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Peptide Corresponding (peptide + corresponding)
Kinds of Peptide Corresponding Selected AbstractsMyosin Va phosphorylated on Ser1650 is found in nuclear speckles and redistributes to nucleoli upon inhibition of transcriptionCYTOSKELETON, Issue 6 2008Maria Cristina S. Pranchevicius Abstract Nuclear actin and nuclear myosins have been implicated in the regulation of gene expression in vertebrate cells. Myosin V is a class of actin-based motor proteins involved in cytoplasmic vesicle transport and anchorage, spindle-pole alignment and mRNA translocation. In this study, myosin-Va, phosphorylated on a conserved serine in the tail domain (phospho-ser1650 MVa), was localized to subnuclear compartments. A monoclonal antibody, 9E6, raised against a peptide corresponding to phosphoserine1650 and flanking regions of the murine myosin Va sequence, was immunoreactive to myosin Va heavy chain in cellular and nuclear extracts of HeLa cells, PC12 cells and B16-F10 melanocytes. Immunofluorescence microscopy with this antibody revealed discrete irregular spots within the nucleoplasm that colocalized with SC35, a splicing factor that earmarks nuclear speckles. Phospho-ser1650 MVa was not detected in other nuclear compartments, such as condensed chromatin, Cajal bodies, gems and perinucleolar caps. Although nucleoli also were not labeled by 9E6 under normal conditions, inhibition of transcription in HeLa cells by actinomycin D caused the redistribution of phospho-ser1650 MVa to nucleoli, as well as separating a fraction of phospho-ser1650 MVa from SC35 into near-neighboring particles. These observations indicate a novel role for myosin Va in nuclear compartmentalization and offer a new lead towards the understanding of actomyosin-based gene regulation. Cell Motil. Cytoskeleton 2008. © 2008 Wiley-Liss, Inc. [source] Failure of immunocompetitive capillary electrophoresis assay to detect disease-specific prion protein in buffy coat from humans and chimpanzees with Creutzfeldt-Jakob diseaseELECTROPHORESIS, Issue 5 2003Larisa Cervenakova Abstract The emergence of a new environmentally caused variant of Creutzfeldt-Jakob disease (vCJD), the result of food-born infection by the causative agent of bovine spongiform encephalopathy (BSE), has stimulated research on a practical diagnostic screening test. The immunocompetitive capillary electrophoresis (ICCE) assay has been reported to detect disease-specific, proteinase-resistant prion protein (PrPres) in the blood of scrapie-infected sheep. We have applied this method to blood from CJD-infected chimpanzees and humans. The threshold of detection achieved with our ICCE was 0.6 nM of synthetic peptide corresponding to the prion protein (PrP) C -terminus, and 2 nM of recombinant human PrP at the optimized conditions. However, the test was unable to distinguish between extracts of leucocytes from healthy and CJD-infected chimpanzees, and from healthy human donors and patients affected with various forms of CJD. Thus, the ICCE assay as presently performed is not suitable for use as a screening test in human transmissible spongiform encephalopathies (TSEs). [source] The cytosolic domain of APP induces the relocalization of dynamin 3 in hippocampal neuronsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 9 2006X. Meckler Abstract Amyloid precursor protein (APP) has been the subject of intense research to uncover its implication in Alzheimer's disease. Its physiological function is, however, still poorly understood. Herein, we investigated its possible influence on the development of cultured hippocampal neurons. A peptide corresponding to the APP intracellular domain linked to a cell-penetrating peptide was used to alter the interactions of APP with its cytosolic partners. This treatment promoted the concentration of the cytosolic GTPase dynamin 3 (Dyn3) in neurite segments when most untreated cells displayed a homogenous punctate distribution of Dyn3. The Dyn3-labelled segments were excluded from those revealed by APP staining after aldehyde fixation. Interestingly, after aldehyde fixation MAP2 also labelled segments excluded from APP-stained segments. Thus APP is also a marker for the spacing pattern of neurites demonstrated by Taylor & Fallon (2006)J. Neurosci., 26, 1154,4463. [source] Disruption of structural and functional integrity of ,2 -macroglobulin by cathepsin EFEBS JOURNAL, Issue 6 2003Mitsue Shibata ,2 -Macroglobulin (,2M) is an