Comparative analysis of the kinetic parameters and thermal stability of two matrix metalloproteinases expressed in the developing sea urchin embryo

The extracellular matrix is now recognized as a dynamic structure which inf luences cellular properties. Many matrix metalloproteinase activities have been identified and characterized in vertebrates and constitute important a gents in controlling the composition of the extracellular matrix. We have b egun a study of matrix metalloproteinase activities in the developing sea u rchin embryo. Using sea urchin peristome collagen or gelatin as physiologic al substrates we have determined the kinetic parameters, K-m and V-max, for an 87 kDa gelatinase activity expressed in late stage sea urchin embryos. We also determined the kinetic parameters K-m, V-max and k(cat), for a 41 k Da species, expressed in the early sea urchin embryo, which possesses both collagenase and gelatinase activities. All values determined were similar t o those reported in the literature for vertebrate collagenases and gelatina ses and K-m values in the micromolar range suggest that bot species possess physiologically relevant activities. Both activities have previously bee s hown to require Ca2+ for activity. Using an assay for quantitating the clea vage of gelatin into trichloroacetic acid soluble peptides were report here markedly different effects of Ca2+ on the thermal denaturation profiles of the gelatinases. This latter finding may be indicative of different modes of action for this activating cation. Collectively these results demonstrat e both similarities and differences between vertebrate and invertebrate sea urchin gelatinases. (C) 1999 Elsevier Science Ltd. All rights reserved.