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)

Human platelets were surface-labeled by the periodate/NaB3H4 method or by lactoperoxidase-catalysed iodination with 125I. The labeled platelets were treated with chymotrypsin under conditions known to give platelets which aggregate with fibrinogen without stimulation with ADP. Platelets and supernatant were then analysed by various gel electrophoretic techniques including isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing or non-reducing conditions and two-dimensional non-reduced/reduced sodium dodecyl sulfate polyacrylamide gel electrophoresis followed by fluorography or indirect autoradiography. Chymotrypsin-treatment of surface-labeled platelets degraded the major glycoproteins Ib, IIb and IIIa but also GP120(4.9-5.4), GPIc and GPV. The membrane-bound fragments of GPIb, IIb and IIIa could be identified and also the supernatant fragments of GPIb and GPV. GPIIIa was also cleaved within a loop structure formed by disulfide bond(s). The fact that remnants of both GPIIb and IIIa are left on chymotrypsin-treated platelets which aggregate spontaneously with fibrinogen may indicate that a complex formed by these remnants constitutes the fibrinogen-binding site on platelets.
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PMID:Identification and characterization of fragments of major glycoproteins from platelet membrane after chymotrypsin treatment. 397 99

Fusion of splenic lymphocytes from Lewis rats, immunized with affinity-purified estrogen receptor from the cytosol of MCF-7 human breast cancer cells, with two different mouse myeloma lines, has provided 13 monoclonal hybridoma lines secreting antiestrophilin antibodies, each of which (with one possible exception) recognizes a different antigenic determinant in the human receptor molecule. Of this library of monoclonal antibodies, some react with estrophilin from all sources tested, some react with mammalian but not avian receptors, whereas one preparation appears specific for estrophilin from primate sources. By proteolytic digestion under controlled conditions with mercury-deactivated papain, chymotrypsin, and trypsin, respectively, it is possible to remove sequentially the determinants recognized by one, two or three of the monoclonal antibodies, leaving the epitopes for the six remaining antibodies investigated on the steroid-binding portion of the receptor. The proteolytic fragment containing the epitope most readily removed (by mercuripapain) also contains the DNA-binding domain of the activated receptor molecule. Immunocytochemical staining, using the peroxidase procedure with various monoclonal antibody preparations, of frozen sections of human breast cancer tissue, fixed in ethanol or in picric acid-formaldehyde reagent, shows clearly that the majority of the native receptor, which appears in the cytosol after tissue homogenization, is actually localized within the nuclear compartment in the intact cell. The immunocytochemical technique also permits the identification of mixed populations of receptor-containing and non-containing cells in human breast cancers.
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PMID:Immunochemical studies of estrogen receptors. 620 Jul

We have demonstrated that incubation of rat liver microsomes with N-hydroxy-2-acetylaminofluorene (N-OH-AAF) leads to formation of a 2-nitrosofluorene-membrane lipid adduct. This adduct exists as a nitroxyl free radical, termed N-O-LAF, in its oxidized state. When microsomes were incubated with the sulfhydryl binding agent, rho-hydroxymercuribenzoate, a larger amount of N-OL-LAF formed. We interpret this as a slowdown in the rate of endogenous chemical reduction of carcinogen-membrane lipid adduct. In this paper we present evidence that N-OH-AAF is deacetylated by a microsomal enzyme to form N-hydroxy-2-aminofluorene and this is then oxidized to 2-nitrosofluorene which adds covalently to membrane lipid double bonds to form N-O-LAF. Various antioxidants, peroxidase inhibitors, and P450 substrates and inhibitors were ineffective in altering the amount of N-O-LAF formed from N-OH-AAF; but two esterase inhibitors, dietyl-rho-nitrophenylphosphate and alpha-toluene-sulfonyl fluoride, prevented N-O-LAF formation. Of the following purified enzymes tested: porcine liver carboxyl esterase, pepsin, chymotrypsin, cathepsin D, ficin, papain, leucine aminopeptidase, Naja naja phospholipase, acetylcholinesterase (type I), trypsin (type I and V) and epoxide hydrase; only carboxyl esterase was effective in deacetylating N-OH-AAF.
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PMID:The deacetylation of N-hydroxy-2-acetylaminofluorene by rat liver microsomes and carboxyl esterase. 626 Mar 32