abundant glycoprotein with the intrinsic capacity for capturing diverse proteins for rapid delivery into cells. After internalization by the receptor- mediated endocytosis, ,2M-protein complexes were rapidly degraded in the endolysosome system. Although this is an important pathway for clearance of both ,2M and biological targets, little is known about the nature of ,2M degradation in the endolysosome system. To investigate the possible involvement of intracellular aspartic proteinases in the disruption of structural and functional integrity of ,2M in the endolysosome system, we examined the capacity of ,2M for interacting with cathepsin E and cathepsin D under acidic conditions and the nature of its degradation. ,2M was efficiently associated with cathepsin E under acidic conditions to form noncovalent complexes and rapidly degraded through the generation of three major proteins with apparent molecular masses of 90, 85 and 30 kDa. Parallel with this reaction, ,2M resulted in the rapid loss of its antiproteolytic activity. Analysis of the N-terminal amino-acid sequences of these proteins revealed that ,2M was selectively cleaved at the Phe811-Leu812 bond in about 100mer downstream of the bait region. In contrast, little change was observed for ,2M treated by cathepsin D under the same conditions. Together, the synthetic SPAFLA peptide corresponding to the Ser808,Ala813 sequence of human ,2M, which contains the cathepsin E-cleavage site, was selectively cleaved by cathepsin E, but not cathepsin D. These results suggest the possible involvement of cathepsin E in disruption of the structural and functional integrity of ,2M in the endolysosome system. [source] All or none fibrillogenesis of a prion peptideFEBS JOURNAL, Issue 18 2001Wen-Quan Zou Amyloid proteins and peptides comprise a diverse group of molecules that vary both in size and amino-acid sequence, yet assemble into amyloid fibrils that have a common core structure. Kinetic studies of amyloid fibrillogenesis have revealed that certain amyloid proteins form oligomeric intermediates prior to fibril formation. We have investigated fibril formation with a peptide corresponding to residues 195,213 of the human prion protein. Through a combination of kinetic and equilibrium studies, we have found that the fibrillogenesis of this peptide proceeds as an all-or-none reaction where oligomeric intermediates are not stably populated. This variation in whether oligomeric intermediates are stably populated during fibril formation indicates that amyloid proteins assemble into a common fibrillar structure; however, they do so through different pathways. [source] CCN2, connective tissue growth factor, stimulates collagen deposition by gingival fibroblasts via module 3 and ,6- and ,1 integrinsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2006Edwin C.K. Heng Abstract CCN2, (connective tissue growth factor, CTGF) is a matricellular factor associated with fibrosis that plays an important role in the production and maintenance of fibrotic lesions. Increased collagen deposition and accumulation is a common feature of fibrotic tissues. The mechanisms by which CCN2/CTGF contributes to fibrosis are not well understood. Previous studies suggest that CTGF exerts some of its biological effects at least in part by integrin binding, though this mechanism has not been previously shown to contribute to fibrosis. Utilizing full length CCN2/CTGF, CCN2/CTGF fragments, and integrin neutralizing antibodies, we provide evidence that the effects of CCN2/CTGF to stimulate extracellular matrix deposition by gingival fibroblasts are mediated by the C-terminal half of CCN2/CTGF, and by ,6 and ,1 integrins. In addition, a synthetic peptide corresponding to a region of CCN2/CTGF domain 3 that binds ,6,1 inhibits the collagen-deposition assay. These studies employed a new and relatively rapid assay for CCN2/CTGF-stimulated collagen deposition based on Sirius Red staining of cell layers. Data obtained support a pathway in which CCN2/CTGF could bind to ,6,1 integrin and stimulate collagen deposition. These findings provide new experimental methodologies applicable to uncovering the mechanism and signal transduction pathways of CCN2/CTGF-mediated collagen deposition, and may provide insights into potential therapeutic strategies to treat gingival fibrosis and other fibrotic conditions. J. Cell. Biochem. 