein production, which has resulted in enhanced yields of multiple protein classes. PMID:24704806

  6. Protein Interactions from Complexes: A Structural Perspective

    PubMed Central

    Hakes, Luke; Robertson, David L.; Oliver, Stephen G.; Lovell, Simon C.

    2007-01-01

    By combining crystallographic information with protein-interaction data obtained through traditional experimental means, this paper determines the most appropriate method for generating protein-interaction networks that incorporate data derived from protein complexes. We propose that a combined method should be considered; in which complexes composed of five chains or less are decomposed using the matrix model, whereas the spoke model is used to derive pairwise interactions for those with six chains or more. The results presented here should improve the accuracy and relevance of studies investigating the topology of protein-interaction networks. PMID:17538689

  7. Proximal protein tyrosine kinases in immunoreceptor signaling

    Microsoft Academic Search

    Sylvain Latour; André Veillette

    2001-01-01

    Immunoreceptor engagement results in the sequential activation of several classes of protein tyrosine kinases, including the Src and Syk\\/Zap-70 families. Recent progress has been made in our understanding of the regulation and function of these molecules. First, it was revealed that membrane compartmentation of protein tyrosine kinases may be essential for their proper biological function. Second, Src family kinases were

  8. Protein solubility: sequence based prediction and experimental

    Microsoft Academic Search

    Pawel Smialowski; Antonio J. Martin-Galiano; Aleksandra Mikolajka; Tobias Girschick; Tad A. Holak; Dmitrij Frishman

    Motivation: Obtaining soluble proteins in sufficient concentrations is a recurring limiting factor in various experimental studies. Solubility is an individual trait of proteins which, under a given set of experi- mental conditions, is determined by their amino acid sequence. Accurate theoretical prediction of solubility from sequence is instru- mental for setting priorities on targets in large-scale proteomics pro- jects. Results:

  9. In vitro protein refolding by chromatographic procedures

    Microsoft Academic Search

    Ming Li; Zhi-Guo Su; Jan-Christer Jansonb

    2004-01-01

    In vitro protein refolding is still a bottleneck in both structural biology and in the development of new biopharmaceuticals, especially for commercially important polypeptides that are overexpressed in Escherichia coli. This review focuses on protein refolding methods based on column procedures because recent advances in chromatographic refolding have shown promising results.

  10. Discovering functional interaction patterns in protein-protein interaction networks

    PubMed Central

    Turanalp, Mehmet E; Can, Tolga

    2008-01-01

    Background In recent years, a considerable amount of research effort has been directed to the analysis of biological networks with the availability of genome-scale networks of genes and/or proteins of an increasing number of organisms. A protein-protein interaction (PPI) network is a particular biological network which represents physical interactions between pairs of proteins of an organism. Major research on PPI networks has focused on understanding the topological organization of PPI networks, evolution of PPI networks and identification of conserved subnetworks across different species, discovery of modules of interaction, use of PPI networks for functional annotation of uncharacterized proteins, and improvement of the accuracy of currently available networks. Results In this article, we map known functional annotations of proteins onto a PPI network in order to identify frequently occurring interaction patterns in the functional space. We propose a new frequent pattern identification technique, PPISpan, adapted specifically for PPI networks from a well-known frequent subgraph identification method, gSpan. Existing module discovery techniques either look for specific clique-like highly interacting protein clusters or linear paths of interaction. However, our goal is different; instead of single clusters or pathways, we look for recurring functional interaction patterns in arbitrary topologies. We have applied PPISpan on PPI networks of Saccharomyces cerevisiae and identified a number of frequently occurring functional interaction patterns. Conclusion With the help of PPISpan, recurring functional interaction patterns in an organism's PPI network can be identified. Such an analysis offers a new perspective on the modular organization of PPI networks. The complete list of identified functional interaction patterns is available at . PMID:18547430

  11. Seo 312 Reitoria /Tcnico-Administrativos Local: Auditrio da Reitoria

    E-print Network

    Floeter, Sergio Ricardo

    VALENTIM BRAZ REITORIA-servidor 312 ALEXANDRE GAVA MENEZES REITORIA-servidor 312 ALEXANDRE PEDRO OLIVEIRA DIAS REITORIA-servidor 312 ANDREA FIGUEIREDO LEAO GRANTS REITORIA-servidor 312 ANDREIA ALVES DOS SANTOS-servidor 312 ARAQUIRI BOTELHO RODRIGUES REITORIA-servidor 312 ARI ABILIO DA SILVA REITORIA-servidor 312 ARI

  12. Seo 102 CCA/DOCENTES local : Hall do Prdio CCA

    E-print Network

    Floeter, Sergio Ricardo

    PEDRO LUIZ MANIQUE BARRETO CCA-professor 102 RENATA DIAS DE MELLO CASTANHO AMBONI CCA-professor 102 CCA-professor 102 CLARILTON EDZARD DAVOINE CARDOSO RIBAS CCA-professor 102 CLAUDIO MANOEL RODRIGUES DE MAURICIO SEDREZ DOS REIS CCA-professor 102 MIGUEL PEDRO GUERRA CCA-professor 102 MONICA YUMI TSUZUKI CCA

  13. Yoonjung Seo Phone: +1(347)287-7022

    E-print Network

    Operating Procedure) · Familiar to cGMP (Current Good Manufacturing Practice) and GLP (Good Laboratory Practice) regulations · Authorized to handle controlled substances such as Morphine Sulfate University

  14. Seo 107 CSE/DOCENTES Local : Hall do CSE

    E-print Network

    Floeter, Sergio Ricardo

    FACHINELLO CSE-professor 107 ARMANDO DE MELO LISBOA CSE-professor 107 ARNO DAL RI JUNIOR CSE-professor 107 BEATRIZ AUGUSTO DE PAIVA CSE-professor 107 BERNADETE LIMONGI CSE-professor 107 BRENA PAULA MAGNO FERNANDEZ

  15. updated 10/08/10 seo5 Graduate Student Activities

    E-print Network

    Yang, Sichun

    Chair's Asst 368.6252 kar18@case.edu SOM E653 Nutrition http://www.case.edu/med/nutrition/phd_nutrition, WRB5136 Blood Club (Cancer Center) Fridays, noon, BRB105 Neurodegeneration Journal Club Fridays, noon

  16. Seo 103 CCB/DOCENTES Local : Sala do Conselho -CCB

    E-print Network

    Floeter, Sergio Ricardo

    REICH CORSEUIL CCB-professor 103 ANGÉLICA FRANCESCA MARIS CCB-professor 103 ANICLETO POLI CCB-professor 103 ANTONIO CARLOS DE SOUZA CCB-professor 103 ANTONIO CARLOS GARDEL LEITAO CCB-professor 103 ANTONIO

  17. Increased protein intake in military special operations.

    PubMed

    Ferrando, Arny A

    2013-11-01

    Special operations are so designated for the specialized military missions they address. As a result, special operations present some unique metabolic challenges. In particular, soldiers often operate in a negative energy balance in stressful and demanding conditions with little opportunity for rest or recovery. In this framework, findings inferred from the performance literature suggest that increased protein intake may be beneficial. In particular, increased protein intake during negative caloric balance maintains lean body mass and blood glucose production. The addition of protein to mixed macronutrient supplements is beneficial for muscle endurance and power endpoints, and the use of amino acids improves gross and fine motor skills. Increasing protein intake during periods of intense training and/or metabolic demand improves subsequent performance, improves muscular recovery, and reduces symptoms of psychological stress. Consumption of protein before sleep confers the anabolic responses required for the maintenance of lean mass and muscle recovery. A maximal response in muscle protein synthesis is achieved with the consumption of 20-25 g of protein alone. However, higher protein intakes in the context of mixed-nutrient ingestion also confer anabolic benefits by reducing protein breakdown. Restricted rations issued to special operators provide less than the RDA for protein ( ? 0.6 g/kg), and these soldiers often rely on commercial products to augment their rations. The provision of reasonable alternatives and/or certification of approved supplements by the U.S. Department of Defense would be prudent. PMID:24027188

  18. Adjusting protein graphs based on graph entropy

    PubMed Central

    2014-01-01

    Measuring protein structural similarity attempts to establish a relationship of equivalence between polymer structures based on their conformations. In several recent studies, researchers have explored protein-graph remodeling, instead of looking a minimum superimposition for pairwise proteins. When graphs are used to represent structured objects, the problem of measuring object similarity become one of computing the similarity between graphs. Graph theory provides an alternative perspective as well as efficiency. Once a protein graph has been created, its structural stability must be verified. Therefore, a criterion is needed to determine if a protein graph can be used for structural comparison. In this paper, we propose a measurement for protein graph remodeling based on graph entropy. We extend the concept of graph entropy to determine whether a graph is suitable for representing a protein. The experimental results suggest that when applied, graph entropy helps a conformational on protein graph modeling. Furthermore, it indirectly contributes to protein structural comparison if a protein graph is solid. PMID:25474347

  19. Protein Binding: Do We Ever Learn??

    PubMed Central

    Zeitlinger, Markus A.; Derendorf, Hartmut; Mouton, Johan W.; Cars, Otto; Craig, William A.; Andes, David; Theuretzbacher, Ursula

    2011-01-01

    Although the influence of protein binding (PB) on antibacterial activity has been reported for many antibiotics and over many years, there is currently no standardization for pharmacodynamic models that account for the impact of protein binding of antimicrobial agents in vitro. This might explain the somewhat contradictory results obtained from different studies. Simple in vitro models which compare the MIC obtained in protein-free standard medium versus a protein-rich medium are prone to methodological pitfalls and may lead to flawed conclusions. Within in vitro test systems, a range of test conditions, including source of protein, concentration of the tested antibiotic, temperature, pH, electrolytes, and supplements may influence the impact of protein binding. As new antibiotics with a high degree of protein binding are in clinical development, attention and action directed toward the optimization and standardization of testing the impact of protein binding on the activity of antibiotics in vitro become even more urgent. In addition, the quantitative relationship between the effects of protein binding in vitro and in vivo needs to be established, since the physiological conditions differ. General recommendations for testing the impact of protein binding in vitro are suggested. PMID:21537013

  20. Bioengineering strategies to generate artificial protein complexes.

    PubMed

    Kim, Heejae; Siu, Ka-Hei; Raeeszadeh-Sarmazdeh, Maryam; Sun, Qing; Chen, Qi; Chen, Wilfred

    2015-08-01

    For many applications, increasing synergy between distinct proteins through organization is important for the specificity, regulation, and overall reaction efficiency. Although there are many examples of protein complexes in nature, a generalized method to create these complexes remains elusive. Many conventional techniques such as random chemical conjugation, physical adsorption onto surfaces, and encapsulation within matrices are imprecise approaches and can lead to deactivation of protein native functionalities. More "bio-friendly" approaches such as genetically fused proteins and biological scaffolds often can result in low yields and low complex stability. Alternatively, site-specific protein conjugation or ligation can generate artificial protein complexes that preserve the native functionalities of protein domains and maintain stability through covalent bonds. In this review, we describe three distinct methods to synthesize artificial protein complexes (genetic incorPoration of unnatural amino acids to introduce bio-orthogonal azide and alkyne groups to proteins, split-intein based expressed protein ligation, and sortase mediated ligation) and highlight interesting applications for each technique. Biotechnol. Bioeng. 2015;112: 1495-1505. © 2015 Wiley Periodicals, Inc. PMID:25943909

  1. Selenium inhibits LPS-induced pro-inflammatory gene expression by modulating MAPK and NF-?B signaling pathways in mouse mammary epithelial cells in primary culture.