The interaction of human blood platelets with collagenase-treated rabbit subendothelium was studied by histochemical ultrastructural methods and by morphometric semi-quantitative analysis. Aortas were deendothelialized and incubated: 1) with a highly purified bacterial collagenase whose specificity was controlled; and 2) with the same collagenase followed by chymotrypsin. For histochemical studies, tannic acid, ruthenium red, and peroxidase-labeled Ricinus communis and concanavalin A were used. Electron microscopy showed that after digestion of fibrillar collagen by collagenase, adherent and aggregated platelets were observed on Ricinus communis-, concanavalin A-, and ruthenium red-positive glycoprotein microfibrils. After successive incubation with collagenase and chymotrypsin, the microfibrils disappeared. No platelets were observed on the remnant amorphous elastin. Morphometric analysis confirmed the interaction of platelets with collagenase-treated subendothelium. In addition, glycoproteins were extracted from collagenase-treated rabbit aortas using 5 M guanidine. Using an in vitro quantitative test, significant platelet adhesion to these glycoproteins was observed. Our results show an interaction between platelets and noncollagenic glycoprotein microfibrils.
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PMID:Histochemical and ultrastructural characterization of subendothelial glycoprotein microfibrils interacting with platelets. 627 53

The borate-insoluble chitin-protein complex, CB-I, from prepupal sarcophagid larvae was cleaved with chymotrypsin and trifluoromethanesulfonic acid releasing a polypeptide fragment of Mr 68 000. The intact glycoprotein was blocked at the C terminus; the N-terminal sequence of Asp-Val-Ala-His-Tyr was not homologous with seven of the borate-soluble nonglycosylated structural proteins. Bityrosine was identified as a component of the primary chain, both half-residues occupied in peptide linkages. Sclerotization initiated a decline in bityrosine coincident with the addition of soluble proteins to the tanned matrix. The chitin-protein complex also included bound peroxidase, propolyphenol oxidase, and an o-diphenol subject to oxidation on activation of the zymogen. In the course of the oxidation N termini declined in accordance with the formation of 1,4 quinonoid cross-links.
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PMID:Chitin-bound protein of sarcophagid larvae: metabolism of covalently linked aromatic constituents. 629 68

A beta 1-serum component, beta 1-anticollagenase, capable of inhibiting various mammalian tissue collagenases, was isolated from human plasma by gel filtration, affinity chromatography and ion-exchange chromatography. The inhibitor contains 1-2 free sulfhydryl groups, which are a prerequiste for inhibitory activity and for binding to the thiol-Sepharose affinity support. Alkylation of beta 1-anticollagenase by iodoacetamide blocks inhibitory activity. The inhibitor was purified to apparent homogeneity and exhibited a Mr = 30500 determined by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The amino acid and carbohydrate composition was determined. According to its composition and the isoelectric focussing beta 1-anticollagenase is an acidic protein with an isoelectric point of 5.6. Inhibition of human leukocyte collagenase proceeds in a strong 1 : 1 stoichiometric reaction. The mechanism of this association takes place by a disulfide/thiol interchange reaction as has been previously indicated for human leukocyte collagenases in forming the latent enzyme [Macartney, H. W. and Tschesche, H. (1980) FEBS Lett. 119, 327-332]. The beta 1-anticollagenase--leukocyte-collagenase complex (latent enzyme) is activatable by disulfide-containing compounds such as cystine, oxidised glutathione, insulin, relaxin, trypsinogen and others, but not by 179,203-di(S-carboxymethyl)trypsinogen, or its trypsin derivative. Compounds containing inaccessible disulfide bonds, e.g. chymotrypsin, or sulfhydryl groups, e.g. D-penicillamine, do not activate the complex. Activation is, however, easily obtained with the oxidised-glutathione-generating system myeloperoxidase/H2O2/glutathione as was previously demonstrated for the human leukocyte latent collagenase activatable in a phagocytosis-simulated respiratory burst [Tschesche, H. and Macartney, H. W. (1981) Eur. J. Biochem. 120, 183-190].
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PMID:Characterisation of beta 1-anticollagenase from human plasma and its reaction with polymorphonuclear leukocyte collagenase by disulfide/thiol interchange. 629 99

Two cytochemical methods for detection of granulocytic elastase and chymotrypsin employing alanine and phenylalanine naphthyl esters were developed. Specificity of reaction with the ester substrates was proven by chloromethyl ketone inhibitors. The results of both staining methods were almost identical with the staining for naphthol AS-D chloroacetate (Cl Ac-O Nap AS-D) esterase, since Cl Ac-O Nap AS-D also reacts with granulocyte elastase and chymotrypsin. Mature neutrophils and myeloid precursors except myeloblasts are stained with all three substrates in peripheral blood and bone marrow. Mast cells, however, only react with Cl Ac-O Nap AS-D and the chymotrypsin substrate and not with the elastase substrate. In acute myeloid leukemia the three esterases appear in parallel at a somewhat later stage of maturation than myeloperoxidase. In blood smears from 380 hospital patients no hereditary elastase or chymotrypsin deficiency could be demonstrated. Staining for elastase and chymotrypsin was also normal in hereditary myeloperoxidase deficiency and chronic granulomatous disease. On the other hand 6% of the hospital patients and about two-thirds of patients with acute myeloid leukemia showed a partial elastase deficiency in more than 25% of the peripheral neutrophils.
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PMID:Cytochemical determination of granulocyte elastase and chymotrypsin in human myeloid cells and its application in acquired deficiency states and diagnosis of myeloid leukemia. 630 May 11

