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N-terminal Peptide (n-terminal + peptide)
Selected AbstractsProtein tyrosine phosphatase SHP-1 specifically recognizes C-terminal residues of its substrates via helix ,0JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2001Jian Yang Abstract The catalytic domain of protein tyrosine phosphatase SHP-1 possesses distinct substrate specificity. It recognizes the P-3 to P-5 residues of its substrates via the ,5-loop-,6 region. To study the substrate specificity further, we determined the structure of the catalytic domain of SHP-1 (C455S) complexed with a less-favorable-substrate peptide originated from SIRP,. The complex has disordered N-terminal peptide structure and reduced interactions between the N-terminal peptide and the ,5-loop-,6 region. This could be the basis for the lower affinity of peptide pY427 for the catalytic domain of SHP-1. In addition, by comparing the SHP-1/less-favorable peptide complex structure with the SHP-1/substrate complex structures, we identified a novel substrate-recognition site in the catalytic domain of SHP-1. This site was formed by helix ,0 and the ,5-loop-,6 motif of SHP-1, and specifically bound residues at the P,+,4 and further C-terminal positions of peptide substrates. © 2001 Wiley-Liss, Inc. [source] Munc18-1 as a key regulator of neurosecretionJOURNAL OF NEUROCHEMISTRY, Issue 1 2010Gayoung A. Han J. Neurochem. (2010) 115, 1,10. Abstract Munc18-1 plays essential roles in neurosecretion by interacting with syntaxin-1 and controlling the formation of the soluble N -ethylmaleimide-sensitive factor attachment protein receptors (SNARE) complex. At least three important functions of Munc18-1 have been proposed: (i) molecular chaperone of syntaxin-1 for appropriate localization and expression of syntaxin-1, (ii) priming/stimulation of the SNARE-mediated membrane fusion, and (iii) docking of large dense-core vesicles to the plasma membrane. Similarly, at least two different binding modes have been proposed for the interaction between Munc18-1 and syntaxin-1: (i) binary binding to a ,closed' conformation of syntaxin-1, and (ii) binding to the N-terminal peptide of syntaxin-1, which is thought to enable an interaction with the quaternary SNARE complex and/or further stabilize the binary interaction between Munc18-1 and closed syntaxin-1. Recent structural analyses have identified critical Munc18-1 residues implicated in these different modes of binding. These have recently been tested functionally in rescue experiments using Munc18-1 null neurons, chromaffin cells and Munc18-1/-2 knockdown PC12 cells, allowing remarkable progress to be made in the structural/functional understanding of Munc18-1. In this review, we summarize these recent advances and attempt to propose an updated model of the pleiotropic functions of Munc18-1 in neuroexocytosis. [source] Longitudinal evaluation of a fibrosis index combining MMP-1 and PIIINP compared with MMP-9, TIMP-1 and hyaluronic acid in patients with chronic hepatitis C treated by interferon-alpha and ribavirinJOURNAL OF VIRAL HEPATITIS, Issue 10 2006C. Trocme Summary., We have recently described a fibrosis index combining serum procollagen type III N-terminal peptide (PIIINP) and matrix metalloproteinase 1 (MMP-1) concentrations for evaluating the amount of liver fibrosis in chronic hepatitis C patients. The aims of the present study were to validate this score in another cohort of patients and to assess its variations along those of TIMP-1, hyaluronic acid (HA) and MMP-9 during antiviral treatment. Seventy-nine patients treated by interferon-alpha and ribavirin for 24 or 48 weeks were included. A liver biopsy was performed within the 6 months before the start of treatment. Serum markers were measured in serum collected the day of the liver biopsy, at start of treatment, and every 3 months during treatment and a 6-month follow-up period. The PIIINP/MMP-1 index was significantly correlated to the METAVIR fibrosis (r = 0.68, P < 0.001). Its overall diagnostic value defined by the area under the receiver operating characteristics curves was 0.77 for discriminating