Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Suspensions of oviduct cells were prepared by subjecting oviduct tissue to sequential incubations with EDTA, alpha-chymotrypsin, and crude collagenase, followed by a final incubation with EDTA. Cells isolated in this way incorporate mannose from exogenous GDP-mannose into mannosyl-lipid, oligosaccharide-lipid, and glycoprotein(s). Based on several criteria, the mannosyl-lipid is identical with mannosyl-phosphoryldolichol. Similarly, the oligosaccharide-lipid appears to be identical with the oligosaccharide-lipid synthesized in vitro (Lucas, J. J., Waechter, C. J., and Lennarz, W. J. (1975) J. Biol. Chem. 250, 1992-2002). In contrast, the glycoproteins are much lower in molecular weight than those labeled in cell-free preparations. Using intact oviduct cell suspensions it was found that: (a) exogenous GDP-mannose, not its breakdown products, serves as the direct mannosyl donor; (b) experiments using mixtures of known proportions of broken and intact cells, as well as studies with metabolic inhibitors, indicate that greater than 50% of the observed incorporation of mannose from GDP-mannose was catalyzed by enzymes associated with intact cells, rather than broken cells or membrane fragments; (c) incorporation of mannose from GDP-mannose into the mannosyl acceptors does not require energy and proceeds without significant uptake of GDP-mannose into trichloroacetic acid-soluble components of the cells; (d) under conditions where labeled guanosine incorporation into nucleic acids is readily detected, no incorporation of the guanosine moiety of [3H]GDP-mannose is observed. These results indicate that the enzymes catalyzing synthesis of lipid-linked intermediates involved in glycoprotein synthesis are not only associated with intracellular membranes, but with the plasma membrane as well.
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PMID:Utilization of exogenous GDP-mannose for the synthesis of mannose-containing lipids and glycoproteins by oviduct cells. 77 Apr 69

Binding of a denaturated polypeptide chain derived from chick skin collagen, the alpha 1(I) chain, by isolated membranes of human platelets has been demonstrated. The process is reversible, and time- and protein concentration-dependent. The binding is specific, with an association constant of 1.88 X 10(-6) M. Prior treatment of the isolated membranes with trypsin, chymotrypsin, and pronase, resulted in significant inhibition of the 14C-labeled alpha 1 chain binding, but neuraminidase or collagenase treatment had no effect. Dissociation of the bound radioactivity and subsequent chromatographic analyses on carboxymethylcellulose and agarose A-1.5m revealed that the alpha 1 chain was unaltered. Scatchard plot analysis suggested that there are approximately 20,000 binding sites per platelet. The binding of the alpha 1 chain was inhibited by a glycopeptide derived from alpha 1, alpha 1-CB5 and by purified glucosylgalactosyl hydroxylysine, but was not affected by other cyanogen bromide peptides of alpha 1, namely alpha 1-CB3, -CB4, -CB7, and -CB8. Kinetic studies demonstrated that inhibition by the hydroxylysine glycoside is competitive. Dose-response curves of platelet aggregation induced by alpha 1 and the binding of alpha 1 by platelet membranes correlate closely. These results indicate that there are specific binding sites for collagen alpha 1 chain on platelet membranes, and that the carbohydrate moiety of the alpha 1 chain plays a role in the binding. The findings also support the hypothesis that the chick skin alpha 1 chain mediates platelet aggregation and the release reaction by acting on platelet membranes.
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PMID:Binding of chick skin collagen alpha 1 chain by isolated membranes from human platelets. 97 74

Cells obtained from chick embryo tendons incorporate isotopically labeled glucosamine and mannose into the pro-alpha1 and pro-alpha2 chains of procollagen as judged by sodium dodecyl sulfate-gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The label was further localized to the propeptides of pro-alpha1 and pro-alpha2 by its chromatographic behavior after digestion with bacterial collagenase or alpha-chymotrypsin. Carbohydrate analysis of isolated pro-alpha chains showed the presence of labeled galactosamine in addition to mannose and glucosamine. Resistance to mild alkaline hydrolysis suggested that greater than 90% of the oligosaccharide units are not linked to the propeptide backbone by either serine or threonine.
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PMID:Carbohydrate moieties of procollagen: incorporation of isotopically labeled mannose and glucosamine into propeptides of procollagen secreted by matrix-free chick embryo tendon cells. 106 Nov 25