98: 409,420, 2006. © 2006 Wiley-Liss, Inc. [source] Molecular characterization of protein kinase C-, binding to lamin AJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2002Alberto M. Martelli Abstract Previous results from our laboratory have identified lamin A as a protein kinase C (PKC)-binding protein. Here, we have identified the regions of PKC-, that are crucial for this binding. By means of overlay assays and fusion proteins made of glutathione-S-transferase (GST) fused to elements of rat PKC-,, we have established that binding occurs through both the V5 region and a portion of the C2 region (i.e., the calcium-dependent lipid binding (CaLB) domain) of the kinase. In particular, we have found that amino acid 200,217 of the CaLB domain are essential for binding lamin A, as a synthetic peptide corresponding to this stretch of amino acids prevented the interaction between the CaLB domain and lamin A. We also show that the presence of four lysine residues of the CaLB domain (K205, K209, K211, and K213) was essential for the binding. We have determined that binding of elements of PKC-, to lamin A does not require the presence of cofactors such as phosphatidylserine (PS) and Ca2+. We have also found that the binding site of lamin A for the CaLB domain of PKC-, is localized in the carboxyl-terminus of the lamin, downstream of amino acid 499. Our findings may prove to be important to clarify the mechanisms regulating PKC function within the nucleus and may also lead to the synthesis of isozyme-specific drugs to attenuate or reverse PKC-dependent nuclear signaling pathways important for the pathogenesis of cancer. © 2002 Wiley-Liss, Inc. [source] Role of atypical protein kinase C isozymes and NF-,B in IL-1,-induced expression of cyclooxygenase-2 in human myometrial smooth muscle cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Sara V. Duggan Increased myometrial expression of cyclooxygenase-2 (Cox-2) at term results from elevated local levels of inflammatory cytokines, and its inhibition provides a potential route for intervention in human pre-term labor. We have identified a role for atypical protein kinase C (PKC) isozymes in IL-1,-induced Cox-2 expression in human myometrial smooth muscle cells (HMSMC). The PKC inhibitor GF109203X (10 µM) inhibited IL-1,-induced Cox-2 protein and RNA expression, which were also reduced by MAPK and nuclear factor ,B (NF-,B) inhibitors. GF109203X did not affect MAPK activities, and neither did it replicate the effect of p38 MAPK inhibition on Cox-2 mRNA stability, suggesting that PKC operates through an independent mechanism. The effect of GF109203X remained intact after depletion of conventional and novel PKC isozymes by phorbol ester pre-treatment. In contrast LY379196 (10 µM), which at micromolar concentrations inhibits all but atypical PKCs, did not affect Cox-2 expression. A peptide corresponding to the pseudosubstrate sequence of atypical PKCs blocked Cox-2 protein expression, whereas the sequence from conventional PKCs was ineffective. GF109203X did not affect NF-,B binding to nuclear proteins, but strongly reduced NF-,B-dependent transcription in luciferase reporter assays. Our findings indicate that IL-1,-induced Cox-2 expression in HMSMC in culture requires p38-MAPK-mediated mRNA stabilization and an independent activation of Cox-2 transcription which is dependent on the action of atypical PKCs, probably through direct stimulation of the transactivating activity of NF-,B. J. Cell. Physiol. 210: 637,643, 2007. © 2006 Wiley-Liss, Inc. [source] Structural characterization of a neurotoxic threonine-rich peptide corresponding to the human prion protein ,2-helical 180,195 segment, and comparison with full-length ,2-helix-derived peptides,JOURNAL OF PEPTIDE SCIENCE, Issue 10 2008Luisa Ronga Abstract The 173,195 segment corresponding to the helix 2 of the globular PrP domain is a good candidate to be one of the several ,spots' of intrinsic structural flexibility, which might induce local destabilization and concur to protein transformation, leading to aggregation-prone conformations. Here, we report CD and NMR studies on the ,2-helix-derived peptide of maximal length (hPrP[180,195]) that is able to exhibit a regular structure different from the prevalently random arrangement of other ,2-helix-derived peptides. This peptide, which has previously been shown to be affected by buffer composition via the ion charge density dependence typical of Hofmeister effects, corresponds to the C -terminal sequence of the PrPC full-length ,2-helix and includes the highly conserved