PURIFICATION AND PROPERTIES OF EXTRACELLULAR MATRIX-DEGRADING METALLOPROTEINASE OVERPRODUCED BY ROUS-SARCOMA VIRUS-TRANSFORMED RAT-LIVER CELL-LINE, AND ITS IDENTIFICATION AS TRANSIN

Rous sarcoma virus-transformed rat liver cell line RSV-BRL secreted a neutral proteinase in a latent precursor form with a molecular weight (Mr) of 57,000 (57k) as a major secreted protein. This enzyme was a calcium-dependent metallo-proteinase. The proenzyme was purified from the serum-free conditioned medium of the transformed cells by affinity chromatographies on a zinc chelate Sepharose column and a reactive red agarose column. When activated by treatment with trypsin or p-aminophenylmercuric acetate (APMA) in the presence of Ca2+, the purified enzyme effectively hydrolyzed casein, fibronectin, and laminin. Type IV collagen was hydrolyzed at 37°C but not at 30°C by the enzyme, whereas type I and type III collagens were hardly hydrolyzed even at 37°C. The treatment with trypsin or AMP A in the presence of Ca2+ converted this 57k proenzyme to an active and stable enzyme with Mr. In the absence of Ca2+, however, APMA converted the proenzyme to an intermediate form with Mr 45k, while trypsin digested it to an inactive peptide with Mr 30k. These results demonstrate that calcium ion is essential for the activation, activity expression, and stabilization of this metallo-proteinase. Analysis of its partial amino acid sequence and amino acid composition showed that the 57k proenzyme was identical or closely related to the putative protein transin, a rat homologue of stromelysin. © 1990 Copyright, 1990 by the Journal of Biochemistry.