Plasmin inhibited the biosynthesis of tissue-type plasminogen activator (tPA) antigen by human umbilical vein endothelial cells (HUVEC) in a dose-dependent manner. The amount of tPA antigen found in the 24-h conditioned medium of cells treated with 100 nM plasmin for 1 h was 20-30% of that in the control group. However, in contrast to tPA, such treatment led to a 3-fold increase in plasminogen activator inhibitor (PAI) activity, whereas the amount of PAI type 1 antigen was unchanged. The effects of plasmin on HUVEC were binding- and catalytic activity-dependent and were specifically blocked by epsilon-aminocaproic acid. Microplasmin, which has no kringle domains, was less effective in reducing tPA antigen biosynthesis or enhancing PAI activity in HUVEC. Kringle domains of plasmin affected neither tPA antigen nor PAI activity of the cells. Other proteases including chymotrypsin, trypsin, and collagenase at comparable concentrations did not have a significant effect on the biosynthesis of tPA antigen or PAI activity of HUVEC. Thrombin stimulated the biosynthesis of tPA and PAI-1 antigens by HUVEC. Thrombin also stimulated an increase in the protein kinase activity in HUVEC, whereas plasmin inhibited the protein kinase activity of the cells. It is possible that plasmin regulates the biosynthesis of tPA in HUVEC through the signal transduction pathway involving protein kinase.
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PMID:Plasmin and the regulation of tissue-type plasminogen activator biosynthesis in human endothelial cells. 138 68

Matrix metalloproteinase 9 (MMP-9) has been purified as an inactive zymogen of M(r) 92,000 (proMMP-9) from the culture medium of HT 1080 human fibrosarcoma cells. The NH2-terminal sequence of proMMP-9 is Ala-Pro-Arg-Gln-Arg-Gln-Ser-Thr-Leu-Val-Leu-Phe-Pro, which is identical to that of the 92-kDa type IV collagenase/gelatinase. The zymogen can be activated by 4-aminophenylmercuric acetate, yielding an intermediate form of M(r) 83,000 and an active species of M(r) 67,000, the second of which has a new NH2 terminus of Met-Arg-Thr-Pro-Arg-(Cys)-Gly-Val-Pro-Asp-Leu-Gly-Arg-Phe-Gln-Thr- Phe-Glu. Immunoblot analyses demonstrate that this activation process is achieved by sequential processing of both NH2- and COOH-terminal peptides. TIMP-1 complexed with proMMP-9 inhibits the conversion of the intermediate form to the active species of M(r) 67,000. The proenzyme is fully activated by cathepsin G, trypsin, alpha-chymotrypsin, and MMP-3 (stromelysin 1) but not by plasmin, leukocyte elastase, plasma kallikrein, thrombin, or MMP-1 (tissue collagenase). During the activation by MMP-3, proMMP-9 is converted to an active species of M(r) 64,000 that lacks both NH2- and COOH-terminal peptides. In addition, HOCl partially activates the zymogen by reacting with an intermediate species of M(r) 83,000. The enzyme degrades type I gelatin rapidly and also cleaves native collagens including alpha 2 chain of type I collagen, collagen types III, IV, and V at undenaturing temperatures. These results indicate that MMP-9 has different activation mechanisms and substrate specificity from those of MMP-2 (72-kDa gelatinase/type IV collagenase).
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PMID:Matrix metalloproteinase 9 (92-kDa gelatinase/type IV collagenase) from HT 1080 human fibrosarcoma cells. Purification and activation of the precursor and enzymic properties. 140 Apr 81