F1 vs F2F3F4, and 0.81 for discriminating F1F2 vs F3F4, and was better than that observed for HA and TIMP-1. At the end of follow-up, the PIIINP/MMP-1 index significantly decreased in responders and remained stable in nonresponder patients. This decrease occurred early and continued regularly during the treatment period. This variation was because of both a decrease of PIIINP and an increase of MMP-1 concentrations. HA and TIMP-1 serum concentrations were also significantly lower at the end of follow-up in responder patients, but early changes were minimal and not influenced by the response to treatment. Our study shows that a noninvasive index combining PIIINP and MMP-1 is a useful tool to follow-up fibrosis change during and after antiviral therapy chronic hepatitis C patients. [source] Mite serine protease activates protease-activated receptor-2 and induces cytokine release in human keratinocytesALLERGY, Issue 9 2009T. Kato Background:, House dust mites produce serine and cysteine proteases. Mite-derived proteases have been suggested to be involved in the pathogenesis of allergies; however, whether mite-derived serine protease activity can stimulate keratinocytes remains unknown. Methods:, We examined the activation of primary human keratinocytes by serine protease-rich extract of whole mite culture and compared with that by recombinant group 1 allergens (rDer f 1 and rDer p 1), which exclusively exhibit cysteine protease activity. Results:, Protease activity of whole mite culture extract (WCE), rDer f 1 and rDer p 1 induced the release of IL-8 and granulocyte-macrophage colony-stimulating factor. Protease activity of WCEs induced a significant upregulation of their mRNA expression but rDer f 1 had much less effect. Protease activity of the WCE stimulated intracellular Ca2+ mobilization but rDer f 1 and rDer p 1 did not. The mobilization induced by agonists for the human protease-activated receptor (PAR)-2, an agonist peptide or trypsin, was diminished by pre-incubation of keratinocytes with WCE. rDer f 1 inefficiently cleaved a synthetic N-terminal peptide of PAR-2 at different sites from trypsin, but the resultant peptides did not stimulate the release of interleukin-8. Conclusions:, The results suggest that mite-derived serine protease activity may contribute to the pathogenesis of atopic dermatitis by activating keratinocytes via PAR-2 activation but cysteine protease activity of Der f 1 and Der p 1 acts via another mechanism. [source] Amino Acid Residues in GRK1/GRK7 Responsible for Interaction with S-Modulin/Recoverin,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2008Aya Torisawa GRK1 is a visual pigment kinase in rods and is essential for inactivation of light-activated rhodopsin. The GRK1 activity is inhibited by binding of the Ca2+ -bound form of S-modulin/recoverin. We previously identified the S-modulin/recoverin site to interact with GRK1. In the present study, we identified its counterpart in GRK1. We synthesized 29 of GRK1 or GRK7 partial peptides that cover the entire sequence of GRK1/GRK7, and examined whether these peptides inhibit S-modulin/recoverin activity most probably by preoccupying the binding site for GRK1. The inhibition was the greatest with the N-terminal peptide (p1, aa 3,23 in GRK7). On mutation of each of eight amino acid residues highly conserved in the p1 region of more than 10 orthologs, the inhibition was significantly reduced in the mutation of Leu6, Asn12 and Tyr15. We further examined the binding of the peptides, including mutated ones, to S-modulin/recoverin with a resonance mirror biosensor. The binding correlated well with the degree of the inhibition by a peptide. The inhibition, therefore, seemed to be due to a direct binding of the kinase peptide to the binding site of active S-modulin/recoverin. A GRK1 region close to its C-terminus also seemed to be the binding site for S-modulin/recoverin. [source] Glucose-regulated protein 78: A new partner of p53 in trophoblastPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 23 2009Serge Arnaudeau Abstract Although wild-type p53 protein is overexpressed in first trimester trophoblast, it