    PubMed

    Zhang, Wen; Zhang, Runxiang; Wang, Tiancheng; Jiang, Haichao; Guo, Mengyao; Zhou, Ershun; Sun, Yong; Yang, Zhengtao; Xu, Shiwen; Cao, Yongguo; Zhang, Naisheng

    2014-04-01

    Mastitis is characterized by an inflammation of the mammary gland of dairy animals and humans; this condition is one of the major causes of economic losses in dairy industries. Selenium (Se), a biological trace element, modulates the functions of many regulatory proteins in signal transduction and provides advantages for animals with inflammatory diseases, including mastitis. The current study aimed to assess the protective effects and the active mechanism of Na(2)SeO(3) against lipopolysaccharide (LPS)-induced inflammation in mouse mammary epithelial cells (MMECs). Our results showed that LPS-induced expressions of cyclooxygenase-2 and tumor necrosis factor-? significantly decreased after Se was supplemented to Se-deficient MMECs. Na(2)SeO(3) also suppressed LPS-induced nuclear factor-?B activation, inhibitory kappa B degradation, and ERK, JNK, and P38 phosphorylation in a dose-dependent manner. These results suggested that Se functions as an anti-inflammatory agent in mastitis. PMID:24202549

  2. Localization of peroxisomal matrix proteins by photobleaching

    SciTech Connect

    Buch, Charlotta [Department of Biosciences and Nutrition, Karolinska Institutet, SE-141 57 Huddinge (Sweden) [Department of Biosciences and Nutrition, Karolinska Institutet, SE-141 57 Huddinge (Sweden); Soedertoerns University, Life Sciences, SE-141 89 Huddinge (Sweden); Hunt, Mary C. [Department of Genetics, Microbiology and Toxicology, Stockholm University, Arrhenius Laboratory, Svante Arrhenius vaeg 16F, SE-106 91 Stockholm (Sweden)] [Department of Genetics, Microbiology and Toxicology, Stockholm University, Arrhenius Laboratory, Svante Arrhenius vaeg 16F, SE-106 91 Stockholm (Sweden); Alexson, Stefan E.H. [Karolinska Institutet, Department of Laboratory Medicine, Division of Clinical Chemistry C1-74, Karolinska University Hospital at Huddinge, SE-141 86 Stockholm (Sweden)] [Karolinska Institutet, Department of Laboratory Medicine, Division of Clinical Chemistry C1-74, Karolinska University Hospital at Huddinge, SE-141 86 Stockholm (Sweden); Hallberg, Einar, E-mail: einar.hallberg@sh.se [Soedertoerns University, Life Sciences, SE-141 89 Huddinge (Sweden)] [Soedertoerns University, Life Sciences, SE-141 89 Huddinge (Sweden)

    2009-10-16

    The distribution of some enzymes between peroxisomes and cytosol, or a dual localization in both these compartments, can be difficult to reconcile. We have used photobleaching in live cells expressing green fluorescent protein (GFP)-fusion proteins to show that imported bona fide peroxisomal matrix proteins are retained in the peroxisome. The high mobility of the GFP-fusion proteins in the cytosol and absence of peroxisomal escape makes it possible to eliminate the cytosolic fluorescence by photobleaching, to distinguish between exclusively cytosolic proteins and proteins that are also present at low levels in peroxisomes. Using this technique we found that GFP tagged bile acid-CoA:amino acid N-acyltransferase (BAAT) was exclusively localized in the cytosol in HeLa cells. We conclude that the cytosolic localization was due to its carboxyterminal non-consensus peroxisomal targeting signal (-SQL) since mutation of the -SQL to -SKL resulted in BAAT being efficiently imported into peroxisomes.

  3. 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

  4. Preparation of Soluble Proteins from Escherichia coli.

    PubMed

    Wingfield, Paul T

    2014-01-01

    Purification of human IL-1? is used in this unit as an example of the preparation of a soluble protein from E. coli. Bacteria containing IL-1? are lysed, and IL-1 ? in the resulting supernatant is purified by anion-exchange chromatography, salt precipitation, and cation-exchange chromatography, and then concentrated. Finally, the IL-1 ? protein is applied to a gel-filtration column to separate it from remaining higher- and lower-molecular-weight contaminants, the purified protein is stored frozen or is lyophilized. The purification protocol described is typical for a protein that is expressed in fairly high abundance (i.e., >5% total protein) and accumulates in a soluble state. In addition, the purification procedure serves as an example of how to use classical protein purifications methods, which may also be used in conjunction with the affinity-based methods now more commonly used. © 2014 by John Wiley & Sons, Inc. PMID:25367009

  5. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2010 BINF 731 http://binf.gmu.edu/vaisman/binf731/ Protein sequence (helices, beta-sheets, turns) ·Tertiary - the three-dimensional fold of a protein subunit prediction Protein structure classification Protein representations Protein representations Protein

  6. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2009 BINF 731 http://binf.gmu.edu/vaisman/binf731/ Protein sequence (helices, beta-sheets, turns) ·Tertiary - the three-dimensional fold of a protein subunit prediction Protein structure classification Protein representations Protein representations Protein

  7. The Scope of Phage Display for Membrane Proteins Rosemarie Vithayathil 1

    E-print Network

    Weiss, Gregory A.

    ; protein engineering; mutagenesis; -barrel membrane proteins Numerous examples of phage display applied to soluble proteins demonstrate the power of the technique for protein engineering, affinity reagent and KO7+ phages. The results provide a guide for protein engineering and large-scale mutagenesis enabled

  8. Premature termination codon mutations in the type VII collagen gene in recessive dystrophic epidermolysis bullosa result in nonsense-mediated mRNA decay and absence of functional protein.

    PubMed

    Christiano, A M; Amano, S; Eichenfield, L F; Burgeson, R E; Uitto, J

    1997-09-01

    The severe mutilating Hallopeau-Siemens type of recessive dystrophic epidermolysis bullosa (HS-RDEB) is characterized by the absence of anchoring fibrils that consist of type VII collagen. We have previously identified premature termination codon (PTC) mutations in both alleles of the type VII collagen gene (COL7A1) in HS-RDEB patients. In this study we have defined the mechanism by which these mutations elicit their phenotypic consequences in a family. The extent of nonsense-mediated mRNA decay induced by these mutations was assessed by quantitation of the level of expression of the corresponding mRNA from each of the mutant alleles by RT-PCR of parental RNA. The level of expression of the paternal mutant allele with a PTC in exon 2 was approximately 30% of that of the wild-type allele whereas that of the maternal mutant allele with a PTC in exon 104 was reduced to about 80% of the normal allele. Immunoprecipitation of newly synthesized type VII collagen with a monoclonal antibody revealed reduced quantities of alpha1(VII) polypeptides in both parents' cells, whereas their synthesis was entirely absent in the proband's keratinocytes. Thus, a consequence of these premature termination codon mutations in COL7A1 is nonsense-mediated mRNA decay, with a dramatic reduction in type VII collagen synthesis, and the absence of anchoring fibrils in the proband. These results establish a mechanistic link between the presence of premature termination codon mutations in both alleles of COL7A1 and the clinical phenotype of HS-RDEB. PMID:9284110

  9. Regulation of Organelle Movement in Melanophores by Protein Kinase A (PKA), Protein Kinase C (PKC), and Protein Phosphatase 2A (PP2A)

    Microsoft Academic Search

    Amy R. Reilein; Irina S. Tint; Natalia I. Peunova; Grigori N. Enikolopov; Vladimir I. Gelfand

    1998-01-01

    We used melanophores, cells specialized for regulated organelle transport, to study signaling path- ways involved in the regulation of transport. We trans- fected immortalized Xenopus melanophores with plas- mids encoding epitope-tagged inhibitors of protein phosphatases and protein kinases or control plasmids encoding inactive analogues of these inhibitors. Expres- sion of a recombinant inhibitor of protein kinase A (PKA) results in

  10. Molecular dynamics and protein function

    PubMed Central

    Karplus, M.; Kuriyan, J.

    2005-01-01

    A fundamental appreciation for how biological macromolecules work requires knowledge of structure and dynamics. Molecular dynamics simulations provide powerful tools for the exploration of the conformational energy landscape accessible to these molecules, and the rapid increase in computational power coupled with improvements in methodology makes this an exciting time for the application of simulation to structural biology. In this Perspective we survey two areas, protein folding and enzymatic catalysis, in which simulations have contributed to a general understanding of mechanism. We also describe results for the F1 ATPase molecular motor and the Src family of signaling proteins as examples of applications of simulations to specific biological systems. PMID:15870208

  11. Protein content in diabetes nutrition plan.

    PubMed

    Hamdy, Osama; Horton, Edward S

    2011-04-01

    Medical nutrition therapy plays a major role in diabetes management. Macronutrient composition has been debated for a long time. However, there is increasing evidence that a modest increase in dietary protein intake above the current recommendation is a valid option toward better diabetes control, weight reduction, and improvement in blood pressure, lipid profile, and markers of inflammation. Increasing the absolute protein intake to 1.5-2 g/kg (or 20-30% of total caloric intake) during weight reduction has been suggested for overweight and obese patients with type 2 diabetes and normal kidney function. Increased protein intake does not increase plasma glucose, but increases the insulin response and results in a significant reduction in hemoglobin A(1c). In addition, a higher dietary protein intake reduces hunger, improves satiety, increases thermogenesis, and limits lean muscle mass loss during weight reduction using a reduced calorie diet and increased physical activity. It is preferable to calculate protein intake for patients with diabetes as grams per kilogram of body weight and not as a fixed percentage of total energy intake to avoid protein malnutrition when a hypocaloric diet is used. The relationship between protein intake as grams per kilogram of body weight and albumin excretion rate is very weak, except in hypertensive patients and particularly in those with uncontrolled diabetes. A protein intake of 0.8-1 g/kg should be recommended only for patients with diabetes and chronic kidney disease. Other patients with diabetes should not reduce protein intake to less than 1 g/kg of body weight. This review discusses the effects of different amounts of protein intake in a diabetes meal plan. It particular, it discusses the effects of protein intake on renal function, the effects of protein content on diabetes control, and the effects of increased dietary protein on body weight. PMID:21207203

  12. Protein Dynamism and Evolvability

    NSDL National Science Digital Library

    Nobuhiko Tokuriki (Weizmann Institute of Science; Department of Biological Chemistry,)

    2009-04-10

    The traditional view that proteins possess absolute functional specificity and a single, fixed structure conflicts with their marked ability to adapt and evolve new functions and structures. We consider an alternative, â??avant-garde viewâ?ť in which proteins are conformationally dynamic and exhibit functional promiscuity. We surmise that these properties are the foundation stones of protein evolvability; they facilitate the divergence of new functions within existing folds and the evolution of entirely new folds. Packing modes of proteins also affect their evolvability, and poorly packed, disordered, and conformationally diverse proteins may exhibit high evolvability. This dynamic view of protein structure, function, and evolvability is extrapolated to describe hypothetical scenarios for the evolution of the early proteins and future research directions in the area of protein dynamism and evolution.

  13. Structure-Based Druggability Assessment of the Mammalian Structural Proteome with Inclusion of Light Protein Flexibility

    PubMed Central

    Loving, Kathryn A.; Lin, Andy; Cheng, Alan C.

    2014-01-01

    Advances reported over the last few years and the increasing availability of protein crystal structure data have greatly improved structure-based druggability approaches. However, in practice, nearly all druggability estimation methods are applied to protein crystal structures as rigid proteins, with protein flexibility often not directly addressed. The inclusion of protein flexibility is important in correctly identifying the druggability of pockets that would be missed by methods based solely on the rigid crystal structure. These include cryptic pockets and flexible pockets often found at protein-protein interaction interfaces. Here, we apply an approach that uses protein modeling in concert with druggability estimation to account for light protein backbone movement and protein side-chain flexibility in protein binding sites. We assess the advantages and limitations of this approach on widely-used protein druggability sets. Applying the approach to all mammalian protein crystal structures in the PDB results in identification of 69 proteins with potential druggable cryptic pockets. PMID:25079060

  14. Sequence repeats and protein structure

    NASA Astrophysics Data System (ADS)

    Hoang, Trinh X.; Trovato, Antonio; Seno, Flavio; Banavar, Jayanth R.; Maritan, Amos

    2012-11-01

    Repeats are frequently found in known protein sequences. The level of sequence conservation in tandem repeats correlates with their propensities to be intrinsically disordered. We employ a coarse-grained model of a protein with a two-letter amino acid alphabet, hydrophobic (H) and polar (P), to examine the sequence-structure relationship in the realm of repeated sequences. A fraction of repeated sequences comprises a distinct class of bad folders, whose folding temperatures are much lower than those of random sequences. Imperfection in sequence repetition improves the folding properties of the bad folders while deteriorating those of the good folders. Our results may explain why nature has utilized repeated sequences for their versatility and especially to design functional proteins that are intrinsically unstructured at physiological temperatures.

  15. Neutron protein crystallography in JAERI

    NASA Astrophysics Data System (ADS)

    Tanaka, I.

    2004-07-01

    Neutron diffraction provides an experimental method of directly locating hy- drogen atoms in proteins. After developing an original neutron detector (neutron imaging plate) and a novel practical neutron monochromator (elastically bent perfect Si monochro- mator), BIX-type diffractometers which were equipped with these tools were e+/-ciently constructed at JRR-3 in Japan Atomic Energy Research Institute (JAERI), Japan and they have finished many protein crystallographic measurements and interesting results have come one after another. At the same time a method of growing large protein single crystals and a database of hydrogen and hydration have also been developed. In the near future, a pulsed neutron diffractometer for biological macromolecules has been proposed at J-PARC in JAERI.

  16. Proteins and Complexity

    NASA Astrophysics Data System (ADS)

    Murray, Joelle; Gibbon, Dana; Runyon, Alissa; Bajracharya, Arun

    2015-03-01

    A protein's tertiary structure determines its function in living organisms. The different functions proteins serve necessitate variety in native structures. How is variation in tertiary structure created from a common set of amino acids and molecular forces? In other words, what generates complexity in structures across all types of native proteins? To explore this question, a simple HP model of protein folding was explored for evidence of self-organized criticality, a potential generator of complexity.

  17. Protein Explorer Tutorial

    NSDL National Science Digital Library

    Protein Explorer is free software for visualizing the three-dimensional structures of protein, DNA, and RNA macromolecules, and their interactions and binding of ligands, inhibitors, and drugs. It is arguably the easiest-to-use software of its kind. It is suitable for high school and college students (ages 16 years and older), yet it is also widely used by graduate students and researchers. This site provides instruction in the use of Protein Explorer and simplifies queries of the Protein Data Bank.