threonine-rich 188,195 segment. At neutral pH, its conformation is dominated by ,-type contributions, which only very strong environmental modifications are able to modify. On TFE addition, an increase of ,-helical content can be observed, but a fully helical conformation is only obtained in neat TFE. However, linking of the 173,179 segment, as occurring in wild-type and mutant peptides corresponding to the full-length ,2-helix, perturbs these intrinsic structural propensities in a manner that depends on whether the environment is water or TFE. Overall, these results confirm that the 180,195 parental region in hPrPC makes a strong contribution to the chameleon conformational behavior of the segment corresponding to the full-length ,2-helix, and could play a role in determining structural rearrangements of the entire globular domain. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd. [source] G,s protein C -terminal ,-helix at the interface: does the plasma membrane play a critical role in the G,s protein functionality?JOURNAL OF PEPTIDE SCIENCE, Issue 10 2005Stefania Albrizio Abstract The heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins, G,,,) mediate the signalling process of a large number of receptors, known as G protein-coupled receptors. The C -terminal domain of the heterotrimeric G protein ,-subunit plays a key role in the selective activation of G proteins by their cognate receptors. The interaction of this domain can take place at the end of a cascade including several successive conformational modifications. G,s(350,394) is the 45-mer peptide corresponding to the C -terminal region of the G,s subunit. In the crystal structure of the G,s subunit it encompasses the ,4/,6 loop, the ,6 ,-sheet segment and the ,5 helix region. Following a previous study based on the synthesis, biological activity and conformational analysis of shorter peptides belonging to the same G,s region, G,s(350,394) was synthesized and investigated. The present study outlines the central role played by the residues involved in the ,4/,6 loop and ,6/,5 loops in the stabilization of the C -terminal G,s,-helix. H2O/2H2O exchange experiments, and NMR diffusion experiments show interesting evidence concerning the interaction between the SDS micelles and the polypeptide. These data prompt intriguing speculations on the role of the intracellular environment/cellular membrane interface in the stabilization and functionality of the C -terminal G,s region. Copyright © 2005 European Peptide Society and John Wiley & Sons, Ltd. [source] Identification of single amino acid residues essential for the binding of lipopolysaccharide (LPS) to LPS binding protein (LBP) residues 86,99 by using an Ala-scanning libraryJOURNAL OF PEPTIDE SCIENCE, Issue 4 2002Dr O. Reyes Abstract Lipopolysaccharide binding protein (LBP) is a 60 kDa acute phase glycoprotein capable of binding to LPS of Gram-negative bacteria and facilitating its interaction with cellular receptors. This process is thought to be of great importance in systemic inflammatory reactions such as septic shock. A peptide corresponding to residues 86,99 of human LBP (LBP86,99) has been reported to bind specifically with high affinity the lipid A moiety of LPS and to inhibit the interaction of LPS with LBP. We identified essential amino acids in LBP86,99 for binding to LPS by using a peptide library corresponding to the Ala-scanning of human LBP residues 86,99. Amino acids Trp91 and Lys92 were indispensable for peptide,LPS interaction and inhibition of LBP,LPS binding. In addition, several alanine-substituted synthetic LBP-derived peptides inhibited LPS,LBP interaction. Substitution of amino acids Arg94, Lys95 and Phe98 by Ala increased the inhibitory effect. The mutant Lys95 was the most active in blocking LPS binding to LBP. These findings emphasize the importance of single amino acids in the LPS binding capacity of small peptides and may contribute to the development of new drugs for use in the treatment of Gram-negative bacterial sepsis. Copyright © 2002 European Peptide Society and John Wiley & Sons, Ltd. [source] A personal account of the role of peptide research in drug discovery: the case of hepatitis C,JOURNAL OF PEPTIDE SCIENCE, Issue 1 2001Antonello Pessi Abstract Although peptides themselves are not usually the end products of a drug discovery effort, peptide research often plays a key role in many aspects of this process. This will be illustrated by reviewing the experience of peptide