Evidence for de novo synthesis of collagenase by tadpole skin explant

Abstract

3-day culture medium from tadpole back skins incubated with 14C-labeled amino acid mixture contains labeled collagenase. The molecular weight of the labeled enzyme was estimated as approximately 40 000–43 000 by molecular-sieve chromatography and by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. The relative content of labeled collagenase was less than 60 % of the total enzyme which is released in an active form from a tadpole skin explant incubated with 14C-labeled amino acid mixture. This suggested that a significant amount of collagenase in the culture medium was derived from the preexisting enzyme, either in an active or inactive form in the tissue. Affinity chromatography on a collagen-polyacrylamide gel column, followed by gel filtration on a Sephadex G-75 superfine column, can be used for the isolation of collagenase in the pure form from the crude enzyme preparation.