  18. Represent tight Disheveled protein

    E-print Network

    Fig. 4.19 #12;Fig. 5.6 #12;#12;Represent tight junctions #12;Fig. 5.6 #12;Fig. 5.8 #12;Fig. 5.9 #12;Fig. 5.10 #12;Disheveled protein 16 Cell EmbryoOocyte Fig. 5.11 Normal ß-catenin protein Overexpressed ß-catenin protein Underexpressed ß-catenin protein #12;Eric Davidson, Caltech #12;Fig. 5.12 Pmar m

  19. Slow cooling of protein crystals

    PubMed Central

    Warkentin, Matthew; Thorne, Robert E.

    2009-01-01

    Cryoprotectant-free thaumatin crystals have been cooled from 300 to 100?K at a rate of 0.1?K?s?1 – 103–104 times slower than in conventional flash cooling – while continuously collecting X-ray diffraction data, so as to follow the evolution of protein lattice and solvent properties during cooling. Diffraction patterns show no evidence of crystalline ice at any temperature. This indicates that the lattice of protein molecules is itself an excellent cryoprotectant, and with sodium potassium tartrate incorporated from the 1.5?M mother liquor ice nucleation rates are at least as low as in a 70% glycerol solution. Crystal quality during slow cooling remains high, with an average mosaicity at 100?K of 0.2°. Most of the mosaicity increase occurs above ?200?K, where the solvent is still liquid, and is concurrent with an anisotropic contraction of the unit cell. Near 180?K a crossover to solid-like solvent behavior occurs, and on further cooling there is no additional degradation of crystal order. The variation of B factor with temperature shows clear evidence of a protein dynamical transition near 210?K, and at lower temperatures the slope dB/dT is a factor of 3–6 smaller than has been reported for any other protein. These results establish the feasibility of fully temperature controlled studies of protein structure and dynamics between 300 and 100?K. PMID:19798409

  20. Phylogenetic analysis of AAA proteins.

    PubMed

    Frickey, Tancred; Lupas, Andrei N

    2004-01-01

    AAA ATPases form a large protein family with manifold cellular roles. They belong to the AAA+ superfamily of ringshaped P-loop NTPases, which exert their activity through the energy-dependent unfolding of macromolecules. Phylogenetic analyses have suggested the existence of five major clades of AAA domains (proteasome subunits, metalloproteases, domains D1 and D2 of ATPases with two AAA domains, and the MSP1/katanin/spastin group), as well as a number of deeply branching minor clades. These analyses however have been characterized by a lack of consistency in defining the boundaries of the AAA family. We have used cluster analysis to delineate unambiguously the group of AAA sequences within the AAA+ superfamily. Phylogenetic and cluster analysis of this sequence set revealed the existence of a sixth major AAA clade, comprising the mitochondrial, membrane-bound protein BCS1 and its homologues. In addition, we identified several deep branches consisting mainly of hypothetical proteins resulting from genomic projects. Analysis of the AAA N-domains provided direct support for the obtained phylogeny for most branches, but revealed some deep splits that had not been apparent from phylogenetic analysis and some unexpected similarities between distant clades. It also revealed highly degenerate D1 domains in plant MSP1 sequences and in at least one deeply branching group of hypothetical proteins (YC46), showing that AAA proteins with two ATPase domains arose at least three times independently. PMID:15037233

  1. Compressive genomics for protein databases

    PubMed Central

    Daniels, Noah M.; Gallant, Andrew; Peng, Jian; Cowen, Lenore J.; Baym, Michael; Berger, Bonnie

    2013-01-01

    Motivation: The exponential growth of protein sequence databases has increasingly made the fundamental question of searching for homologs a computational bottleneck. The amount of unique data, however, is not growing nearly as fast; we can exploit this fact to greatly accelerate homology search. Acceleration of programs in the popular PSI/DELTA-BLAST family of tools will not only speed-up homology search directly but also the huge collection of other current programs that primarily interact with large protein databases via precisely these tools. Results: We introduce a suite of homology search tools, powered by compressively accelerated protein BLAST (CaBLASTP), which are significantly faster than and comparably accurate with all known state-of-the-art tools, including HHblits, DELTA-BLAST and PSI-BLAST. Further, our tools are implemented in a manner that allows direct substitution into existing analysis pipelines. The key idea is that we introduce a local similarity-based compression scheme that allows us to operate directly on the compressed data. Importantly, CaBLASTP’s runtime scales almost linearly in the amount of unique data, as opposed to current BLASTP variants, which scale linearly in the size of the full protein database being searched. Our compressive algorithms will speed-up many tasks, such as protein structure prediction and orthology mapping, which rely heavily on homology search. Availability: CaBLASTP is available under the GNU Public License at http://cablastp.csail.mit.edu/ Contact: bab@mit.edu PMID:23812995

  2. Combining random gene fission and rational gene fusion to discover near-infrared fluorescent protein fragments that report on protein-protein interactions.

    PubMed

    Pandey, Naresh; Nobles, Christopher L; Zechiedrich, Lynn; Maresso, Anthony W; Silberg, Jonathan J

    2015-05-15

    Gene fission can convert monomeric proteins into two-piece catalysts, reporters, and transcription factors for systems and synthetic biology. However, some proteins can be challenging to fragment without disrupting function, such as near-infrared fluorescent protein (IFP). We describe a directed evolution strategy that can overcome this challenge by randomly fragmenting proteins and concomitantly fusing the protein fragments to pairs of proteins or peptides that associate. We used this method to create libraries that express fragmented IFP as fusions to a pair of associating peptides (IAAL-E3 and IAAL-K3) and proteins (CheA and CheY) and screened for fragmented IFP with detectable near-infrared fluorescence. Thirteen novel fragmented IFPs were identified, all of which arose from backbone fission proximal to the interdomain linker. Either the IAAL-E3 and IAAL-K3 peptides or CheA and CheY proteins could assist with IFP fragment complementation, although the IAAL-E3 and IAAL-K3 peptides consistently yielded higher fluorescence. These results demonstrate how random gene fission can be coupled to rational gene fusion to create libraries enriched in fragmented proteins with AND gate logic that is dependent upon a protein-protein interaction, and they suggest that these near-infrared fluorescent protein fragments will be suitable as reporters for pairs of promoters and protein-protein interactions within whole animals. PMID:25265085

  3. Comparative transcriptional analysis of Bacillus subtilis cells overproducing either secreted proteins, lipoproteins or membrane proteins

    PubMed Central

    2012-01-01

    Background Bacillus subtilis is a favorable host for the production of industrially relevant proteins because of its capacity of secreting proteins into the medium to high levels, its GRAS (Generally Recognized As Safe) status, its genetic accessibility and its capacity to grow in large fermentations. However, production of heterologous proteins still faces limitations. Results This study aimed at the identification of bottlenecks in secretory protein production by analyzing the response of B. subtilis at the transcriptome level to overproduction of eight secretory proteins of endogenous and heterologous origin and with different subcellular or extracellular destination: secreted proteins (NprE and XynA of B. subtilis, Usp45 of Lactococcus lactis, TEM-1 ?-lactamase of Escherichia coli), membrane proteins (LmrA of L. lactis and XylP of Lactobacillus pentosus) and lipoproteins (MntA and YcdH of B. subtilis). Responses specific for proteins with a common localization as well as more general stress responses were observed. The latter include upregulation of genes encoding intracellular stress proteins (groES/EL, CtsR regulated genes). Specific responses include upregulation of the liaIHGFSR operon under Usp45 and TEM-1 ?-lactamase overproduction; cssRS, htrA and htrB under all secreted proteins overproduction; sigW and SigW-regulated genes mainly under membrane proteins overproduction; and ykrL (encoding an HtpX homologue) specifically under membrane proteins overproduction. Conclusions The results give better insights into B. subtilis responses to protein overproduction stress and provide potential targets for genetic engineering in order to further improve B. subtilis as a protein production host. PMID:22624725

  4. UMBCUMBCUMBC Protein Image Alignment

    E-print Network

    Potra, Florian

    . Gels must be aligned before comparison. Bottleneck: Gel Alignment Protein Image Alignment via Quadratic (Compugen) All are deficient on gel alignment Protein Image Alignment via Quadratic Programming ­ p.3/47 #12 (2) , I (3) , I (4) Aligned gels Protein Image Alignment via Quadratic Programming ­ p.5/47 #12

  5. Protein folding and misfolding

    Microsoft Academic Search

    Christopher M. Dobson

    2003-01-01

    The manner in which a newly synthesized chain of amino acids transforms itself into a perfectly folded protein depends both on the intrinsic properties of the amino-acid sequence and on multiple contributing influences from the crowded cellular milieu. Folding and unfolding are crucial ways of regulating biological activity and targeting proteins to different cellular locations. Aggregation of misfolded proteins that

  6. Protein engineering of lantibiotics

    Microsoft Academic Search

    Oscar P. Kuipers; Gabriele Bierbaum; Birgit Ottenwälder; Helen M. Dodd; Nicky Horn; Jörg Metzger; Thomas Kupke; Volker Gnau; Roger Bongers; Patrick van den Bogaard; Hans Kosters; Harry S. Rollema; Willem M. de Vos; Roland J. Siezen; Günther Jung; Friedrich Götz; Hans-Georg Sahl; Michael J. Gasson

    1996-01-01

    Whereas protein engineering of enzymes and structural proteins nowadays is an established research tool for studying structure-function relationships of polypeptides and for improving their properties, the engineering of posttranslationally modified peptides, such as the lantibiotics, is just coming of age. The engineering of lantibiotics is less straightforward than that of unmodified proteins, since expression systems should be developed not only

  7. Protein Preservation BIOMATERIALS

    E-print Network

    Protein Preservation BIOMATERIALS Our goal is to develop measurements for characterizing sugar-based glasses with respect to their ability to serve as preservation media for therapeutic proteins to the stability of proteins in stabilizing sugar-based glass. This property had been completely overlooked

  8. Creating functional artificial proteins.

    PubMed

    Razeghifard, Reza; Wallace, Brett B; Pace, Ron J; Wydrzynski, Tom

    2007-02-01

    Much is now known about how protein folding occurs, through the sequence analysis of proteins of known folding geometry and the sequence/structural analysis of proteins and their mutants. This has allowed not only the modification of natural proteins but also the construction of de novo polypeptides with predictable folding patterns. Structure/function analysis of natural proteins is used to construct derived versions that retain a degree of biological activity. The constructed versions made of either natural or artificial sequences contain critical residues for activity such as receptor binding. In some cases, the functionality is introduced by incorporating binding sites for other elements, such as organic cofactors or transition metals, into the protein scaffold. While these modified proteins can mimic the function of natural proteins, they can also be constructed to have novel activities. Recently engineered photoactive proteins are good examples of such systems in which a light-induced electron transfer can be established in normally light-insensitive proteins. The present review covers some aspects of protein design that have been used to investigate protein receptor binding, cofactor binding and biological electron transfer. PMID:17305556

  9. Destabilized bioluminescent proteins

    DOEpatents

    Allen, Michael S. (Knoxville, TN); Rakesh, Gupta (New Delhi, IN); Gary, Sayler S. (Blaine, TN)

    2007-07-31

    Purified nucleic acids, vectors and cells containing a gene cassette encoding at least one modified bioluminescent protein, wherein the modification includes the addition of a peptide sequence. The duration of bioluminescence emitted by the modified bioluminescent protein is shorter than the duration of bioluminescence emitted by an unmodified form of the bioluminescent protein.