research carried out at IRBM in the course of our study of hepatitis C virus (HCV). The target of our work is the NS3/4A protease, which is essential for maturation of the viral polyprotein. After a thorough examination of its substrate specificity we fine-tuned several substrate-derived peptides for enzymology studies, high-throughput screening and as fluorescent probes for secondary binding assays. In the course of these studies we made the key observation: that the protease is inhibited by its own cleavage products. Single analog and combinatorial optimization then derived potent peptide inhibitors. The crucial role of the NS4A cofactor was also addressed. NS4A is a small transmembrane protein, whose central domain is the minimal region sufficient for enzyme activation. Structural studies were performed with a peptide corresponding to the minimal activation domain, with a series of product inhibitors and with both. We found that NS3/4A is an induced fit enzyme, requiring both the cofactor and the substrate to acquire its bioactive conformation; this explained some puzzling results of ,serine-trap' type inhibitors. A more complete study on NS3 activation, however, requires the availability of the full-length NS4A protein. This was prepared by native chemical ligation, after sequence engineering to enhance its solubility; structural studies are in progress. Current work is focused on the P, region of the substrate, which, at variance with the P region, is not used for ground state binding to the enzyme and might give rise to inhibitors showing novel interactions with the enzyme. Copyright © 2001 European Peptide Society and John Wiley & Sons, Ltd. [source] Copper binding to octarepeat peptides of the prion protein monitored by mass spectrometryPROTEIN SCIENCE, Issue 2 2000Randy M. Whittal Abstract Electrospray ionization mass spectrometry (ESI-MS) was used to measure the binding of Cu2+ ions to synthetic peptides corresponding to sections of the sequence of the mature prion protein (PrP). ESI-MS demonstrates that Cu2+ is unique among divalent metal ions in binding to PrP and defines the location of the major Cu2+ binding site as the octarepeat region in the N-terminal domain, containing multiple copies of the repeat ProHisGlyGlyGlyTrpGlyGln. The stoichiometries of the complexes measured directly by ESI-MS are pH dependent: a peptide containing four octarepeats chelates two Cu2+ ions at pH 6 but four at pH 7.4. At the higher pH, the binding of multiple Cu2+ ions occurs with a high degree of cooperativity for peptides C-terminally extended to incorporate a fifth histidine. Dissociation constants for each Cu2+ ion binding to the octarepeat peptides, reported here for the first time, are mostly in the low micromolar range; for the addition of the third and fourth Cu2+ ions to the extended peptides at pH 7.4, KD's are <100 nm. n-terminal acetylation of the peptides caused some reduction in the stoichiometry of binding at both ph's. cu2+ also binds to a peptide corresponding to the extreme N-terminus of PrP that precedes the octarepeats, arguing that this region of the sequence may also make a contribution to the Cu2+ complexation. Although the structure of the four-octarepeat peptide is not affected by pH changes in the absence of Cu2+, as judged by circular dichroism, Cu2+ binding induces a modest change at pH 6 and a major structural perturbation at pH 7.4. It is possible that PrP functions as a Cu2+ transporter by binding Cu2+ ions from the extracellular medium under physiologic conditions and then releasing some or all of this metal upon exposure to acidic pH in endosomes or secondary lysosomes. [source] The role of synaptotagmin I C2A calcium-binding domain in synaptic vesicle clustering during synapse formationTHE JOURNAL OF PHYSIOLOGY, Issue 1 2007Peter Gardzinski Synaptic vesicles aggregate at the presynaptic terminal during synapse formation via mechanisms that are poorly understood. Here we have investigated the role of the putative calcium sensor synaptotagmin I in vesicle aggregation during the formation of soma,soma synapses between identified partner cells using a simple in vitro synapse model in the mollusc Lymnaea stagnalis. Immunocytochemistry, optical imaging and electrophysiological recording techniques were used to monitor synapse formation and vesicle localization. Within 6 h, contact between appropriate synaptic partner cells up-regulated global synaptotagmin I expression, and induced a localized