Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

By using an AtT20 cell line transfected with complementary DNA for preproTRH, we have identified the proTRH polyeptide precursor [26 kilodaltons (kDa)] and shown that this molecule gives rise to the proTRH derived sequences as determined by pulse-chase and trypsinization studies. The predicted proTRH precursor composed of 231 amino acids with 5 copies of a TRH progenitor sequence (Gln-His-Pro-Gly) and 7 other cryptic peptides was demonstrated by: 1) Western Blot analysis of an AtT20 cell extract with anti-pCC10 antibodies (an antibody that recognizes the intact prohormone as well as some intermediate products of processing); 2) Immunoprecipitation of the radiolabelled 26 kDa protein with anti-pCC10 followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis; 3) Gel filtration chromatography of the radiolabeled 26 kDa extracted from SDS-PAGE. 4) RIA with anti-pCC10 antiserum against peptides extracted from adult rat hypothalamus and olfactory lobe after SDS-PAGE. 5) Trypsinization of the proTRH precursor which generated the proTRH cryptic peptides preproTRH25-50 (pYE27) and preproTRH53-74 (pFT22). These moieties were also produced during trypsinization of intermediate products of processing. By means of pulse-chase studies, the 26 kDa polypeptide was shown to be the biosynthetic precursor to all the proTRH derived cryptic peptides. Cleavage at two positions in the center of the molecule (Lys107-Arg108 and Lys152-Arg153) generated two moieties of 16.5 and 15 kDa. The 15 kDa N-terminal fragment is later cleaved to a 6 kDa peptide that includes the proTRH derived peptides, pYE27, pFT22, and pEH24. The carboxy-terminal 16.5 kDa fragment of the prohormone is processed to a 9.6 kDa fragment which contains the proTRH derived peptide pST10 (preproTRH160-169) and a fragment of 5.4 kDa that may be the C-terminal peptide preproTRH208-255 recognized by antisera pAC12 and pYE17. In further processing, the 9.6 kDa molecule is cleaved to produce a 5.4 kDa peptide from either sequences 115-169 or 160-199.
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PMID:Identification of the thyrotropin-releasing hormone-prohormone and its posttranslational processing in a transfected AtT20 tumoral cell line. 844 Jan 87

An acetylated polypeptide corresponding to residues 2-26 of human lipocortin 1 was synthesized and the anti-inflammatory activity assessed in three models of acute inflammation in rat and mouse. In the carrageenin rat paw oedema test, the peptide produced a maximal inhibition of approximately 41% at the 3 h time point with a 10 micrograms dose. When rat paw oedema was induced by the injection of venom phospholipase A2, the peptide produced a significant inhibition (31%) at the top dose of 20 micrograms per paw. In the mouse air-pouch model, systemic treatment with the peptide produced a dramatic reduction in cytokine-induced leukocyte migration with an ID50 of approximately 40 micrograms per mouse. The N-terminal peptide 2-26 shares the actions of lipocortin 1 in these acute models of inflammation.
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PMID:Anti-inflammatory actions of an N-terminal peptide from human lipocortin 1. 846 52

We have reported previously that the nucleoprotein (N), the phosphoprotein (P), and the 22-kDa protein of human respiratory syncytial virus (HRSV) are components of the cytoplasmic inclusion bodies observed in HEp-2-infected cells. In addition, coexpression of N and P was sufficient to induce the formation of N-P complexes detectable by either coimmunoprecipitation with anti-P antibodies or generation of cytoplasmic inclusions. We now report the identification of protein regions required for these interactions. Deletion mutant analysis of the P protein gene indicated that its C-terminal end was essential for interacting with N. This conclusion was strengthened by the finding that an anti-P monoclonal antibody (021/12P), reacting with a 21-residue P protein C-terminal peptide, apparently displaced N from N-P complexes. The same effect was observed with high concentrations of the C-terminal peptide. However, sequence requirements for the P protein C-terminal end were not absolute, and mutants with the substitution Ser-237-->Ala or Ser-237-->Thr were as efficient as the wild type in interacting with N. In addition, P and N proteins from strains of different HRSV antigenic groups, with sequence differences in the P protein C-terminal end, were able to coimmunoprecipitate and formed cytoplasmic inclusions. Deletion mutant analysis of the N gene indicated that large segments of this polypeptide were required for interacting with P. The relevance of these interactions for HRSV is discussed in comparison with those of analogous proteins from related viruses.
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PMID:Identification of protein regions involved in the interaction of human respiratory syncytial virus phosphoprotein and nucleoprotein: significance for nucleocapsid assembly and formation of cytoplasmic inclusions. 855 18