F (fusion) and HANA (hemagglutinin and neuraminidase) glycoproteins of HVJ (Sendai virus) were purified and characterized. The NH2-terminal hydrophobic region of the F1 (larger) subunit of F (fusion)-glycoprotein seems to be required for the hemolytic and cell fusion-inducing activity of the virus for the following reasons. (1) Selective splitting off of a 2,500-3,500 dalton segment from the NH2-terminal region of F1 by chymotrypsin or thermolysin resulted in inactivation of the biological activities of HVJ. (2) At least a part of this region may be exposed to the surrounding medium, since it is preferentially iodinated and is easily split by aminopeptidase M, chymotrypsin, and thermolysin. Tryptic digestion, which does not remove the NH2-terminal region but produce nicking of F1 subunit to subfragments F1a (larger one) and F1b (smaller one), resulted in substantial structural changes evidenced by circular dichroism measurement and iodination by lactoperoxidase method. Trypsin-digested F seems to have the NH2-terminal hydrophobic region buried within hydrophobic interior of the protein (or in the lipid bilayers). Based on these and other results, we propose a hypothesis featuring direct interaction of the hydrophbic region with the lipid bilayers of the target-cell membrane as an important step in fusion reactions between the viral envelope and cell membranes.
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PMID:Viral proteins in cell fusion. 631 Aug 22

The extracellular matrix of cultured chicken embryo fibroblasts undergoes a number of modifications during the early stages of oncogenic transformation. One alteration is increased production of a small protein (Mr approximately 21,000) which is transiently deposited in the matrix by transforming cells infected with LA24, a temperature-sensitive mutant of Rous sarcoma virus (RSV) (Blenis, J., and Hawkes, S.P. (1983) Proc. Natl. Acad. Sci. U.S.A. 80, 770-774). This protein is a major component of substratum-associated material (material which remains attached to culture dishes after removal of cells with ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid). Its synthesis is stimulated by transformation of cells with NY68, another ts mutant of RSV, and also by treatment of normal, uninfected cells with the tumor promoter, phorbol myristate acetate. Accessibility of the 21-kDa protein to lactoperoxidase-catalyzed iodination indicates an exposed location within the matrix. The protein binds strongly to the culture dish and/or other matrix components. This interaction can be disrupted by sodium dodecyl sulfate but not by several nonionic detergents, unless beta-mercaptoethanol or KCl (0.5 M) are also present. High concentrations of urea or guanidine hydrochloride also remove the protein from the matrix. The 21-kDa protein is resistant to trypsin, collagenase, and the hydrolytic enzymes associated with cells transformed by the wild-type Prague A RSV but not to Pronase or chymotrypsin. A 21-kDa protein with properties similar to those described above is also detected in the medium and binds to the matrix, suggesting that a potential route of deposition of the 21-kDa protein in the matrix may be via shedding and subsequent interaction with other matrix components.
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PMID:Characterization of a transformation-sensitive protein in the extracellular matrix of chicken embryo fibroblasts. 643 99

Whole cells and isolated outer membranes (OMs) of four strains of gonococci were surface radioiodinated with either lactoperoxidase or Iodogen (Pierce Chemical Co., Rockford, Ill.). These preparations were solubilized in sodium dodecyl sulfate and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Surface-radioiodinated protein I (PI) and PIII bands were excised from the sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and digested with alpha-chymotrypsin, and the resultant 125I-peptide fragments were resolved by high-voltage electrophoresis and thin-layer chromatography (i.e., surface peptide mapping). Radioemitting peptidic fragments were visualized by autoradiography. Results demonstrated that the PI molecule of each gonococcal strain studied had unique iodinatable peptides exposed on the surface of whole cells and OMs, whereas PIIIs appeared to have the same portion of the molecule exposed on the surface of bacteria or OMs, regardless of the gonococcal strain from which they were isolated. Many more radiolabeled peptides were seen in surface peptide maps of PIs from radiolabeled OMs than in those from radioiodinated whole cells, whereas different peptidic fragments were seen in the surface peptide maps of PIIIs from radiolabeled OMs than were seen in those from radiolabeled whole cells. These data suggest that PI may contribute strain-specific antigenic determinants and PIII may contribute cross-reactive determinants and that the surface exposure of PI and PIII is different in isolated OMs than in the OM of intact gonococci.
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PMID:Surface peptide mapping of protein I and protein III of four strains of Neisseria gonorrhoeae. 681 36


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