aggregation of synaptotagmin I at the contact site. Cell contacts between non-synaptic partner cells did not affect synaptotagmin I expression. Application of an human immunodeficiency virus type-1 transactivator (HIV-1 TAT)-tagged peptide corresponding to loop 3 of the synaptotagmin I C2A domain prevented synaptic vesicle aggregation and synapse formation. By contrast, a TAT-tagged peptide containing the calcium-binding motif of the C2B domain did not affect synaptic vesicle aggregation or synapse formation. Calcium imaging with Fura-2 demonstrated that TAT,C2 peptides did not alter either basal or evoked intracellular calcium levels. These results demonstrate that contact with an appropriate target cell is necessary to initiate synaptic vesicle aggregation during nascent synapse formation and that the initial aggregation of synaptic vesicles is dependent on loop 3 of the C2A domain of synaptotagmin I. [source] The Arabidopsis MAP kinase kinase MKK1 participates in defence responses to the bacterial elicitor flagellinTHE PLANT JOURNAL, Issue 4 2006Tamás Mészáros Summary Plants sense pathogens through both pathogen-associated molecular patterns and recognition of race-specific virulence factors, which induce basal defence or an accelerated defence (often manifest in the form of local cell death), respectively. A mitogen-activated protein kinase (MAPK) module in Arabidopsis was previously proposed to signal from perception of the bacterial elicitor flagellin to the activation of basal defence-related genes. Here, we present evidence for a parallel MAPK-signalling pathway involved in the response to flg22, a peptide corresponding to the most conserved domain of flagellin. The endogenous Arabidopsis MAP kinase kinase MKK1 is activated in cells treated with flg22, phosphorylates the MAPK MPK4 in vitro, and activates it in vivo in protoplasts. In mkk1 mutant plants, the activation by flg22 of MPK4 and two other flg22-induced MAPKs (MPK3 and MPK6) is impaired. In the mkk1 mutant, a battery of both flg22-induced and flg22-repressed genes show altered expression, indicating that MKK1 negatively regulates the activity of flagellin-responsive genes. Intriguingly, in contrast to the mpk4 mutant, mkk1 shows no morphological anomalies and is compromised in resistance to both virulent and avirulent Pseudomonas syringae strains. Thus, the MKK1 signalling pathway modulates the expression of genes responding to elicitors and plays an important role in pathogen defence. [source] A Synthetic Mechano Growth Factor E Peptide Enhances Myogenic Precursor Cell Transplantation SuccessAMERICAN JOURNAL OF TRANSPLANTATION, Issue 10 2007P. Mills Myogenic precursor cell (MPC) transplantation is a good strategy to introduce dystrophin expression in muscles of Duchenne muscular dystrophy (DMD) patients. Insulin-like growth factor (IGF-1) promotes MPC activities, such as survival, proliferation, migration and differentiation, which could enhance the success of their transplantation. Alternative splicing of the IGF-1 mRNA produces different muscle isoforms. The mechano growth factor (MGF) is an isoform, especially expressed after a mechanical stress. A 24 amino acids peptide corresponding to the C-terminal part of the MGF E domain (MGF-Ct24E peptide) was synthesized. This peptide had been shown to enhance the proliferation and delay the terminal differentiation of C2C12 myoblasts. The present study showed that the MGF-Ct24E peptide improved human MPC transplantation by modulating their proliferation and differentiation. Indeed, intramuscular or systemic delivery of this synthetic peptide significantly promoted engraftment of human MPCs in mice. In vitro experiments demonstrated that the MGF-Ct24E peptide enhanced MPC proliferation by a different mechanism than the binding to the IGF-1 receptor. Moreover, MGF-Ct24E peptide delayed human MPC differentiation while having no outcome on survival. Those combined effects are probably responsible for the enhanced transplantation success. Thus, the MGF-Ct24E peptide is an interesting agent to increase MPC transplantation success in DMD patients. [source] Bim,Bcl-2 homology 3 mimetic therapy is effective at suppressing inflammatory arthritis through the activation of myeloid cell apoptosisARTHRITIS & RHEUMATISM, Issue 2 2010John C. Scatizzi Objective Rheumatoid arthritis (RA) is a destructive autoimmune disease characterized by an increased inflammation in the joint. Therapies that activate the apoptotic