Purification and characterization of human 92-kDa type IV collagenase (gelatinase B)

Abstract

Human 92-kDa type IV collagenase (gelatinase B), a family of matrix metalloproteinases (MMP), play important roles in the degradation of the basement membrane and the migration of leukocytes and metastatic tumor cells during inflammation and invasion. To investigate the biochemical and enzymatic characteristics of human neutrophil type IV collagenase, the enzyme was extracted from human leukocytes and purified by a combination of Ultrogel AcA 54 and Bio-Rex 70 chromatographies. The purified enzyme showed a single band of molecular weight of 92 kDa on SDS-PAGE. Human neutrophil type IV collagenase degraded gelatin by cleaving the specific sites, but did not affect intact type I collagen. Human 92-kDa type IV collagenase activity was inhibited by EGTA, EDTA and tetracycline. Tetracycline showed the strongest inhibitory effect on the gelatinolytic activity of the 92-kDa type IV collagenase. These inhibitory effects may be due to the chelation effect of these agents since 92-kDa type IV collagenase is a metalloenzyme.