  10. IGSF9 family proteins.

    PubMed

    Hansen, Maria; Walmod, Peter Schledermann

    2013-06-01

    The Drosophila protein Turtle and the vertebrate proteins immunoglobulin superfamily (IgSF), member 9 (IGSF9/Dasm1) and IGSF9B are members of an evolutionarily ancient protein family. A bioinformatics analysis of the protein family revealed that invertebrates contain only a single IGSF9 family gene, whereas vertebrates contain two to four genes. In cnidarians, the gene appears to encode a secreted protein, but transmembrane isoforms of the protein have also evolved, and in many species, alternative splicing facilitates the expression of both transmembrane and secreted isoforms. In most species, the longest isoforms of the proteins have the same general organization as the neural cell adhesion molecule family of cell adhesion molecule proteins, and like this family of proteins, IGSF9 family members are expressed in the nervous system. A review of the literature revealed that Drosophila Turtle facilitates homophilic cell adhesion. Moreover, IGSF9 family proteins have been implicated in the outgrowth and branching of neurites, axon guidance, synapse maturation, self-avoidance, and tiling. However, despite the few published studies on IGSF9 family proteins, reports on the functions of both Turtle and mammalian IGSF9 proteins are contradictory. PMID:23417431

  11. Protein purification and preparation.

    E-print Network

    Lebendiker, Mario

    Protein purification and preparation. High purity and recovery for better discovery. #12 discovery is only a step away. From protein isolation to purification, you can count on Merck Millipore. Scale-up compatibility It's easy to scale up to high-throughput recombinant protein purification

  12. Protein crystallization Mirjam Leunissen

    E-print Network

    Leunissen, Mirjam

    in the pharmaceutical industry. In this report, an overview of the unique physical-chemical characteristics of protein of proteins? While, in the past, purification of mixtures was the main goal in crystallizing experimentsProtein crystallization Mirjam Leunissen #12;Mirjam Leunissen Department of solid state chemistry

  13. Protein Expression Xpress System

    E-print Network

    Lebendiker, Mario

    and Elution Under Denaturing Conditions..................................8 4.6 Cleavage Of The Fusion PeptideXpressŞ System Protein Expression pEBVHis Version D 180208 25-0042 XpressŞ System Protein Expression pEBVHis A Manual of Methods for Expression of Polyhistidine - Containing Recombinant Proteins

  14. Energetics of Protein Folding

    Microsoft Academic Search

    Robert L. Baldwin

    2007-01-01

    The energetics of protein folding determine the 3D structure of a folded protein. Knowledge of the energetics is needed to predict the 3D structure from the amino acid sequence or to modify the structure by protein engineering. Recent developments are discussed: major factors are reviewed and auxiliary factors are discussed briefly. Major factors include the hydrophobic factor (burial of non-polar

  15. Nanoelectrochemical Immunosensors for Protein Detection

    NASA Astrophysics Data System (ADS)

    Carpentiero, Alessandro; de Leo, Manuela; Garcia Romero, Ivan; Pozzi Mucelli, Stefano; Reuther, Freimut; Stanta, Giorgio; Tormen, Massimo; Ugo, Paolo; Zamuner, Martina

    Nanoelectrochemical immunosensors fabricated by templated electrodeposition of gold nanoelectrodes inside the pores of polycarbonate (PC) track-etched membranes, followed by the immobilization of the biorecognition elements on the surrounding PC, have proven high sensitivity and specificity for protein detection. The signal transduction scheme involves a suitable redox mediator added to the sample solution to shuttle electrons from the gold nanoelectrodes to the biorecognition layer, both elements being in strict spatial proximity. Highly improved signal-to-background current ratio, which are peculiar of NEEs with respect to other electrochemical transducers, can be exploited in this way. Two detection schemes were tested: one based on the direct immobilization of the target protein on the PC of the NEE (approach A) and the other based on the immobilisation on PC of an antibody to capture the target protein (approach B). The biorecognition process was completed by adding a primary antibody and a secondary antibody with horse radish peroxidase (HRP) as enzyme label; methylene blue was the redox mediator added to the electrolyte solution. Typical target analytes were single chain fragment variable proteins, for approach A, and trastuzumab (also known as Herceptin®), for approach B. NEE-based capture sensors were tested successfully to detect small amounts of the receptor protein HER2 in biological samples. Finally, motivated by the target of a better control of the geometrical characteristics of ensembles of nanoelectrodes (size, density, geometrical arrangement, and degree of recession), and by the positive results obtained with track-etch membranes of PC from the standpoint of protein immobilization, we demonstrated the fabrication of nanobiosensors by patterning ordered arrays of nanoelectrodes (NEAs) by electron beam lithography (EBL) on polycarbonate. EBL results perfectly suitable for the top-down fabrication of arrays of nanobiosensors on thin PC films deposited on gold coated silicon.

  16. Protein-spanning water networks and implications for prediction of protein-protein interactions mediated through hydrophobic effects.

    PubMed

    Cui, Di; Ou, Shuching; Patel, Sandeep

    2014-12-01

    Hydrophobic effects, often conflated with hydrophobic forces, are implicated as major determinants in biological association and self-assembly processes. Protein-protein interactions involved in signaling pathways in living systems are a prime example where hydrophobic effects have profound implications. In the context of protein-protein interactions, a priori knowledge of relevant binding interfaces (i.e., clusters of residues involved directly with binding interactions) is difficult. In the case of hydrophobically mediated interactions, use of hydropathy-based methods relying on single residue hydrophobicity properties are routinely and widely used to predict propensities for such residues to be present in hydrophobic interfaces. However, recent studies suggest that consideration of hydrophobicity for single residues on a protein surface require accounting of the local environment dictated by neighboring residues and local water. In this study, we use a method derived from percolation theory to evaluate spanning water networks in the first hydration shells of a series of small proteins. We use residue-based water density and single-linkage clustering methods to predict hydrophobic regions of proteins; these regions are putatively involved in binding interactions. We find that this simple method is able to predict with sufficient accuracy and coverage the binding interface residues of a series of proteins. The approach is competitive with automated servers. The results of this study highlight the importance of accounting of local environment in determining the hydrophobic nature of individual residues on protein surfaces. PMID:25204743

  17. Protein – Which is Best?

    PubMed Central

    Hoffman, Jay R.; Falvo, Michael J.

    2004-01-01

    Protein intake that exceeds the recommended daily allowance is widely accepted for both endurance and power athletes. However, considering the variety of proteins that are available much less is known concerning the benefits of consuming one protein versus another. The purpose of this paper is to identify and analyze key factors in order to make responsible recommendations to both the general and athletic populations. Evaluation of a protein is fundamental in determining its appropriateness in the human diet. Proteins that are of inferior content and digestibility are important to recognize and restrict or limit in the diet. Similarly, such knowledge will provide an ability to identify proteins that provide the greatest benefit and should be consumed. The various techniques utilized to rate protein will be discussed. Traditionally, sources of dietary protein are seen as either being of animal or vegetable origin. Animal sources provide a complete source of protein (i.e. containing all essential amino acids), whereas vegetable sources generally lack one or more of the essential amino acids. Animal sources of dietary protein, despite providing a complete protein and numerous vitamins and minerals, have some health professionals concerned about the amount of saturated fat common in these foods compared to vegetable sources. The advent of processing techniques has shifted some of this attention and ignited the sports supplement marketplace with derivative products such as whey, casein and soy. Individually, these products vary in quality and applicability to certain populations. The benefits that these particular proteins possess are discussed. In addition, the impact that elevated protein consumption has on health and safety issues (i.e. bone health, renal function) are also reviewed. Key Points Higher protein needs are seen in athletic populations. Animal proteins is an important source of protein, however potential health concerns do exist from a diet of protein consumed from primarily animal sources. With a proper combination of sources, vegetable proteins may provide similar benefits as protein from animal sources. Casein protein supplementation may provide the greatest benefit for increases in protein synthesis for a prolonged duration. PMID:24482589

  18. Simultaneous dual protein labeling using a triorthogonal reagent.

    PubMed

    Rashidian, Mohammad; Kumarapperuma, Sidath C; Gabrielse, Kari; Fegan, Adrian; Wagner, Carston R; Distefano, Mark D

    2013-11-01

    Construction of heterofunctional proteins is a rapidly emerging area of biotherapeutics. Combining a protein with other moieties, such as a targeting element, a toxic protein or small molecule, and a fluorophore or polyethylene glycol (PEG) group, can improve the specificity, functionality, potency, and pharmacokinetic profile of a protein. Protein farnesyl transferase (PFTase) is able to site-specifically and quantitatively prenylate proteins containing a C-terminal CaaX-box amino acid sequence with various modified isoprenoids. Here, we describe the design, synthesis, and application of a triorthogonal reagent, 1, that can be used to site-specifically incorporate an alkyne and aldehyde group simultaneously into a protein. To illustrate the capabilities of this approach, a protein was enzymatically modified with compound 1 followed by oxime ligation and click reaction to simultaneously incorporate an azido-tetramethylrhodamine (TAMRA) fluorophore and an aminooxy-PEG moiety. This was performed with both a model protein [green fluorescent protein (GFP)] as well as a therapeutically useful protein [ciliary neurotrophic factor (CNTF)]. Next, a protein was enzymatically modified with compound 1 followed by coupling to an azido-bis-methotrexate dimerizer and aminooxy-TAMRA. Incubation of that construct with a dihydrofolate reductase (DHFR)-DHFR-anti-CD3 fusion protein resulted in the self-assembly of nanoring structures that were endocytosed into T-leukemia cells and visualized therein. These results highlight how complex multifunctional protein assemblies can be prepared using this facile triorthogonal approach. PMID:24134212

  19. Structure and Protein-Protein Interaction Studies on Chlamydia trachomatis Protein CT670 (YscO Homolog)

    SciTech Connect

    Lorenzini, Emily; Singer, Alexander; Singh, Bhag; Lam, Robert; Skarina, Tatiana; Chirgadze, Nickolay Y.; Savchenko, Alexei; Gupta, Radhey S. (Toronto); (McMaster U.); (OCI)

    2010-07-28

    Comparative genomic studies have identified many proteins that are found only in various Chlamydiae species and exhibit no significant sequence similarity to any protein in organisms that do not belong to this group. The CT670 protein of Chlamydia trachomatis is one of the proteins whose genes are in one of the type III secretion gene clusters but whose cellular functions are not known. CT670 shares several characteristics with the YscO protein of Yersinia pestis, including the neighboring genes, size, charge, and secondary structure, but the structures and/or functions of these proteins remain to be determined. Although a BLAST search with CT670 did not identify YscO as a related protein, our analysis indicated that these two proteins exhibit significant sequence similarity. In this paper, we report that the CT670 crystal, solved at a resolution of 2 {angstrom}, consists of a single coiled coil containing just two long helices. Gel filtration and analytical ultracentrifugation studies showed that in solution CT670 exists in both monomeric and dimeric forms and that the monomer predominates at lower protein concentrations. We examined the interaction of CT670 with many type III secretion system-related proteins (viz., CT091, CT665, CT666, CT667, CT668, CT669, CT671, CT672, and CT673) by performing bacterial two-hybrid assays. In these experiments, CT670 was found to interact only with the CT671 protein (YscP homolog), whose gene is immediately downstream of ct670. A specific interaction between CT670 and CT671 was also observed when affinity chromatography pull-down experiments were performed. These results suggest that CT670 and CT671 are putative homologs of the YcoO and YscP proteins, respectively, and that they likely form a chaperone-effector pair.

  20. From Topology to Phenotype in Protein-Protein Interaction Networks

    NASA Astrophysics Data System (ADS)

    Pržulj, Nataša

    We have recently witnessed an explosion in biological network data along with the development of computational approaches for their analyses. This new interdisciplinary research area is an integral part of systems biology, promising to provide new insights into organizational principles of life, as well as into evolution and disease. However, there is a danger that the area might become hindered by several emerging issues. In particular, there is typically a weak link between biological and computational scientists, resulting in the use of simple computational techniques of limited potential to explain these complex biological data. Hence, there is a danger that the community might view the topological features of network data as mere statistics, ignoring the value of the information contained in these data. This might result in the imposition of scientific doctrines, such as scale-free-centric (on the modelling side) and genome-centric (on the biological side) opinions onto this nascent research area. In this chapter, we take a network science perspective and present a brief, high-level overview of the area, commenting on possible challenges ahead. We focus on protein-protein interaction networks (PINs) in which nodes correspond to proteins in a cell and edges to physical bindings between the proteins.

  1. A Second-generation Protein–Protein Interaction Network of Helicobacter pylori*

    PubMed Central

    Häuser, Roman; Ceol, Arnaud; Rajagopala, Seesandra V.; Mosca, Roberto; Siszler, Gabriella; Wermke, Nadja; Sikorski, Patricia; Schwarz, Frank; Schick, Matthias; Wuchty, Stefan; Aloy, Patrick; Uetz, Peter

    2014-01-01

    Helicobacter pylori infections cause gastric ulcers and play a major role in the development of gastric cancer. In 2001, the first protein interactome was published for this species, revealing over 1500 binary protein interactions resulting from 261 yeast two-hybrid screens. Here we roughly double the number of previously published interactions using an ORFeome-based, proteome-wide yeast two-hybrid screening strategy. We identified a total of 1515 protein–protein interactions, of which 1461 are new. The integration of all the interactions reported in H. pylori results in 3004 unique interactions that connect about 70% of its proteome. Excluding interactions of promiscuous proteins we derived from our new data a core network consisting of 908 interactions. We compared our data set to several other bacterial interactomes and experimentally benchmarked the conservation of interactions using 365 protein pairs (interologs) of E. coli of which one third turned out to be conserved in both species. PMID:24627523

  2. The vaccinia virus E6 protein influences virion protein localization during virus assembly.

    PubMed

    Condit, Richard C; Moussatche, Nissin

    2015-08-01

    Vaccinia virus mutants in which expression of the virion core protein gene E6R is repressed are defective in virion morphogenesis. E6 deficient infections fail to properly package viroplasm into viral membranes, resulting in an accumulation of empty immature virions and large aggregates of viroplasm. We have used immunogold electron microscopy and immunofluorescence confocal microscopy to assess the intracellular localization of several virion structural proteins and enzymes during E6R mutant infections. We find that during E6R mutant infections virion membrane proteins and virion transcription enzymes maintain a normal localization within viral factories while several major core and lateral body proteins accumulate in aggregated virosomes. The results support a model in which vaccinia virions are assembled from at least three substructures, the membrane, the viroplasm and a "pre-nucleocapsid", and that the E6 protein is essential for maintaining proper localization of the seven-protein complex and the viroplasm during assembly. PMID:25863879

  3. A second-generation protein-protein interaction network of Helicobacter pylori.

    PubMed

    Häuser, Roman; Ceol, Arnaud; Rajagopala, Seesandra V; Mosca, Roberto; Siszler, Gabriella; Wermke, Nadja; Sikorski, Patricia; Schwarz, Frank; Schick, Matthias; Wuchty, Stefan; Aloy, Patrick; Uetz, Peter

    2014-05-01

    Helicobacter pylori infections cause gastric ulcers and play a major role in the development of gastric cancer. In 2001, the first protein interactome was published for this species, revealing over 1500 binary protein interactions resulting from 261 yeast two-hybrid screens. Here we roughly double the number of previously published interactions using an ORFeome-based, proteome-wide yeast two-hybrid screening strategy. We identified a total of 1515 protein-protein interactions, of which 1461 are new. The integration of all the interactions reported in H. pylori results in 3004 unique interactions that connect about 70% of its proteome. Excluding interactions of promiscuous proteins we derived from our new data a core network consisting of 908 interactions. We compared our data set to several other bacterial interactomes and experimentally benchmarked the conservation of interactions using 365 protein pairs (interologs) of E. coli of which one third turned out to be conserved in both species. PMID:24627523

  4. Statistically Inferring Protein-Protein Asociations with Affinity Isolation LC-MS/MS Assays

    SciTech Connect

    Sharp, Julia L.; Anderson, Kevin K.; Hurst, G. B.; Daly, Don S.; Pelletier, Dale A.; Cannon, William R.; Auberry, Deanna L.; Schmoyer, Denise D.; McDonald, W. Hayes; White, Amanda M.; Hooker, Brian S.; Victry, Kristin D.; Buchanan, M. V.; Kery, Vladimir; Wiley, H. S.