cascade may have potential for use in RA; however, few therapeutic agents fit this category. The purpose of this study was to examine the potential of Bim, an agent that mimics the action of Bcl-2 homology 3 (BH3) domain,only proteins that have shown success in preclinical studies of cancer, in the treatment of autoimmune disease. Methods Synovial tissues from RA and osteoarthritis patients were analyzed for the expression of Bim and CD68 using immunohistochemistry. Macrophages from Bim,/, mice were examined for their response to lipopolysaccharide (LPS) using flow cytometry, real-time polymerase chain reaction analysis, enzyme-linked immunosorbent assay, and immunoblotting. Bim,/, mice were stimulated with thioglycollate or LPS and examined for macrophage activation and cytokine production. Experimental arthritis was induced using the K/BxN serum,transfer model. A mimetic peptide corresponding to the BH3 domain of Bim (TAT-BH3) was administered as a prophylactic agent and as a therapeutic agent. Edema of the ankles and histopathologic analysis of ankle tissue sections were used to determine the severity of arthritis, its cellular composition, and the degree of apoptosis. Results The expression of Bim was reduced in RA synovial tissue as compared with controls, particularly in macrophages. Bim,/, macrophages displayed elevated expression of markers of inflammation and secreted more interleukin-1, following stimulation with LPS or thioglycollate. TAT-BH3 ameliorated arthritis development, reduced the number of myeloid cells in the joint, and enhanced apoptosis without inducing cytotoxicity. Conclusion These data demonstrate that BH3 mimetic therapy may have significant potential for the treatment of RA. [source] Deiminated Epstein-Barr virus nuclear antigen 1 is a target of anti,citrullinated protein antibodies in rheumatoid arthritisARTHRITIS & RHEUMATISM, Issue 3 2006Federico Pratesi Objective To test the hypothesis that deimination of viral sequences containing Arg,Gly repeats could generate epitopes recognized by anti,citrullinated protein antibodies (ACPAs) that are present in rheumatoid arthritis (RA) sera. Methods Multiple antigen peptides derived from Epstein-Barr virus (EBV),encoded Epstein-Barr nuclear antigen 1 (EBNA-1) were synthesized, substituting the arginines with citrulline, and were used to screen RA sera. Anti,cyclic citrullinated peptide antibodies were purified by affinity chromatography and tested on a panel of in vitro deiminated proteins. Their ability to bind in vivo deiminated proteins was evaluated by immunoprecipitation, using EBV-infected cell lines. Results Antibodies specific for a peptide corresponding to the EBNA-135,58 sequence containing citrulline in place of arginine (viral citrullinated peptide [VCP]) were detected in 50% of RA sera and in <5% of normal and disease control sera. In addition, affinity-purified anti-VCP antibodies from RA sera reacted with filaggrin-derived citrullinated peptides, with deiminated fibrinogen, and with deiminated recombinant EBNA-1. Moreover, anti-VCP antibodies immunoprecipitated, from the lysate of calcium ionophore,stimulated lymphoblastoid cell lines, an 80-kd band that was reactive with a monoclonal anti,EBNA-1 antibody and with anti,modified citrulline antibodies. Conclusion These data indicate that ACPAs react with a viral deiminated protein and suggest that EBV infection may play a role in the induction of these RA-specific antibodies. [source] Trifluoroethanol and binding to model membranes stabilize a predicted turn in a peptide corresponding to the first extracellular loop of the angiotensin II AT1A receptorBIOPOLYMERS, Issue 1 2002Roberto K. Salinas Abstract Homology modeling of the angiotensin II AT1A receptor based on rhodopsin,s crystal structure has assigned the 92,100 (YRWPFGNHL) sequence of the receptor to its first extracellular loop. Solution and membrane-bound conformational properties of a peptide containing this sequence (EL1) were examined by CD, fluorescence, and 1H-NMR. CD spectra in aqueous solution revealed an equilibrium between less organized and folded conformers. NMR spectra indicated the coexistence of trans and cis isomers of the Trp3,Pro4 bond. A positive band at 226 nm in the CD spectra suggested aromatic ring stacking, modulated by EL1's ionization degree. CD spectra showed that trifluoroethanol (TFE), or binding to detergent micelles and phospholipid bilayers, shifted the equilibrium toward conformers with