To study the behavior of the neuroendocrine polypeptide 7B2 in the presence of calcium, various fragments of this molecule were produced in Escherichia coli as fusion proteins to glutathione S-transferase (GST). Addition of millimolar concentrations of Ca2+ to purified preparations of hybrid molecules carrying the N-terminal segment of 7B2 induced precipitation in a manner dependent on protein and cation concentrations. This precipitation occurred at pH 7.5 but not at pH 5.2. It was augmented by 4 and 8 mM ATP, and reduced by 12 and 24 mM ATP. ADP had a similar but weaker effect. Calcium failed to cause precipitation of GST alone or of GST fused to the C-terminal peptide 7B2(156-186). However, when the latter protein was mixed with a GST protein carrying a short fragment of the N-terminal region of 7B2, both proteins were precipitated by calcium. Except for the pH dependence, the behavior of 7B2 fusion proteins in the presence of calcium and adenosine nucleotides are reminiscent of those exhibited by chromogranins and secretogranins, which, like 7B2, are acidic proteins found in the secretory granules of a variety of neuroendocrine cells. As suggested for other granins, this property may underlie the segregation of 7B2 fragments into secretory granules.
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PMID:Calcium-induced aggregation of neuroendocrine protein 7B2 in vitro and its modulation by ATP. 858 12

The extracellular hemoglobin of Lumbricus terrestris comprises four oxygen binding chains, a, b, c, d, and three linker chains L1, L2, L3 as major components. A stoichiometry of the whole molecule has been proposed on the basis of these chains, with a total number of 216 chains: forty-eight chains of each oxygen binding chain and eight molecules of each linker chain. We have isolated additional minor components by HPLC and two-dimensional gel electrophoresis. The following biochemical characterizations have been made. (i) All components so far reported, the heme-containing chains a, b, c, d, and linker chains L1, L2, L3 and a new minor polypeptide, L4, were mapped on a two-dimensional gel. Their estimated isoelectric points were between 4.7 and 5.9. (ii) The sequences of several peptides including the unique N-terminal peptide from linker L4 show that it can be considered as a duplicated gene product with a similar mass. (iii) Chain d2 was isolated and found to correspond to the minor chain previously pointed out by Shishikura et al. (J. Biol. Chem. 262 (1987) 3123-3131). (iv) The major chain d1 has serine at position 7 from the N-terminus. This is not consistent with previously reported glycine (Shishikura et al., J. Biol. Chem. 262 (1987) 3123-3131).
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PMID:Characterization of the constituent polypeptides of the extracellular hemoglobin from Lumbricus terrestris: heterogeneity and discovery of a new linker chain L4. 859 73

The neuroendocrine-specific polypeptide 7B2 is a constituent of the regulated secretary pathway. Recently, 7B2 was found to function as a molecular chaperone for prohormone convertase PC2. This report describes the genomic organization of the 7B2 gene which consists of six exons. Exon I corresponds to the 5'-untranslated mRNA region, while exons 2 and 3 encode the signal peptide and the amino-terminal half of the 7B2 protein that is distantly related to a subclass of molecular chaperones. The carboxy-terminal half of 7B2, responsible for its inhibitory action on PC2, is encoded by exons 4-6. Primer-extension analysis showed that the human 7B2 gene is transcribed from multiple transcription-initiation sites. The human 7B2 gene promoter contains a cAMP-responsive element, an AP-1 site, and several Pit-1/GHF-1-binding domains and heat-shock-element-like sequences but no obvious TATA or CAAT boxes. Of further interest is the finding of two DNA elements which are common to the promoter regions of the 7B2 gene and other genes selectively expressed in neuroendocrine tissues.
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PMID:Structural organization of the gene encoding the neuroendocrine chaperone 7B2. 861 87

Protein splicing involves the self-catalyzed formation of a branched intermediate, which then resolves into the excised intervening sequence and the spliced protein. A possible mechanism for branched intermediate formation is an N-O rearrangement of the peptide bond involving the amino group of the conserved serine/cysteine residue at the upstream splice junction to yield a linear peptide ester intermediate. This possibility was examined in using an in vitro splicing system involving the intervening sequence from the DNA polymerase of the extremely thermophilic archeon, Pyrococcus sp. GB-D. Because thioesters react much more rapidly with nitrogen nucleophiles at neutral pH than do oxygen esters, protein-splicing precursors in which the serine residue of interest was replaced by cysteine were constructed and purified. In the presence of 0.25 M hydroxylamine or 0.1 M ethylene diamine at pH 6 or higher, these constructs underwent rapid cleavage at the upstream splice junction, consistent with the aminolysis of a thioester. The site of hydroxylaminolysis was identified by analysis of the C-terminus of the polypeptide cleavage products. Comparison of the C-terminal peptide hydroxamate with the synthetic peptide hydroxamates with respect to chromatographic mobility, colorimetric assay, amino acid composition, and high-resolution mass spectrometry showed that the hydroxylamine-sensitive site in the splicing precursor was the peptide bond adjacent to the serine residue at the upstream splice junction. These results provide evidence that the peptide bond at the upstream splice junction can undergo a self-catalyzed N-O or N-S acyl rearrangement to yield a linear polypeptide ester intermediate and suggest that this kind of rearrangement constitutes the first step in protein splicing.
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PMID:Protein splicing: evidence for an N-O acyl rearrangement as the initial step in the splicing process. 862 3

