Characterization of the individual collagenases from Clostridium histolyticum.
The six collagenases (alpha, beta, gamma, delta, epsilon, and zeta) from Clostridium histolyticum isolated in the preceding paper [Bond, M. D., & Van Wart, H. E. (1984) Biochemistry (first paper of three in this issue)] have been characterized in detail. The molecular weights determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis range from 68 000 to 125 000. Isoelectric focusing experiments demonstrate that the isoelectric points of the collagenases are in the 5.35-6.20 range. These experiments also reveal that the subspecies of alpha- and gamma-collagenases (alpha1 vs. alpha 2 and gamma 1 vs. gamma 2) have different isoelectric points but the same molecular weights. Microheterogeneity is also observed for the beta- and epsilon-collagenases. The amino acid compositions of all six collagenases have been determined, and analysis for neutral sugars and hexosamines shows that none of the enzymes have a significant carbohydrate content. Zinc and calcium are the only metals that copurify with the collagenases. The purified enzymes contain approximately 1 mol of zinc/mol of protein and a calcium content that varies from about 2 mol/mol for alpha-collagenase to about 7 mol/mol for beta-collagenase. All of the collagenases are 5-10 times more active against gelatin than collagen. The alpha-, beta-, and gamma-collagenases are significantly less active toward the synthetic peptide substrates examined than the delta-, epsilon, and zeta-collagenases. This property, taken together with data on the stabilities and amino acid compositions of these enzymes, strongly supports their assignment to two distinct classes. This establishes clearly that C. histolyticum does, indeed, produce more than one different type of collagenase.
D000596:Amino Acids / Q000032:analysis; D002413:Cations, Divalent; D003013:Clostridium / Q000201:enzymology*; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D037102:Lectins; D003012:Microbial Collagenase / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight
D000596:Amino Acids; D002413:Cations, Divalent; D007527:Isoenzymes; D037102:Lectins; D003012:Microbial Collagenase