higher secondary structure content. Different media gave rise to spectra suggestive of different ,-turns. Chemical shift changes in the NMR spectra corroborated the stabilization of different conformations. Thus, environments of lower polarity or binding to interfaces probably favored the formation of hydrogen bonds, stabilizing ,-turns, predicted for this sequence in the whole receptor. Increases in Trp3 fluorescence intensity and anisotropy, blue shifts of the maximum emission wavelength, and pK changes also evinced the interaction between EL1 and model membranes. Binding was seen to depend on both hydrophobic and electrostatic interactions, as well as lipid phase packing. Studies with water-soluble and membrane-bound fluorescence quenchers demonstrated that Trp3 is located close to the water,membrane interface. The results are discussed with regard to possible implications in receptor folding and function. © 2002 Wiley Periodicals, Inc. Biopolymers 65: 21,31, 2002 [source] Involvement of molecular mimicry between human T-cell leukemia virus type 1 gp46 and osteoprotegerin in induction of hypercalcemiaCANCER SCIENCE, Issue 3 2009Yasuko Sagara Human T-cell leukemia virus type-1 (HTLV-1) causes adult T-cell leukemia/lymphoma (ATL), frequently associated with hypercalcemia and bone destruction. A positive correlation between the appearance of an antibody recognizing the central region (Asp197 to Leu216) on Gp46, gp46-197, and the severity of ATL has been demonstrated. In this study, five male Nihon Hakusyoku rabbits were immunized with a synthetic peptide corresponding to the gp46-197 region to clarify its action and mechanism. Two of the rabbits showed piloerection, anorexia, and somnolence, and died soon after booster administration. The serum calcium level of the dead rabbits was significantly high, compared to those of surviving rabbits. Interestingly, amino acid sequences homologous with gp46-197 were found in the carboxyl-terminal half of osteoprotegerin (OPG), an osteoclast inhibitory factor. To confirm the effect of the gp46-197 region on osteogenesis in vivo, the peptide was intraperitoneally administered to male Sprague-Dawley rats. The administration of the gp46-197 peptide resulted in a decrease of bone mineral density (BMD), a significant increase of serum calcium level, and inhibition of normal bone growth in both short- and long-term experiments. In rats, femoral growth inhibition by the gp46-197 peptide was restored by the coadministration of recombinant human OPG. Improvement by OPG in the adverse effect indicates that the central region of HTLV-1 Gp46 acts as an antagonist for OPG and leads to hypercalcemia. (Cancer Sci 2009; 100: 490,496) [source] Studies on the conformational properties of CP-1042,55, the hinge region of CP-10, using circular dichroism and RP-HPLCCHEMICAL BIOLOGY & DRUG DESIGN, Issue 6 2000E. Lazoura Abstract: The conformational properties of CP-1042,55, a peptide corresponding to the hinge region of CP-10, were investigated using circular dichroism spectroscopy and reverse-phase high-performance liquid chromatography (RP-HPLC). The circular dichroism studies indicated that CP-1042,55 formed considerable secondary structure in the presence of hydrophobic solution environments including 50% acetonitrile, 50% trifluoroethanol and 200 mm sodium dodecyl sulfate, which comprised a mixture of ,-helix and ,-sheet. The effect of temperature on the conformation of CP-1042,55 was investigated between 5 and 40°C, with very small changes in the spectra being observed.RP-HPLC was then used to investigate the effect of temperature on the conformation of CP-1042,55 in the presence of a hydrophobic surface. Using a C18 -adsorbent, CP-1042,55 exhibited a conformational transition at 25°C, which was associated with an increase in the chromatographic contact area and the binding affinity of the peptide for the stationary phase. In addition, near-planar bandbroadening behaviour indicated that conformational species interconverted with rapid rate constants compared with the chromatographic time scale. These results indicated that the conformational change at 25°C in theRP-HPLC system most likely corresponds to the unfolding of an ,-helical and/or ,-sheet structure to an extended coil structure. Therefore, the strong chemotactic properties of this peptide may be attributed to its ability to form considerable secondary structure in the presence of a hydrophobic environment. [source] | ||||||||||||||||