    2007-09-30

    Affinity isolation of protein complexes followed by protein identification by LC-MS/MS is an increasingly popular approach for mapping protein interactions. However, systematic and random assay errors from multiple sources must be considered to confidently infer authentic protein-protein interactions. To address this issue, we developed a general, robust statistical method for inferring authentic interactions from protein prey-by-bait frequency tables using a binomial-based likelihood ratio test (LRT) coupled with Bayes’ Odds estimation. We then applied our LRT-Bayes’ algorithm experimentally using data from protein complexes isolated from Rhodopseudomonas palustris. Our algorithm, in conjunction with the experimental protocol, inferred with high confidence authentic interacting proteins from abundant, stable complexes, but few or no authentic interactions for lower-abundance complexes. The algorithm can discriminate against a background of prey proteins that are detected in association with a large number of baits as an artifact of the measurement. We conclude that the experimental protocol including the LRT-Bayes’ algorithm produces results with high confidence but moderate sensitivity. We also found that Monte Carlo simulation is a feasible tool for checking modeling assumptions, estimating parameters, and evaluating the significance of results in protein association studies.

  5. Thermodynamic study of protein phases formation and clustering in model water-protein-salt solutions.

    PubMed

    Rozhkov, Sergey P; Goryunov, Andrey S

    2010-09-01

    Thermodynamic analysis of the water-protein-salt system, based on the description of the spinodal curve, has been carried out in various coordinate systems: (water chemical potential, protein concentration m(2)); (protein "solubility" log S, salt concentration m(3)); (effective temperature, critical composition of the system m(2)/m(3)). Such presentations explain the existence of diagrams with normal and retrograde protein solubility as a result of straightforward effect of ions present in solution as well as some features of the widely used phase diagram in coordinates (temperature, protein concentration). Analytic expressions for coefficients K and b of the salting out equation log S=-K.m(3)+b as functions of protein charge and protein adsorbed ions have been obtained and identified with the spinodal characteristic points reflecting quasi-equilibrium between protein-lean phase and dense protein-rich phase. Liquid-liquid, liquid-solid phase transitions, dynamic protein clusters and second virial coefficient that characterize interaction between solution components have been thus interrelated. The results of our thermodynamic analysis have been compared with the data reported for lysozyme . PMID:20494508

  6. ComPPI: a cellular compartment-specific database for protein-protein interaction network analysis.

    PubMed

    Veres, Daniel V; Gyurkó, Dávid M; Thaler, Benedek; Szalay, Kristóf Z; Fazekas, Dávid; Korcsmáros, Tamás; Csermely, Peter

    2015-01-01

    Here we present ComPPI, a cellular compartment-specific database of proteins and their interactions enabling an extensive, compartmentalized protein-protein interaction network analysis (URL: http://ComPPI.LinkGroup.hu). ComPPI enables the user to filter biologically unlikely interactions, where the two interacting proteins have no common subcellular localizations and to predict novel properties, such as compartment-specific biological functions. ComPPI is an integrated database covering four species (S. cerevisiae, C. elegans, D. melanogaster and H. sapiens). The compilation of nine protein-protein interaction and eight subcellular localization data sets had four curation steps including a manually built, comprehensive hierarchical structure of >1600 subcellular localizations. ComPPI provides confidence scores for protein subcellular localizations and protein-protein interactions. ComPPI has user-friendly search options for individual proteins giving their subcellular localization, their interactions and the likelihood of their interactions considering the subcellular localization of their interacting partners. Download options of search results, whole-proteomes, organelle-specific interactomes and subcellular localization data are available on its website. Due to its novel features, ComPPI is useful for the analysis of experimental results in biochemistry and molecular biology, as well as for proteome-wide studies in bioinformatics and network science helping cellular biology, medicine and drug design. PMID:25348397

  7. Characterization of essential proteins based on network topology in proteins interaction networks

    NASA Astrophysics Data System (ADS)

    Bakar, Sakhinah Abu; Taheri, Javid; Zomaya, Albert Y.

    2014-06-01

    The identification of essential proteins is theoretically and practically important as (1) it is essential to understand the minimal surviving requirements for cellular lives, and (2) it provides fundamental for development of drug. As conducting experimental studies to identify essential proteins are both time and resource consuming, here we present a computational approach in predicting them based on network topology properties from protein-protein interaction networks of Saccharomyces cerevisiae. The proposed method, namely EP3NN (Essential Proteins Prediction using Probabilistic Neural Network) employed a machine learning algorithm called Probabilistic Neural Network as a classifier to identify essential proteins of the organism of interest; it uses degree centrality, closeness centrality, local assortativity and local clustering coefficient of each protein in the network for such predictions. Results show that EP3NN managed to successfully predict essential proteins with an accuracy of 95% for our studied organism. Results also show that most of the essential proteins are close to other proteins, have assortativity behavior and form clusters/sub-graph in the network.

  8. ComPPI: a cellular compartment-specific database for protein–protein interaction network analysis

    PubMed Central

    Veres, Daniel V.; Gyurkó, Dávid M.; Thaler, Benedek; Szalay, Kristóf Z.; Fazekas, Dávid; Korcsmáros, Tamás; Csermely, Peter

    2015-01-01

    Here we present ComPPI, a cellular compartment-specific database of proteins and their interactions enabling an extensive, compartmentalized protein–protein interaction network analysis (URL: http://ComPPI.LinkGroup.hu). ComPPI enables the user to filter biologically unlikely interactions, where the two interacting proteins have no common subcellular localizations and to predict novel properties, such as compartment-specific biological functions. ComPPI is an integrated database covering four species (S. cerevisiae, C. elegans, D. melanogaster and H. sapiens). The compilation of nine protein–protein interaction and eight subcellular localization data sets had four curation steps including a manually built, comprehensive hierarchical structure of >1600 subcellular localizations. ComPPI provides confidence scores for protein subcellular localizations and protein–protein interactions. ComPPI has user-friendly search options for individual proteins giving their subcellular localization, their interactions and the likelihood of their interactions considering the subcellular localization of their interacting partners. Download options of search results, whole-proteomes, organelle-specific interactomes and subcellular localization data are available on its website. Due to its novel features, ComPPI is useful for the analysis of experimental results in biochemistry and molecular biology, as well as for proteome-wide studies in bioinformatics and network science helping cellular biology, medicine and drug design. PMID:25348397

  9. Improving objectivity and scalability in protein crystallization

    Microsoft Academic Search

    Igor Jurisica; Patrick Rogers; Janice I. Glasgow; Robert J. Collins; Jennifer R. Wolfley; Joseph R. Luft; George T. DeTitta

    2001-01-01

    This article describes the application of image analysis techniques to protein crystallization experiment classification. By applying knowledge discovery techniques to the analysis results, we can extract important crystallographic knowledge.

  10. BIOCHEMISTRY: Metamorphic Proteins

    NSDL National Science Digital Library

    Alexey G. Murzin (MRC Centre for Protein Engineering; )

    2008-06-27

    Access to the article is free, however registration and sign-in are required. Proteins that can adopt more than one native folded conformation may be more common than previously thought. A small but growing number of "metamorphic" proteins adopt different folded conformations for the same amino acid sequence in native conditions. Unlike prions, they undergo reversible conformational changes. Most of these proteins have been treated as special cases, because the transitions between their alternative conformations are triggered by environmental factors. The discoveries of a new metamorphic protein capable of independent interconversion (1) and of an abrupt fold change in a protein lineage (2) point to a more general nature of this phenomenon.

  11. Highly thermostable fluorescent proteins

    DOEpatents

    Bradbury, Andrew M. (Santa Fe, NM); Waldo, Geoffrey S. (Santa Fe, NM); Kiss, Csaba (Los Alamos, NM)

    2011-11-29

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  12. Highly thermostable fluorescent proteins

    DOEpatents

    Bradbury, Andrew M. (Santa Fe, NM); Waldo, Geoffrey S. (Santa Fe, NM); Kiss, Csaba (Los Alamos, NM)

    2012-05-01

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  13. Highly thermostable fluorescent proteins

    DOEpatents

    Bradbury, Andrew M. (Santa Fe, NM); Waldo, Geoffrey S. (Santa Fe, NM); Kiss, Csaba (Los Alamos, NM)

    2011-03-22

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  14. Prediction of functions for two LEA proteins from mung bean.

    PubMed

    Rajesh, Subramanian; Manickam, Ayyanar

    2006-01-01

    LEA (late embryogenesis abundant) proteins are associated with tolerance to water stress resulting from desiccation and cold shock. Although various functions have been proposed to LEA proteins, their precise role is not fully defined. In silico analysis of the amino acid sequence of two LEA proteins (early methionine-labeled Vigna, EMV) from the tropical legume crop, Vigna radiata identified a 20 residues motif 'GGQTRKQQLGSEGYHEMGRK' characteristic to group 1 LEA proteins. Structural analyses hypothesize these proteins to function like DNA/RNA binding proteins in protecting macromolecules/ membrane stabilization at the time of dehydration process. PMID:17597874

  15. Prediction of functions for two LEA proteins from mung bean

    PubMed Central

    Rajesh, Subramanian; Manickam, Ayyanar

    2006-01-01

    LEA (late embryogenesis abundant) proteins are associated with tolerance to water stress resulting from desiccation and cold shock. Although various functions have been proposed to LEA proteins, their precise role is not fully defined. In silico analysis of the amino acid sequence of two LEA proteins (early methionine-labeled Vigna, EMV) from the tropical legume crop, Vigna radiata identified a 20 residues motif ‘GGQTRKQQLGSEGYHEMGRK’ characteristic to group 1 LEA proteins. Structural analyses hypothesize these proteins to function like DNA/RNA binding proteins in protecting macromolecules/ membrane stabilization at the time of dehydration process. PMID:17597874

  16. Protein flexibility (Bioforum Europe 2008) How flexible protein structures are?

    E-print Network

    Paris-Sud XI, Université de

    Protein flexibility (Bioforum Europe 2008) 1 How flexible protein structures are? New questions on the protein structure plasticity. Aurélie Bornot1 , Bernard Offmann2,3 & Alexandre G. de Brevern 1 * 1 INSERM Offmann Protein structures and protein structural models are great tools to reach protein function

  17. The Effect of Sequence Complexity on the Construction of Protein-Protein Interaction Networks

    Microsoft Academic Search

    Mehdi Kargar; Aijun An

    2010-01-01

    \\u000a In this paper, the role of sequence complexity in the construction of important nodes in protein-protein interaction (PPI)\\u000a networks is investigated. We use two complexity measures, linguistic complexity and Shanon entropy, to measure the complexity\\u000a of protein sequences. Three different datasets of yeast PPI networks are used to conclude the results. It has been shown that\\u000a there are two important

  18. Multi-level machine learning prediction of protein–protein interactions in Saccharomyces cerevisiae

    PubMed Central

    Zubek, Julian; Tatjewski, Marcin; Boniecki, Adam; Mnich, Maciej; Basu, Subhadip

    2015-01-01

    Accurate identification of protein–protein interactions (PPI) is the key step in understanding proteins’ biological functions, which are typically context-dependent. Many existing PPI predictors rely on aggregated features from protein sequences, however only a few methods exploit local information about specific residue contacts. In this work we present a two-stage machine learning approach for prediction of protein–protein interactions. We start with the carefully filtered data on protein complexes available for Saccharomyces cerevisiae in the Protein Data Bank (PDB) database. First, we build linear descriptions of interacting and non-interacting sequence segment pairs based on their inter-residue distances. Secondly, we train machine learning classifiers to predict binary segment interactions for any two short sequence fragments. The final prediction of the protein–protein interaction is done using the 2D matrix representation of all-against-all possible interacting sequence segments of both analysed proteins. The level-I predictor achieves 0.88 AUC for micro-scale, i.e., residue-level prediction. The level-II predictor improves the results further by a more complex learning paradigm. We perform 30-fold macro-scale, i.e., protein-level cross-validation experiment. The level-II predictor using PSIPRED-predicted secondary structure reaches 0.70 precision, 0.68 recall, and 0.70 AUC, whereas other popular methods provide results below 0.6 threshold (recall, precision, AUC). Our results demonstrate that multi-scale sequence features aggregation procedure is able to improve the machine learning results by more than 10% as compared to other sequence representations. Prepared datasets and source code for our experimental pipeline are freely available for download from: http://zubekj.github.io/mlppi/ (open source Python implementation, OS independent). PMID:26157620