The effects of various reactive oxygen species on latent human neutrophil and fibroblast-type interstitial collagenases were studied. Latent human neutrophil collagenases (proMMP-8) was efficiently activated by hypochlorous acid and hydrogen peroxide and less efficiently by the serine proteinases trypsin and chymotrypsin. Human plasmin and plasma kallikrein did not activate latent human neutrophil collagenase. The activation of latent human neutrophil collagenase by hypochlorous acid and hydrogen peroxide corresponded to the activation obtained with the other known non-proteolytic activators phenylmercuric chloride and gold thioglucose. The activation by hydrogen peroxide was inhibited by mannitol and desferoxamine, suggesting a localized Fenton-type reaction to be responsible for the generation of hydroxyl radical and/or hydroxyl radical-like reactive oxygen pathway of neutrophil procollagenase does not involve plasmin and plasma kallikrein, which are efficient proteolytic activators of latent fibroblast-type procollagenase (proMMP-1). Fibroblast procollagenase was also slightly activated by hypochlorous acid and gold thioglucose. Thus neutrophil procollagenase seems to prefer non-proteolytic means of activation and reactive oxygen species can be regarded as potent activators in vivo. Synovial-fluid neutrophils from rheumatoid arthritis patients were found to release collagenase in 30% active form when compared to same patients' peripheral blood neutrophils, which released collagenase in completely latent form. This may indicate that the triggering of neutrophil at the site of inflammation in vivo involves initial oxidative activation of collagenase upon the degranulation process.
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PMID:Reactive oxygen species as regulators of human neutrophil and fibroblast interstitial collagenases. 144 75

Procollagenase M(r) 85,000 (SDS-PAGE) was purified from buffy coat to homogeneity and represents a stable single polypeptide chain forming the entire proenzyme. The procollagenase can be activated by various proteinases, e.g. trypsin, chymotrypsin, cathepsin G, kallikrein and stromelysin and by different mercurial compounds. Proteolytic conversion of the latent enzyme to the active form by chymotrypsin is accompanied by a molecular weight reduction to an apparent M(r) 64,000. This active enzyme lacks the first 79 N-terminal residues. Activation by trypsin leads to a latent intermediate of apparent M(r) 70,000, lacking 48 N-terminal residues. The active enzyme is therefore generated upon prolonged incubation with trypsin by further cleavage of 22 N-terminal residues. Another latent intermediate form with apparent M(r) 69,000 is generated from the proenzyme upon incubation with leukocyte elastase by N-terminal cleavage of 53 or 64 residues, respectively. However, latent collagenase cannot be activated by plasmin. Activation by different mercurial compounds finally results in the formation of active collagenase with apparent M(r) 64,000. In contrast to the proenzyme, active collagenase can autolyse to give active M(r) 57,000 and 45,000 intermediates and two M(r) 28,000 fragments. Purification of latent leukocyte gelatinase yields three final products with apparent M(r) 98,000, 125,000 and 220,000 (SDS-PAGE; non reduced). Upon reduction, only the M(r) 98,000 form can be detected. The latent gelatinase can be activated in a similar manner as collagenase. Proteolytic activation by trypsin leads after N-terminal cleavage to an active gelatinase with sequence homology to leukocyte collagenase.
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PMID:Latent collagenase and gelatinase from human neutrophils and their activation. 148 34

Two clones were isolated by screening a shrimp hepatopancreas cDNA library with a DNA fragment obtained by PCR amplification using two oligonucleotides based on the partial protein sequence of Penaeus vanameii chymotrypsin purified earlier. One of these clones, PVC 7 contains a complete cDNA coding for a serine protease. The deduced amino acid sequence shows the existence of a 270 residue-long preproenzyme containing a highly hydrophobic signal peptide of 14 amino acids. This suggests the existence of a putative zymogen form of the enzyme containing a 30 amino acid-long peptide which is cleaved to give a mature protein of 226 residues. A highly preferred codon usage is observed for this protein. The other obtained cDNA was found to encode the less predominant variant of the protein. Sequence alignments show that shrimp chymotrypsin is highly homologous with crab collagenase (77% homology taking into account the same amino acid at the same position, and 83% homology taking into account amino acids with conserved function) and that it is more similar to mouse trypsin (41% homology of strictly conserved amino acids) than to hornet chymotrypsin (35% homology).
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PMID:Molecular cloning of a cDNA that encodes a serine protease with chymotryptic and collagenolytic activities in the hepatopancreas of the shrimp Penaeus vanameii (Crustacea, Decapoda). 151 90