is inactive towards its target genes Metalloproteinase 2 and 9. This seems to be due to a complex mechanism of inactivation and stabilization of p53 relying on the formation of protein complexes involving the N-terminus of p53. To detect the proteins associated with this sequence, we incubated biotinylated p53 N-terminal peptide in cytotrophoblastic cell medium 24,h before lysis of cells. We purified the proteins retained on biotinylated peptide using a neutravidin affinity column. Proteins were then identified by peptide mass finger printing followed or not by peptide fragmentation sequencing. Among these proteins, we identified glucose-regulated protein 78 (GRP78) and verified its interaction with p53 in trophoblastic cells by immunoprecipitation and Western blot analysis. Moreover, the decreased expression of GRP78 induced by GRP78siRNA or versipelostatin decreased the formation of high molecular weight p53 complexes and p53 monomer and increased trophoblastic invasion. These results suggest that GRP78 is involved in inactivation and stabilization of p53 and in the regulation of trophoblastic invasion. [source] Role of the transgenic human thyrotropin receptor A-subunit in thyroiditis induced by A-subunit immunization and regulatory T cell depletionCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 3 2008Y. Mizutori Summary Transgenic BALB/c mice that express intrathyroidal human thyroid stimulating hormone receptor (TSHR) A-subunit, unlike wild-type (WT) littermates, develop thyroid lymphocytic infiltration and spreading to other thyroid autoantigens after T regulatory cell (Treg) depletion and immunization with human thyrotropin receptor (hTSHR) adenovirus. To determine if this process involves intramolecular epitope spreading, we studied antibody and T cell recognition of TSHR ectodomain peptides (A,Z). In transgenic and WT mice, regardless of Treg depletion, TSHR antibodies bound predominantly to N-terminal peptide A and much less to a few downstream peptides. After Treg depletion, splenocytes from WT mice responded to peptides C, D and J (all in the A-subunit), but transgenic splenocytes recognized only peptide D. Because CD4+ T cells are critical for thyroid lymphocytic infiltration, amino acid sequences of these peptides were examined for in silico binding to BALB/c major histocompatibility complex class II (IA,d). High affinity subsequences (inhibitory concentration of 50% < 50 nm) are present in peptides C and D (not J) of the hTSHR and mouse TSHR equivalents. These data probably explain why transgenic splenocytes do not recognize peptide J. Mouse TSHR mRNA levels are comparable in transgenic and WT thyroids, but only transgenics have human A-subunit mRNA. Transgenic mice can present mouse TSHR and human A-subunit-derived peptides. However, WT mice can present only mouse TSHR, and two to four amino acid species differences may preclude recognition by CD4+ T cells activated by hTSHR-adenovirus. Overall, thyroid lymphocytic infiltration in the transgenic mice is unrelated to epitopic spreading but involves human A-subunit peptides for recognition by T cells activated using the hTSHR. [source] Toll-like receptor 6-independent signaling by diacylated lipopeptidesEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 1 2005Ute Buwitt-Beckmann Abstract Bacterial lipopeptides are strong immune modulators that activate early host responses after infection as well as initiating adjuvant effects on the adaptive immune system. These lipopeptides induce signaling in cells of the immune system through Toll-like receptor 2 (TLR2),TLR1 or TLR2,TLR6 heteromers. So far it has been thought that triacylated lipopeptides, such as the synthetic N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2RS)-propyl]-(R)-cysteine (Pam3)-CSK4, signal through TLR2,TLR1 heteromers, whereas diacylated lipopeptides, like the macrophage-activating lipopeptide from Mycoplasma fermentans (MALP2) or S-[2,3-bis(palmitoyloxy)-(2RS)-propyl]-(R)-cysteine (Pam2)-CGNNDESNISFKEK, induce signaling through TLR2,TLR6 heteromers. Using new synthetic lipopeptide derivatives we addressed the contribution of