  19. Analysis of Hepatic Glycogen-Associated Proteins

    PubMed Central

    Stapleton, David; Nelson, Chad; Parsawar, Krishna; McClain, Donald; Gilbert-Wilson, Ryan; Barker, Elizabeth; Rudd, Brant; Brown, Kevin; Hendrix, Wayne; O’Donnell, Paul; Parker, Glendon

    2010-01-01

    Glycogen particles are associated with a population of proteins that mediate its biological functions, including: management of glucose flux into and out of the glycogen particle, maintenance of glycogen structure and regulation of particle size, number, and cellular location. A survey of the glycogen-associated proteome would be predicted to identify the relative representation of known members of this population, and associations with unexpected proteins that have the potential to mediate other functions of the glycogen particle. We therefore purified glycogen particles from both mouse and rat liver, using different techniques, and analyzed the resulting tryptic peptides by mass spectrometry. We also specifically eluted glycogen-binding proteins from the pellet using malto-oligosaccharides. Comparison of the rat and mouse populations, and analysis of specifically eluted proteins, allow some conclusions to be made about the hepatic glycogen sub-proteome. With the exception of glycogen branching enzyme all glycogen metabolic proteins were detected. Novel associations were identified, including ferritin and starch-binding domain protein 1, a protein that contains both a transmembrane endoplasmic reticulum signal peptide and a carbohydrate-binding module. This study therefore provides insight into the organization of the glycogen proteome, identifies other associated proteins and provides a starting point to explore the dynamic nature and cellular distribution of this metabolically important protein population. PMID:20391537

  20. Preparative parallel protein purification (P4).

    PubMed

    Strömberg, Patrik; Rotticci-Mulder, Joke; Björnestedt, Robert; Schmidt, Stefan R

    2005-04-15

    In state of the art drug discovery, it is essential to gain structural information of pharmacologically relevant proteins. Increasing the output of novel protein structures requires improved preparative methods for high throughput (HT) protein purification. Currently, most HT platforms are limited to small-scale and available technology for increasing throughput at larger scales is scarce. We have adapted a 10-channel parallel flash chromatography system for protein purification applications. The system enables us to perform 10 different purifications in parallel with individual gradients and UV monitoring. Typical protein purification applications were set up. Methods for ion exchange chromatography were developed for different sample proteins and columns. Affinity chromatography was optimized for His-tagged proteins using metal chelating media and buffer exchange by gel filtration was also tested. The results from the present system were comparable, with respect to resolution and reproducibility, with those from control experiments on an AKTA purifier system. Finally, lysates from 10 E. coli cultures expressing different His-tagged proteins were subjected to a three-step parallel purification procedure, combining the above-mentioned procedures. Nine proteins were successfully purified whereas one failed probably due to lack of expression. PMID:15722038

  1. Longer action means better drug: tuning up protein therapeutics.

    PubMed

    Szlachcic, Anna; Zakrzewska, Malgorzata; Otlewski, Jacek

    2011-01-01

    An increasing number of proteins are currently available on the market as therapeutics and this branch of the pharmaceutical industry will expand substantially during the coming years. As many diseases result from dysfunction of proteins forming multicomponent complexes, protein drugs with their inherent high specificity and affinity seem to be optimal medical agents. On the other hand, proteins are often highly instable and sensitive to degradation, which questions their applicability as effective therapeutics. Therefore, redesign and engineering of proteins is usually a required step in the present day drug development. Several approaches have been applied to optimize the protein properties central to their pharmaceutical use. This review focuses on different strategies that improve two crucial factors influencing protein drug efficiency: protein stability and its in vivo half-life. We provide examples of successful genetic and chemical modifications applied in the design of effective protein therapeutics. PMID:21443940

  2. A new view on the role of intrinsic protein disorder

    NASA Astrophysics Data System (ADS)

    Liu, Jintao; Faeder, James; Camacho, Carlos

    2009-03-01

    Recent studies have found that many proteins do not have stable structure by themselves, i.e., are intrinsically disordered, which challenges the conventional view that structure determines protein function and interaction. We have analyzed the Human Protein Reference Database with the VSL2 protein disorder predictor, and find that the amount of disorder in a protein is the result of evolutionary pressure: catalytic proteins interact with substrates rapidly and highly specifically and thus exhibit low levels of disorder; transcription regulators often slide along DNA, which favors flexible or disordered structures; binding proteins have affinities that depend weakly on folding stability, and thus have a broad disorder distribution. Finally, our findings suggest that sequence/structural features such as phosphotyrosine are better indicators of multiple protein-protein interactions than disorder.

  3. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2013 BINF 731 Secondary Structure: Computational Problems Secondary structure characterization Secondary structure assignment Secondary structure prediction Protein structure classification Structural classes of proteins all all Protein Structure Classification SCOP

  4. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2012 BINF 731 Secondary Structure: Computational Problems Secondary structure characterization Secondary structure assignment Secondary structure prediction Protein structure classification Structural classes of proteins all all Protein Structure Classification SCOP

  5. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2009 BINF 731 Secondary Structure: Computational Problems Secondary structure characterization Secondary structure assignment Secondary structure prediction Protein structure classification Structural classes of proteins all all / Protein Structure Classification SCOP

  6. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2012 BINF 731 http://binf.gmu.edu/vaisman/binf731 Informatics Structural Bioinformatics Computational Structural Biology Protein Engineering Protein Design Drug Design Molecular Modeling Proteomics Structural Genomics Protein Structure and... Business Law Ethics

  7. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2009 BINF 731 http://binf.gmu.edu/vaisman/binf731 Biology Crystallography NMR Spectroscopy Protein Informatics Structural Bioinformatics Computational Structural Biology Protein Engineering Protein Design Drug Design Molecular Modeling Proteomics Structural

  8. The ADAR protein family

    PubMed Central

    2012-01-01

    Adenosine to inosine (A-to-I) RNA editing is a post-transcriptional process by which adenosines are selectively converted to inosines in double-stranded RNA (dsRNA) substrates. A highly conserved group of enzymes, the adenosine deaminase acting on RNA (ADAR) family, mediates this reaction. All ADARs share a common domain architecture consisting of a variable number of amino-terminal dsRNA binding domains (dsRBDs) and a carboxy-terminal catalytic deaminase domain. ADAR family members are highly expressed in the metazoan nervous system, where these enzymes predominantly localize to the neuronal nucleus. Once in the nucleus, ADARs participate in the modification of specific adenosines in pre-mRNAs of proteins involved in electrical and chemical neurotransmission, including pre-synaptic release machineries, and voltage- and ligand-gated ion channels. Most RNA editing sites in these nervous system targets result in non-synonymous codon changes in functionally important, usually conserved, residues and RNA editing deficiencies in various model organisms bear out a crucial role for ADARs in nervous system function. Mutation or deletion of ADAR genes results in striking phenotypes, including seizure episodes, extreme uncoordination, and neurodegeneration. Not only does the process of RNA editing alter important nervous system peptides, but ADARs also regulate gene expression through modification of dsRNA substrates that enter the RNA interference (RNAi) pathway and may then act at the chromatin level. Here, we present a review on the current knowledge regarding the ADAR protein family, including evolutionary history, key structural features, localization, function and mechanism. PMID:23273215

  9. Metastable Mesoscopic Phases in Concentrated Protein Solutions

    NASA Astrophysics Data System (ADS)

    Vekilov, P. G.; Pan, W.; Gliko, O.; Katsonis, P.; Galkin, O.

    It is sometimes claimed that the cytosol around the organelles, tubules, and other cellular structures represents a liquid phase. On the other hand, almost all protein molecules in the cytosol participate in complexes with other proteins, nucleic acids, small molecules, etc. The two pictures of a homogeneous liquid and a granular multiscale mixture appear incompatible. Thus, an important question in physical biology is whether the protein complexes represent a property of the protein solutions, or are the result of complex specific interaction involving multiple biological molecules. We apply light scattering, atomic force microscopy, and other techniques to demonstrate that even solutions of a single protein of moderate concentration do not comply with Gibbs's definition of phase. In such solutions clusters of sizes from several tens to several hundred nanometers exist and have limited lifetimes. These clusters have a higher free energy than the protein solution, and their lifetime is determined by a barrier for their decay. The clusters affect the viscous and visco-elastic behavior of the solution and are an essential part of potential condensation and aggregation pathways. Since the clusters are observed in solutions of single proteins, they indicate that the proteins have an intrinsic propensity to form mesoscopic structures, which likely is utilized in the formation of the protein complexes in the cytosol. Cluster theories developed for colloid systems appear inapplicable to proteins due to the high level of implied Coulombic repulsion. A Monte Carlo model with protein-like potentials reproduces the metastable clusters of dense liquid with limited lifetimes and variable sizes, and suggests that the clusters' sizes are determined by the kinetics of growth and decay, and not by thermodynamics. A microscopic theory, which should account for stabilizing and destabilizing factors involving protein molecules and solvent inside the clusters, is still to be developed.

  10. Pigment-protein complexes

    SciTech Connect

    Siegelman, H W

    1980-01-01

    The photosynthetically-active pigment protein complexes of procaryotes and eucaryotes include chlorophyll proteins, carotenochlorophyll proteins, and biliproteins. They are either integral components or attached to photosynthetic membranes. Detergents are frequently required to solubilize the pigment-protein complexes. The membrane localization and detergent solubilization strongly suggest that the pigment-protein complexes are bound to the membranes by hydrophobic interactions. Hydrophobic interactions of proteins are characterized by an increase in entropy. Their bonding energy is directly related to temperature and ionic strength. Hydrophobic-interaction chromatography, a relatively new separation procedure, can furnish an important method for the purification of pigment-protein complexes. Phycobilisome purification and properties provide an example of the need to maintain hydrophobic interactions to preserve structure and function.

  11. Protein folding tames chaos

    E-print Network

    Xia, Kelin

    2013-01-01

    Protein folding produces characteristic and functional three-dimensional structures from unfolded polypeptides or disordered coils. The emergence of extraordinary complexity in the protein folding process poses astonishing challenges to theoretical modeling and computer simulations. The present work introduces molecular nonlinear dynamics (MND), or molecular chaotic dynamics, as a theoretical framework for describing and analyzing protein folding. We unveil the existence of intrinsically low dimensional manifolds (ILDMs) in the chaotic dynamics of folded proteins. Additionally, we reveal that the transition from disordered to ordered conformations in protein folding increases the transverse stability of the ILDM. Stated differently, protein folding reduces the chaoticity of the nonlinear dynamical system, and a folded protein has the best ability to tame chaos. Additionally, we bring to light the connection between the ILDM stability and the thermodynamic stability, which enables us to quantify the disorderli...

  12. Protein synthesis and translation initiation factor activation in neonatal pigs fed increasing levels of dietary protein.

    PubMed

    Frank, Jason W; Escobar, Jeffery; Suryawan, Agus; Kimball, Scot R; Nguyen, Hanh V; Jefferson, Leonard S; Davis, Teresa A

    2005-06-01

    Limited data suggest that the growth of low-birth-weight infants is enhanced by feeding a high-protein diet; however, the mechanisms involved in the effect have not been delineated. To identify these mechanisms, 34 pigs were fed from 2 to 7 d of age [60 g dry matter/(kg body weight . d)] isocaloric milk diets that contained levels of dietary protein that were marginal, adequate, and in excess of the piglets protein requirement (21, 33, and 45% of dry matter, respectively). Dietary protein replaced lactose and fat on an isocaloric basis. Fractional protein synthesis rates, various biomarkers of translational regulation, and plasma glucose and insulin levels were measured in overnight food-deprived and fed pigs. Mean daily weight gain of pigs fed the 33 and 45% protein diets was greater than that of pigs fed the 21% protein diet (P < 0.01). Plasma glucose (P = 0.07) and insulin (P < 0.01) levels decreased as dietary protein increased 60 min after feeding. Protein synthesis rates in longissimus dorsi, gastrocnemius, masseter, heart, liver, kidney, jejunum, and pancreas were greater in the fed than in the food-deprived state (P < 0.01). Protein synthesis in skeletal muscle did not change with protein intake in the fed state, but decreased quadratically (P < 0.01) with increasing dietary protein in the food-deprived state. Protein kinase B, ribosomal protein S6 kinase 1(S6K1), and eukaryotic initiation factor (eIF) 4E binding protein-1 (4E-BP1) were more phosphorylated, and assembly of the inactive eukaryotic initiation factor 4E . 4E-BP1 complex in muscle and liver was reduced in the fed state (P < 0.001) and were not consistently affected by dietary protein level. The results suggest that feeding stimulates protein synthesis, and this is modulated by the activation of initiation factors that regulate mRNA binding to the ribosomal complex. However, the provision of a high-protein diet that exceeds the protein requirement does not further enhance protein synthesis or translation initiation factor activation. PMID:15930440

  13. Applications of SYPRO Orange and SYPRO Red Protein Gel Stains

    Microsoft Academic Search

    Thomas H. Steinberg; Richard P. Haugland; Victoria L. Singer

    1996-01-01

    We have further characterized the sensitivity and specificity of SYPRO Orange protein gel stain and SYPRO Red protein gel stain with native and 2-dimensional polyacrylamide gels and for staining gels prior to Western blot analysis. We found that nucleic acids are not stained by the SYPRO protein gel stains, in contrast to results obtained with commonly used silver staining techniques.