The cell-extracellular matrix junction, which includes the cell wall and the outer surface of the plasma membrane, may be an essential region for the perception of gravity by the internodal cells of Chara corallina. Typically, when an internodal cell is oriented vertically, the downwardly directed cytoplasmic stream travels at a velocity that is 10% faster than that of the upwardly directed stream. However when the cells are treated with impermeant hydrolytic enzymes that partially digest cellulose or hemicellulose, the cells lose their ability to respond to gravity even though streaming continues. By contrast, enzymes that digest pectins have no effect on the gravity-induced polarity of cytoplasmic streaming. Furthermore, gravisensing is sensitive to protease treatment; Proteinase K, thermolysin and collagenase but not trypsin, alpha-chymotrypsin or carboxypeptidase B, inhibit gravisensing. These findings indicate that proteins in the cell-extracellular matrix junction may be required for gravisensing. Moreover, the tetrapeptide Arg-Gly-Asp-Ser (RGDS) inhibits gravisensing in a concentration-dependent manner, indicating that the gravireceptor may be an integrin-like protein. The macromolecules necessary for gravisensing have been localized to the cell ends. As a consequence of the exoplasmic site of action of the enzymes and the tetrapeptides, we interpret the results to mean that they are acting on the gravireceptor, although we cannot eliminate the possibility that they are acting on the signal transduction chain. On the whole, our observations indicate that the cell-extracellular matrix junction is a sine qua non for graviperception in statolith-free Chara internodal cells and we suggest that the gravireceptor is located in this region.
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PMID:The contribution of the extracellular matrix to gravisensing in characean cells. 152 45

Oversulphated chondroitin sulphate proteoglycan from squid skin was isolated from 4 M guanidine hydrochloride extract by ion-exchange chromatography, gel chromatography and density gradient centrifugation. The proteoglycan had Mr 3.5 x 10(5), contained on average six oversulphated chondroitin sulphate chains (Mr 4 x 10(4)) bound on a polypeptide of Mr 2.8 x 10(4), and oligosaccharides consisting of both hexosamines, glucuronic acid, sulphates and fucose as the only neutral monosaccharide. The major amino acids of the proteoglycan protein core are glycine (corresponding to about one third of the total amino acids), aspartic acid/asparagine and serine, together amounting to 50% of the total. The proteoglycan was resistant to the proteolytic enzymes V8 protease, trypsin (treated with diphenylcarbamoyl chloride), alpha-chymotrypsin and pronase, while it was completely degraded by papain and to a large extent by collagenase. Pretreated proteoglycan with chondroitinase AC was degraded by pronase to a large extent and slightly by V8 protease and trypsin. The proteoglycan did not interact with hyaluronic acid and did not form self-aggregates. Oversulphated chondroitin sulphate chains were composed of unusual sulphated disaccharide units which were isolated and characterized by HPLC. In particular, it contained 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose 4-sulphate (delta di-4S) and disulphated disaccharides (delta di-diS) [90% 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid 2/3-sulphate)-D-galactose 6-sulphate (delta di-diSD) and 10% 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid 2/3-sulphate)-D-galactose 4-sulphate (delta di-diSK)] as the major disaccharides, significant amounts of trisulphated disaccharides (delta di-triS) and small amounts of 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose 6-sulphate (delta di-6S) and 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose (delta di-OS). Trisulphated disaccharides contained sulphate groups at C-4 and C-6 of the galactosamine and at C-2 or C-3 of the glucuronic acid. By HPLC analysis of a pure preparation of oversulphated chondroitin sulphate, it was found that it contains glucose, galactose, mannose and fucose most likely as branches.
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PMID:Isolation, characterization and properties of the oversulphated chondroitin sulphate proteoglycan from squid skin with peculiar glycosaminoglycan sulphation pattern. 154 Dec 70


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