Elastin Degradation by Matrix Metalloproteinases

Abstract

Insoluble elastin was used as a substrate to character- ize the peptide bond specificities of human (HME) and mouse macrophage elastase (MME) and to compare these enzymes with other mammalian metalloprotein- ases and serine elastases. New amino termini detected by protein sequence analysis in insoluble elastin follow- ing proteolytic digestion reveal the P1 residues in the carboxyl-terminal direction from the scissile bond. The relative proportion of each amino acid in this position reflects the proteolytic preference of the elastolytic en- zyme. The predominant amino acids detected by protein sequence analysis following cleavage of insoluble elas- tin with HME, MME, and 92-kDa gelatinase were Leu, Ile, Ala, Gly, and Val. HME and MME were similar in their substrate specificity and showed a stronger pref- erence for Leu/Ile than did the 92-kDa enzyme. Fibro- blast collagenase showed no activity toward elastin. The amino acid residues detected in insoluble elastin follow- ing hydrolysis with porcine pancreatic elastase and hu- man neutrophil elastase were predominantly Gly and Ala, with lesser amounts of Val, Phe, Ile, and Leu. There were interesting specificity differences between the two enzymes, however. For both the serine and matrix met- alloproteinases, catalysis of peptide bond cleavage in insoluble elastin was characterized by temperature ef- fects and water requirements typical of common en- zyme-catalyzed reactions, even those involving soluble substrates. In contrast to what has been observed for collagen, the energy requirements for elastolysis were not extraordinary, consistent with cleavage sites in elas- tin being readily accessible to enzymatic attack.