Interaction of human rheumatoid synovial collagenase (matrix metalloproteinase 1) and stromelysin (matrix metalloproteinase 3) with human alpha 2-macroglobulin and chicken ovostatin. Binding kinetics and identification of matrix metalloproteinase cleavage sites.
The homologous proteinase inhibitors, human alpha 2-macroglobulin (alpha 2M) and chicken ovostatin, have been compared with respect to their "bait" region sequences and interactions with two human matrix metalloproteinases, collagenase and stromelysin. A stretch of 34 amino acid residues of the ovostatin bait region sequence was determined and the matrix metalloproteinase cleavage sites identified. Collagenase cleaved a X-Leu bond where X was unidentified, whereas the major cleavage site by stromelysin was at the Gly-Phe bond, 4 residues on the COOH-terminal side of the collagenase cleavage site. Collagenase cleaved the alpha 2M bait region at the Gly679-Leu680 bond, and stromelysin at Gly679-Leu680 and Phe684-Tyr685 bonds. Sequence similarity in the bait region of members of the alpha-macroglobulin family is strikingly low. The kinetic studies indicate that alpha 2M is a 150-fold better substrate for collagenase than type I collagen. Structural predictions based on the bait region sequences suggest that a collagen-like triple helical structure is not a prerequisite for the efficient binding of tissue collagenase to a substrate. The binding of stromelysin to alpha 2M is slower than that of collagenase. Stromelysin reacts with ovostatin even more slowly. Despite the preference of chicken ovostatin for metalloproteinases, human alpha 2M, a far less selective inhibitor, reacts more rapidly with collagenase and stromelysin. These results suggest that alpha 2M may play an important role in regulating the activities of matrix metalloproteinases in the extracellular space.
Enghild
J J
JJ
Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710.
Salvesen
G
G
Brew
K
K
Nagase
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001172:Arthritis, Rheumatoid / Q000201:enzymology*; D002478:Cells, Cultured; D002645:Chickens; D015387:Egg Proteins, Dietary / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008259:Macroglobulins; D019278:Matrix Metalloproteinase 3; D008666:Metalloendopeptidases / Q000378:metabolism*; D003012:Microbial Collagenase / Q000378:metabolism*; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000378:metabolism*; D011485:Protein Binding; D013045:Species Specificity; D013582:Synovial Fluid / Q000201:enzymology*; D000511:alpha-Macroglobulins / Q000378:metabolism*
D015387:Egg Proteins, Dietary; D008259:Macroglobulins; D011480:Protease Inhibitors; D000511:alpha-Macroglobulins; C036828:ovomacroglobulin; D008666:Metalloendopeptidases; D019278:Matrix Metalloproteinase 3; D003012:Microbial Collagenase