Characterisation of beta 1-anticollagenase from human plasma and its reaction with polymorphonuclear leukocyte collagenase by disulfide/thiol interchange.
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].
D000596:Amino Acids / Q000032:analysis; D002241:Carbohydrates / Q000032:analysis; D004220:Disulfides / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007460:Iodoacetamide / Q000494:pharmacology; D003012:Microbial Collagenase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology; D009195:Peroxidase / Q000494:pharmacology; D013438:Sulfhydryl Compounds / Q000494:pharmacology
D000596:Amino Acids; D002241:Carbohydrates; D004220:Disulfides; D013438:Sulfhydryl Compounds; D006861:Hydrogen Peroxide; D009195:Peroxidase; D003012:Microbial Collagenase; D007460:Iodoacetamide