the lipid and, in particular, the peptide moieties with respect to TLR2 heteromer usage. In contrast to the current model of receptor usage, not only triacylated lipopeptides, but also diacylated lipopeptides like Pam2CSK4 and the elongated MALP2 analog Pam2CGNNDESNISFKEK-SK4 (MALP2-SK4) induced B lymphocyte proliferation and TNF-, secretion in macrophages in a TLR6-independent manner as determined with cells from TLR6-deficient mice. Our results indicate that both the lipid and the N-terminal peptides of lipoproteins contribute to the specificity of recognition by TLR2 heteromers and are responsible for the ligand,receptor interaction on host cells. [source] Identification of the N-termini of NADPH : protochlorophyllide oxidoreductase A and B from barley etioplasts (Hordeum vulgare L.)FEBS JOURNAL, Issue 4 2009Matthias Plöscher The N-termini of the NADPH : protochlorophyllide oxidoreductase (POR) proteins A and B from barley and POR from pea were determined by acetylation of the proteins and selective isolation of the N-terminal peptides for mass spectrometry de novo sequence analysis. We show that the cleavage sites between the transit peptides and the three mature POR proteins are homologous. The N-terminus in PORA is V48, that in PORB is A61, and that in POR from pea is E64. For the PORB protein, two additional N-termini were identified as A62 and A63, with decreased signal intensity of the corresponding N-terminal peptides. The results show that the transit peptide of PORA is considerably shorter than previously reported and predicted by ChloroP. A pentapeptide motif that has been characterized as responsible for binding of protochlorophyllide to the transit peptide of PORA [Reinbothe C, Pollmann S, Phetsarath-Faure P, Quigley F, Weisbeek P & Reinbothe S (2008) Plant Physiol148, 694,703] is shown here to be part of the mature PORA protein. [source] A rapid isolation and identification method for blocked N-terminal peptides by isothiocyanate-coupled magnetic nanoparticles and MSPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 18 2009Liyan Zhao Abstract A quick isolation and identification of N-blocked peptides from protein digest mixtures were achieved by diisothiocyanate or isothiocyanate-coupled magnetic nanoparticles and MS. After protein digests were guanidinated and then mixed with diisothiocyanate or isothiocyanate-coupled magnetic nanoparticles, unmodified N-terminal peptides were covalently bound to magnetic nanoparticles, and can be removed from the mixture under magnetic field. Therefore, N-blocked peptides could be isolated and analyzed by MALDI or ESI MS. This new strategy was demonstrated with model peptides, proteins, and the lysates of HepG2 cells. [source] The structure of the BIR3 domain of cIAP1 in complex with the N-terminal peptides of SMAC and caspase-9ACTA CRYSTALLOGRAPHICA SECTION D, Issue 1 2009Raviraj Kulathila The inhibitor of apoptosis protein (IAP) family of molecules inhibit apoptosis through the suppression of caspase activity. It is known that the XIAP protein regulates both caspase-3 and caspase-9 through direct protein,protein interactions. Specifically, the BIR3 domain of XIAP binds to caspase-9 via a `hotspot' interaction in which the N-terminal residues of caspase-9 bind in a shallow groove on the surface of XIAP. This interaction is regulated via SMAC, the N-terminus of which binds in the same groove, thus displacing caspase-9. The mechanism of suppression of apoptosis by cIAP1 is less clear. The structure of the BIR3 domain of cIAP1 (cIAP1-BIR3) in complex with N-terminal peptides from both SMAC and caspase-9 has been determined. The binding constants of these peptides to cIAP1-BIR3 have also been determined using the surface plasmon resonance technique. The structures show that the peptides interact with cIAP1 in the same way that they interact with XIAP: both peptides bind in a similar shallow groove in the BIR3 surface, anchored at the N-terminus by a charge-stabilized hydrogen bond. The binding data show that the SMAC and caspase-9 peptides bind with comparable affinities (85 and 48,nM, respectively). [source] | ||||||||||||||||