We generated an antiserum to the predicted C-terminal peptide of the A17L open reading frame (ORF), which encodes a 23-kDa polypeptide with hydrophobic regions characteristic of membrane proteins. Immuno-electron microscopy of infected cells indicated that the A17L protein is intimately associated with the earliest characteristic viral membranes, even those formed in the presence of the drug rifampin. To study the role of the A17L protein in morphogenesis, we constructed recombinant vaccinia viruses in which the endogenous A17L ORF was deleted and a copy of the ORF under the control of the bacteriophage T7 RNA polymerase and the Escherichia coli lac repressor was inserted into an alternative site in the vaccinia virus genome. Growth of these recombinant viruses was entirely dependent on the induction of A17L expression by isopropyl-beta-D-thiogalactopyranoside. Electron microscopic examination of cells infected in the absence of inducer revealed the accumulation of large, well-demarcated electron-dense aggregates but no characteristic membrane-associated viral structures. Viral late protein synthesis occurred under these conditions, although the maturational proteolytic processing of structural proteins was inhibited. We conclude that the product of the A17L gene is an essential component of the immature viral membrane and has an early function in viral morphogenesis.
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PMID:Vaccinia virus A17L open reading frame encodes an essential component of nascent viral membranes that is required to initiate morphogenesis. 862 54

An echistatin analogue, designated as des(46-49)-[Ala8,37]-echistatin gamma, was synthesized chemically by solid-phase peptide synthesis. The analogue was made by replacing Cys8 and Cys37 residues with two alanines and the deletion of C-terminal peptide 46-49 of echistatin gamma, resulting in an artificial polypeptide of 45 amino acids with three disulfide bonds. In the platelet aggregation assay, the analogue exhibits almost the same activity as echistatin gamma, indicating that the linear sequence of des(46-49)-[Ala8,37]-echistatin gamma contains all of the primary-structure information that is required for proper folding of this synthetic polypeptide. The tertiary structure of the analogue, as determined from high-resolution nuclear magnetic resonance (NMR) coupled with dynamic simulated annealing, is very similar to that of echistatin alpha1 which differs from echistatin gamma by 8 residues. In particular the two important sites of the Arg-Gly-Asp (RGD) loop and the C-terminal Lys45, both of which show some degree of disorder, are maintained in similar spatial orientation and proximity as those in echistatin alpha 1 even without the constraint provided by the disulfide bond of the (Cys8-Cys37) pair. These results provide new insights in further defining distinct structural features of echistatin gamma, which are involved in supporting the active polypeptide conformation to achieve biological activity in the absence of one pair of disulfide bonds.
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PMID:Structural analysis of a biologically active echistatin analogue des(46-49)-[Ala8,37]-echistatin gamma with three disulfide bonds by 2D-NMR and computer graphics. 864 91

Although previous efforts to produce significant quantities of purified prohormone convertase 2 from either recombinant or natural sources have been unsuccessful, our recent finding that the neuroendocrine polypeptide 7B2 is necessary for the biosynthesis of enzymatically active prohormone convertase 2 (PC2) has enabled us to obtain active recombinant enzyme from the conditioned medium of PC2-producing CHO cells supertransfected with cDNA coding for 21 kDa 7B2. The recombinant enzyme was purified to apparent homogeneity, with a 40% recovery, in milligram quantities. Two protein bands of Mrs 71 and 75 kDa were observed after SDS-PAGE followed by either Coomassie staining or Western blotting with PC2 antiserum. Spontaneous conversion of the 71- and 75-kDa species to the 66-kDa form occurred during incubation at pH 5.0; the degree of conversion correlated with a dramatic increase in activity. Kms of 124 and 131 microM and Kcats of 0.49 and 0.81 s(-1) were obtained for the substrates Cbz-Arg-Ser-Lys-Arg-AMC and Pyr-Arg-Thr-Lys-Arg-AMC, respectively. The pH optimum was 5.0, and the enzyme was inhibited by h7B2(155-185') p-CMS, and EDTA but not by other inhibitors tested. Interestingly, 21 kDa 7B2 was observed to copurify with the enzyme in a molar ratio of about 1:100 (7B2:PC2). Prior addition of recombinant 21 kDa 7B2 to activated 66 kDa PC2 provided significant protection against thermal denaturation. When coassociated 7B2 was mostly removed from activated PC2 through gel filtration, subsequent addition of recombinant 7B2 exerted a significant stabilizing effect on enzyme activity. Millimolar Ca2+ and pHs between 5 and 6 were required to observe this effect. Since these conditions resemble those thought to occur within secretory granules, and since 21 kDa 7B2 represents a stored secretory granule protein, our data suggest a physiological role for 21 kDa 7B2 in the stabilization of PC2 activity.
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PMID:Purification and enzymatic characterization of recombinant prohormone convertase 2: stabilization of activity by 21 kDa 7B2. 866 Jun 52


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