  14. An electrophoretic investigation of mammalian spermatid-specific nuclear proteins

    Microsoft Academic Search

    Maryvonne Lanneau; M. Loir

    1982-01-01

    Summary. Using standardized methods for protein extraction and analysis, the testes of rams, bulls, goats, boars, stallions, rats, cats, hedgehogs, European mink and ferrets were examined for basic spermatid nucleoproteins by electrophoresis. The results suggest that differences exist in the total number of these proteins as well as in the number and amount of the cross-linked cystein-containing proteins. These differences

  15. Factors affecting molecular characteristics of whey protein gelation

    Microsoft Academic Search

    Joyce I. Boye; Inteaz Alli; Ashraf A. Ismail; Bernard F. Gibbs; Yasuo Konishi

    1995-01-01

    The effects of pH, protein concentration, NaCl, heating temperature and time on the gelation of a whey protein concentrate (WPC) and the associated changes in the molecular conformation of the individual whey proteins were studied using polyacrylamide gel electrophoresis, high performance liquid chromatography and Fourier transform infrared spectroscopy. Heat denaturation was studied using differential scanning calorimetry. The results obtained showed

  16. Engineering a uranyl specific binding protein from NikR.

    SciTech Connect

    Wegner, S. V.; Boyaci, H.; Chen, H.; Jensen, M. P.; He, C. (Chemical Sciences and Engineering Division); ( PSC-USR); (Univ. of Chicago)

    2009-03-16

    The first uranyl-selective DNA-binding protein is designed using the E. coli nickel(II)-responsive protein NikR as the template. The resulting NikR? protein binds uranyl (see picture) with a dissociation constant Kd=53?nM and selectively binds to DNA in the presence of uranyl.

  17. Protein Structural Change Upon Ligand Binding: Linear Response Theory

    Microsoft Academic Search

    Mitsunori Ikeguchi; Jiro Ueno; Miwa Sato; Akinori Kidera

    2005-01-01

    A simple formula based on linear response theory is proposed to explain and predict the structural change of proteins upon ligand binding. By regarding ligand binding as an external perturbation, the structural change as a response is described by atomic fluctuations in the ligand-free form and the protein-ligand interactions. The results for three protein systems of various sizes are consistent

  18. An Analysis of Core Deformations in Protein Superfamilies

    Microsoft Academic Search

    Alejandra Leo-Macias; Pedro Lopez-Romero; Dmitry Lupyan; Daniel Zerbino; Angel R. Ortiz

    2005-01-01

    An analysis is presented on how structural cores modify their shape across homologous proteins, and whether or not a relationship exists between these structural changes and the vibrational normal modes that proteins experience as a result of the topological constraints imposed by the fold. A set of 35 representative, well-populated protein families is studied. The evolutionary directions of deformation are

  19. METHIONINE OXIDATION AND PROTEIN PHOSPHORYLATION: INTERACTIVE PARTNERS IN SIGNALING?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Protein phosphorylation can affect the activity, stability or localization of a protein and as result plays a broad role in regulation of processes ranging from metabolism to control of plant growth and development. One aspect of current interest in our lab is how protein kinases target their substr...

  20. A second cytotoxic proteolytic peptide derived from amyloid ?-protein precursor

    Microsoft Academic Search

    Daniel C. Lu; Shahrooz Rabizadeh; Sreeganga Chandra; Rana F. Shayya; Lisa M. Ellerby; Xin Ye; Guy S. Salvesen; Edward H. Koo; Dale E. Bredesen

    2000-01-01

    The amyloid ?-protein precursor gives rise to the amyloid ?-protein, the principal constituent of senile plaques and a cytotoxic fragment involved in the pathogenesis of Alzheimer disease. Here we show that amyloid ?-protein precursor was proteolytically cleaved by caspases in the C terminus to generate a second unrelated peptide, called C31. The resultant C31 peptide was a potent inducer of

  1. BLOOD GROUPS AND POLYMORPHIC PROTEINS IN CATTLE AND SWINE*

    E-print Network

    Paris-Sud XI, Université de

    BLOOD GROUPS AND POLYMORPHIC PROTEINS IN CATTLE AND SWINE* Bent LARSEN Department of Physiology V (Denmark) SUMMARY The well known blood group and polymorphic protein systems in cattle and pigs. The results from studies of genetic linkage among 10 blood group systems and 7 loci control- ling protein

  2. Heat-induced Protein Structure and Subfractions in Relation to Protein Degradation Kinetics and Intestinal Availability in Dairy Cattle

    SciTech Connect

    Doiron, K.; Yu, P; McKinnon, J; Christensen, D

    2009-01-01

    The objectives of this study were to reveal protein structures of feed tissues affected by heat processing at a cellular level, using the synchrotron-based Fourier transform infrared microspectroscopy as a novel approach, and quantify protein structure in relation to protein digestive kinetics and nutritive value in the rumen and intestine in dairy cattle. The parameters assessed included (1) protein structure a-helix to e-sheet ratio; (2) protein subfractions profiles; (3) protein degradation kinetics and effective degradability; (4) predicted nutrient supply using the intestinally absorbed protein supply (DVE)/degraded protein balance (OEB) system for dairy cattle. In this study, Vimy flaxseed protein was used as a model feed protein and was autoclave-heated at 120C for 20, 40, and 60 min in treatments T1, T2, and T3, respectively. The results showed that using the synchrotron-based Fourier transform infrared microspectroscopy revealed and identified the heat-induced protein structure changes. Heating at 120C for 40 and 60 min increased the protein structure a-helix to e-sheet ratio. There were linear effects of heating time on the ratio. The heating also changed chemical profiles, which showed soluble CP decreased upon heating with concomitant increases in nonprotein nitrogen, neutral, and acid detergent insoluble nitrogen. The protein subfractions with the greatest changes were PB1, which showed a dramatic reduction, and PB2, which showed a dramatic increase, demonstrating a decrease in overall protein degradability. In situ results showed a reduction in rumen-degradable protein and in rumen-degradable dry matter without differences between the treatments. Intestinal digestibility, determined using a 3-step in vitro procedure, showed no changes to rumen undegradable protein. Modeling results showed that heating increased total intestinally absorbable protein (feed DVE value) and decreased degraded protein balance (feed OEB value), but there were no differences between the treatments. There was a linear effect of heating time on the DVE and a cubic effect on the OEB value. Our results showed that heating changed chemical profiles, protein structure a-helix to e-sheet ratio, and protein subfractions; decreased rumen-degradable protein and rumen-degradable dry matter; and increased potential nutrient supply to dairy cattle. The protein structure a-helix to e-sheet ratio had a significant positive correlation with total intestinally absorbed protein supply and negative correlation with degraded protein balance.

  3. [The protein quality of commercial products of texturized soybean protein and meat mixtures].

    PubMed

    Elías, L G; Braham, J E; Navarrete, D A; Bressani, R

    1984-06-01

    The purpose of the present study was to evaluate the protein quality of eight commercial samples of texturized soybean protein (TSP) collected in the food markets of various Latin American countries. The study also included experiments designed to establish the limiting amino acids in those products, and studies to evaluate the process of texturization by extrusion cooking. Finally, experiments were also carried out to evaluate the effect of replacement of meat by TSP as well as the supplementary value of the latter to tortilla flour. The eight samples were characterized for their functional properties and protein quality. The results obtained indicate that all samples were different in density, water absorption, and nitrogen solubility. Likewise, all samples were different in their lysine, methionine and threonine content, with PER values ranging from 1.75 to 2.31. Protein quality standard value is 2.3. The amino acid supplementation studies showed a positive significant response to methionine addition, particularly in those products exhibiting low PER values. The addition of both lysine and threonine to the methionine-supplemented product, increased PER significantly. The data also demonstrated that replacement of more than 25% of meat protein by TSP protein decreased protein quality of the product, even when a TSP with an accepted protein quality value was used. This product was also found to supplement the quality of tortilla protein. Finally, it was established that the texturization process did not reduce the protein quality of soybean protein. Based on these results, it is concluded that the variability in the protein quality measured in the eight TSP samples was due to the initial quality of the soy flour texturized, which was already damaged before texturization. For this reason, it is recommended that Latin American countries establish TSP quality standards when this product is used as a protein extender for meat, and that appropriate systems be implemented for the quality control of such products. PMID:6543573

  4. Identification and Characterization of Proteins Involved in Nuclear Organization Using Drosophila GFP Protein Trap Lines

    PubMed Central

    Rohrbaugh, Margaret; Clore, Alyssia; Davis, Julia; Johnson, Sharonta; Jones, Brian; Jones, Keith; Kim, Joanne; Kithuka, Bramwel; Lunsford, Krystal; Mitchell, Joy; Mott, Brian; Ramos, Edward; Tchedou, Maza R.; Acosta, Gilbert; Araujo, Mark; Cushing, Stuart; Duffy, Gabriel; Graves, Felicia; Griffin, Kyler; Gurudatta, B. V.; Jackson, Deaundra; Jaimes, Denis; Jamison, Kendall; Jones, Khali; Kelley, Dhaujee; Kilgore, Marquita; Laramore, Derica; Le, Thuy; Mazhar, Bakhtawar; Mazhar, Muhammad M.; McCrary, Britney; Miller, Teanndras; Moreland, Celethia; Mullins, Alex; Munye, Elyas; Okoorie, Sheila; Pittman, Elisha; Roberts, Nikkita; Rose, De’Warren; Rowland, Alex; Shagarabi, Anwar; Smith, Jamela; Stallworth, Tayler; Stroud, Nicole; Sung, Elizabeth; Sung, Kai; Takenaka, Naomi; Torre, Eduardo; Veira, Jarvis; Vu, Kim; Wagstaff, William; Wood, Ashley M.; Wu, Karen; Yang, Jingping; Corces, Victor G.

    2013-01-01

    Background Strains from a collection of Drosophila GFP protein trap lines express GFP in the normal tissues where the endogenous protein is present. This collection can be used to screen for proteins distributed in the nucleus in a non-uniform pattern. Methodology/Principal Findings We analyzed four lines that show peripheral or punctate nuclear staining. One of these lines affects an uncharacterized gene named CG11138. The CG11138 protein shows a punctate distribution in the nuclear periphery similar to that of Drosophila insulator proteins but does not co-localize with known insulators. Interestingly, mutations in Lamin proteins result in alterations in CG11138 localization, suggesting that this protein may be a novel component of the nuclear lamina. A second line affects the Decondensation factor 31 (Df31) gene, which encodes a protein with a unique nuclear distribution that appears to segment the nucleus into four different compartments. The X-chromosome of males is confined to one of these compartments. We also find that Drosophila Nucleoplasmin (dNlp) is present in regions of active transcription. Heat shock leads to loss of dNlp from previously transcribed regions of polytene chromosome without redistribution to the heat shock genes. Analysis of Stonewall (Stwl), a protein previously found to be necessary for the maintenance of germline stem cells, shows that Stwl is present in a punctate pattern in the nucleus that partially overlaps with that of known insulator proteins. Finally we show that Stwl, dNlp, and Df31 form part of a highly interactive network. The properties of other components of this network may help understand the role of these proteins in nuclear biology. Conclusions/Significance These results establish screening of GFP protein trap alleles as a strategy to identify factors with novel cellular functions. Information gained from the analysis of CG11138 Stwl, dNlp, and Df31 sets the stage for future studies of these proteins. PMID:23341925

  5. Stabilization of Proteins and Noncovalent Protein Complexes during Electrospray Ionization by Amino Acid Additives.

    PubMed

    Zhang, Hua; Lu, Haiyan; Chingin, Konstantin; Chen, Huanwen

    2015-07-21

    Ionization of proteins and noncovalent protein complexes with minimal disturbance to their native structure presents a great challenge for biological mass spectrometry (MS). In living organisms, the native structure of intracellular proteins is commonly stabilized by solute amino acids (AAs) accumulated in cells at very high concentrations. Inspired by nature, we hypothesized that AAs could also pose a stabilizing effect on the native structure of proteins and noncovalent protein complexes during ionization. To test this hypothesis, here we explored MS response for various protein complexes upon the addition of free AAs at mM concentrations into the electrospray ionization (ESI) solution. Thermal activation of ESI droplets in the MS inlet capillary was employed as a model destabilizing factor during ionization. Our results indicate that certain AAs, in particular proline (Pro), pose considerable positive effect on the stability of noncovalent protein complexes in ESI-MS without affecting the signal intensity of protein ions and original protein-ligand equilibrium, even when added at the 20 mM concentration. The data suggest that the degree of protein stabilization is primarily determined by the osmolytic and ampholytic characteristics of AA solutes. The highest stability and visibility of noncovalent protein complexes in ESI-MS are achieved using AA additives with neutral isoelectric point, moderate proton affinity, and unfavorable interaction with the native protein state. Overall, our results indicate that the simple addition of free amino acids into the working solution can notably improve the stability and accuracy of protein analysis by native ESI-MS. PMID:26115185

