Revised Manuscript Received November 10, 1994s abstract: Human 72 kDa type IV collagenase (gelatinase A, MMP-2) was expressed in a baculovirus/insect cell system. The enzyme was produced in the wild-type form and in two mutant forms, where the active site GIU375 was substituted by Asp or Gin. The mutated proteins had strongly reduced or no detectable activity, respectively, allowing detailedanalysis of rapid autoactivation reactions. MMP-2 was readily degraded to a proenzyme form lacking the first fouramino acid residues. This cleavage was shown to be an autolytic process, although enzyme activity was apparently not affected by this truncation. Conversion to the active enzyme form was achieved without external activator in a concentration-dependent manner at 37 C. The activation of MMP-2 was shown to be a stepwise process, probablyvia a A1-50 form as a highly unstable intermediate. The C-terminal hemopexin-like domain is removed rather early at two cleavage sites, and degradation within the Zn-binding site inactivates the enzyme. The fibronectin-and hemopexin-like domains are stable, although the autodegradation pattern did not show any sequence specificity, except for charged residues in the Pi'position. The results indicate that a specific activator may not be essential for MMP-2.

Connective tissue turnover in physiological and pathological conditions is highly dependent on the action of matrix metalloproteases (MMPs). Matrix metalloproteases are a family of homologous extracellular enzymes which are secreted in latent forms and which have a Zn atom at the catalytic site (Matrisian, 1992). The matrix metalloprotein-ases can degrade various components of the extracellular matrix, and they have been subclassified on the basis of their substrate specificity as determined in vitro into interstitial