  6. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2012 BINF 731 Protein Function Prediction Enzyme Function for predicting DNA-binding proteins function from protein structure FINDSITE - a threading-based method sites and their properties in evolutionarily related proteins Pandit et al., PSiFR, 2009 Protein

  7. Protein Structure Analysis Iosif Vaisman

    E-print Network

    Vaisman, Iosif

    Protein Structure Analysis Iosif Vaisman 2012 BINF 731 Protein Engineering Protein Engineering stability Increase proteins resistance to proteases Change codon composition Protein Engineering Bornscheueretal.Nature485,185-194(2012) Protein Engineering Bornscheuer et al. Nature 485, 185-194 (2012

  8. Self-Assembling Protein Microarrays

    Microsoft Academic Search

    Niroshan Ramachandran; Eugenie Hainsworth; Bhupinder Bhullar; Samuel Eisenstein; Benjamin Rosen; Albert Y. Lau; Johannes C. Walter; Joshua LaBaer

    2004-01-01

    Protein microarrays provide a powerful tool for the study of protein function. However, they are not widely used, in part because of the challenges in producing proteins to spot on the arrays. We generated protein microarrays by printing complementary DNAs onto glass slides and then translating target proteins with mammalian reticulocyte lysate. Epitope tags fused to the proteins allowed them

  9. Interplay between chaperones and protein disorder promotes the evolution of protein networks.

    PubMed

    Pechmann, Sebastian; Frydman, Judith

    2014-06-01

    Evolution is driven by mutations, which lead to new protein functions but come at a cost to protein stability. Non-conservative substitutions are of interest in this regard because they may most profoundly affect both function and stability. Accordingly, organisms must balance the benefit of accepting advantageous substitutions with the possible cost of deleterious effects on protein folding and stability. We here examine factors that systematically promote non-conservative mutations at the proteome level. Intrinsically disordered regions in proteins play pivotal roles in protein interactions, but many questions regarding their evolution remain unanswered. Similarly, whether and how molecular chaperones, which have been shown to buffer destabilizing mutations in individual proteins, generally provide robustness during proteome evolution remains unclear. To this end, we introduce an evolutionary parameter ? that directly estimates the rate of non-conservative substitutions. Our analysis of ? in Escherichia coli, Saccharomyces cerevisiae, and Homo sapiens sequences reveals how co- and post-translationally acting chaperones differentially promote non-conservative substitutions in their substrates, likely through buffering of their destabilizing effects. We further find that ? serves well to quantify the evolution of intrinsically disordered proteins even though the unstructured, thus generally variable regions in proteins are often flanked by very conserved sequences. Crucially, we show that both intrinsically disordered proteins and highly re-wired proteins in protein interaction networks, which have evolved new interactions and functions, exhibit a higher ? at the expense of enhanced chaperone assistance. Our findings thus highlight an intricate interplay of molecular chaperones and protein disorder in the evolvability of protein networks. Our results illuminate the role of chaperones in enabling protein evolution, and underline the importance of the cellular context and integrated approaches for understanding proteome evolution. We feel that the development of ? may be a valuable addition to the toolbox applied to understand the molecular basis of evolution. PMID:24968255

  10. Interplay between Chaperones and Protein Disorder Promotes the Evolution of Protein Networks

    PubMed Central

    Pechmann, Sebastian; Frydman, Judith

    2014-01-01

    Evolution is driven by mutations, which lead to new protein functions but come at a cost to protein stability. Non-conservative substitutions are of interest in this regard because they may most profoundly affect both function and stability. Accordingly, organisms must balance the benefit of accepting advantageous substitutions with the possible cost of deleterious effects on protein folding and stability. We here examine factors that systematically promote non-conservative mutations at the proteome level. Intrinsically disordered regions in proteins play pivotal roles in protein interactions, but many questions regarding their evolution remain unanswered. Similarly, whether and how molecular chaperones, which have been shown to buffer destabilizing mutations in individual proteins, generally provide robustness during proteome evolution remains unclear. To this end, we introduce an evolutionary parameter ? that directly estimates the rate of non-conservative substitutions. Our analysis of ? in Escherichia coli, Saccharomyces cerevisiae, and Homo sapiens sequences reveals how co- and post-translationally acting chaperones differentially promote non-conservative substitutions in their substrates, likely through buffering of their destabilizing effects. We further find that ? serves well to quantify the evolution of intrinsically disordered proteins even though the unstructured, thus generally variable regions in proteins are often flanked by very conserved sequences. Crucially, we show that both intrinsically disordered proteins and highly re-wired proteins in protein interaction networks, which have evolved new interactions and functions, exhibit a higher ? at the expense of enhanced chaperone assistance. Our findings thus highlight an intricate interplay of molecular chaperones and protein disorder in the evolvability of protein networks. Our results illuminate the role of chaperones in enabling protein evolution, and underline the importance of the cellular context and integrated approaches for understanding proteome evolution. We feel that the development of ? may be a valuable addition to the toolbox applied to understand the molecular basis of evolution. PMID:24968255

  11. New reporters of protein trafficking and protein-protein interactions in live cells

    E-print Network

    Fernández Suárez, Marta

    2008-01-01

    Here, we describe our attempts to harness the exquisite specificity of natural protein and RNA enzymes to develop improved methods to study protein localization and protein-protein interactions in live cells. We first ...

  12. Perturbation waves in proteins and protein networks: Applications of percolation and game theories in signaling and drug design

    Microsoft Academic Search

    Miklos A. Antal; Csaba Bode; Peter Csermely

    2008-01-01

    The network paradigm is increasingly used to describe the dynamics of complex systems. Here we review the current results and propose future development areas in the assessment of perturbation waves, i.e. propagating structural changes in amino acid networks building individual protein molecules and in protein-protein interaction networks (interactomes). We assess the possibilities and critically review the initial attempts for the

  13. Human T-lymphotropic Virus Type 1-infected Cells Secrete Exosomes That Contain Tax Protein*S

    E-print Network

    Vertes, Akos

    Human T-lymphotropic Virus Type 1-infected Cells Secrete Exosomes That Contain Tax Protein delivery of functional HTLV-1 proteins and mRNA to recipient cells. Discovered in the early 1980s, human T functional elements. Results: Exosomal Tax protein causes phenotypic changes in uninfected cells. Conclusion

  14. Purification and characterization of 30S ribosomal proteins from Bacillus subtilis : correlation to Escherichia coli 30S proteins

    Microsoft Academic Search

    Ken-ichi Higo; Eiko Otaka; Syozo Osawa

    1982-01-01

    Twenty proteins were isolated from the 30S ribosomal subunits of Bacillus subtilis and their amino acid compositions and amino-terminal amino acid sequences were determined. These results were compared with the data of Escherichia coli 30S ribosomal proteins and the structural correspondence of individual ribosomal proteins has been established between B. subtilis and E. coli.

  15. Stable Isotope Labeling Strategy for Protein-Ligand Binding Analysis in Multi-Component Protein Mixtures

    NASA Astrophysics Data System (ADS)

    DeArmond, Patrick D.; West, Graham M.; Huang, Hai-Tsang; Fitzgerald, Michael C.

    2011-03-01

    Described here is a stable isotope labeling protocol that can be used with a chemical modification- and mass spectrometry-based protein-ligand binding assay for detecting and quantifying both the direct and indirect binding events that result from protein-ligand binding interactions. The protocol utilizes an H{2/16}O2 and H{2/18}O2 labeling strategy to evaluate the chemical denaturant dependence of methionine oxidation in proteins both in the presence and absence of a target ligand. The differential denaturant dependence to the oxidation reactions performed in the presence and absence of ligand provides a measure of the protein stability changes that occur as a result of direct interactions of proteins with the target ligand and/or as a result of indirect interactions involving other protein-ligand interactions that are either induced or disrupted by the ligand. The described protocol utilizes the 18O/16O ratio in the oxidized protein samples to quantify the ligand-induced protein stability changes. The ratio is determined using the isotopic distributions observed for the methionine-containing peptides used for protein identification in the LC-MS-based proteomics readout. The strategy is applied to a multi-component protein mixture in this proof-of-principle experiment, which was designed to evaluate the technique's ability to detect and quantify the direct binding interaction between cyclosporin A and cyclophilin A and to detect the indirect binding interaction between cyclosporin A and calcineurin (i.e., the protein-protein interaction between cyclophilin A and calcineurin that is induced by cyclosporin A binding to cyclophilin A).

  16. Aux\\/IAA Proteins Contain a Potent Transcriptional Repression Domain

    Microsoft Academic Search

    Shiv B. Tiwari; Gretchen Hagen; Tom J. Guilfoyle

    2004-01-01

    Aux\\/IAA proteins are short-lived nuclear proteins that repress expression of primary\\/early auxin response genes in protoplast transfection assays. Repression is thought to result from Aux\\/IAA proteins dimerizing with auxin response factor (ARF) transcriptional activators that reside on auxin-responsive promoter elements, referred to as AuxREs. Most Aux\\/IAA proteins contain four conserved domains, designated domains I, II, III, and IV. Domain II

  17. Chapter 6 Plasmon Resonance Methods in Membrane Protein Biology

    Microsoft Academic Search

    Zdzislaw Salamon; Gordon Tollin; Isabel Alves; Victor Hruby

    2009-01-01

    Plasmon waveguide resonance (PWR) spectroscopy, a variant of surface plasmon resonance (SPR) spectrometry, allows one to examine changes in conformation of anisotropic structures such as membranes and membrane?associated proteins such as G?protein–coupled receptors (GPCRs). The binding and resulting structural changes that accompany interactions of membrane protein with ligands (agonists, antagonists, inverse agonist, etc.), G?proteins, and other effectors and modulators of

  18. The utilization of oilseed proteins as encapsulation agents

    E-print Network

    Baker, Grey Wayne

    1979-01-01

    -. pared from oilseed proteins functioned well in baking appli- cations but poorly as coffee whiteners. Protein level and solubility had very little effect on functionality. Overall results indicate oilseed proteins are effective encapsulating agents...THE UTILIZATION OF OILSEED PROTEINS AS ENCAPSULATION AGENTS A Thesis by GREY WAYNE BAKER Submitted to the Graduate College of Texas A&M University in partial fulfillment of the requirement for the degree of MASTER OF SCIENCE August 1979...

  19. Improved method for simultaneous isolation of proteins and nucleic acids

    Microsoft Academic Search

    Soroth Chey; Claudia Claus; Uwe Gerd Liebert

    2011-01-01

    Guanidinium thiocyanate–phenol–chloroform extraction (GTPC extraction) is widely used in molecular biology for isolating DNA, RNA, and proteins. Protein isolation by commercially available GTPC solutions is time consuming and the resulting pellets are only incompletely soluble. In this study ethanol–bromochloropropane–water was used for precipitation of proteins from the phenol–ethanol phase after GTPC extraction of RNA and DNA. The precipitated proteins can

  20. The BRO proteins of Bombyx mori nucleopolyhedrovirus are nucleocytoplasmic shuttling proteins that utilize the CRM1-mediated nuclear export pathway

    SciTech Connect

    Kang, Won Kyung [Molecular Entomology Laboratory, RIKEN, 2-1 Hirosawa, Wako 351-0198 (Japan)]. E-mail: wkkang@riken.jp; Kurihara, Masaaki [Molecular Entomology Laboratory, RIKEN, 2-1 Hirosawa, Wako 351-0198 (Japan)]. E-mail: mkuri@riken.jp; Matsumoto, Shogo [Molecular Entomology Laboratory, RIKEN, 2-1 Hirosawa, Wako 351-0198 (Japan)]. E-mail: smatsu@riken.jp

    2006-06-20

    The BRO proteins of Bombyx mori nucleopolyhedrovirus (BmNPV) display a biphasic pattern of intracellular localization during infection. At early times, they reside in the nucleus but then show both cytoplasmic and nuclear localization as the infection proceeds. Therefore, we examined the possibility of nuclear export. Using inhibitors, we reveal that BmNPV BRO proteins shuttle between the nucleus and cytoplasm. Mutations on the leucine-rich region of BRO proteins resulted in nuclear accumulation of transiently expressed proteins, suggesting that this region functions as a CRM1-dependent nuclear export signal (NES). On the contrary, mutant BRO-D with an altered NES did not show nuclear accumulation in infected cells, although protein production seemed to be reduced. RT-PCR analysis showed that the lower level of protein production was due to a reduction in RNA synthesis. Taken together, our results suggest that BRO proteins are nucleocytoplasmic shuttling proteins that utilize the CRM1-mediated nuclear export pathway.