Localization of cleavage sites for human fibroblast collagenase in the bait region of five mammalian alpha-macroglobulins.
The interaction between human fibroblast collagenase and five mammalian alpha-macroglobulins (human alpha 2-macroglobulin and pregnancy zone protein, rat alpha 1- and alpha 2-macroglobulin and rat alpha 1-inhibitor 3) is discussed. Complex formation involves specific limited proteolysis in the alpha-macroglobulin bait regions, activation of the internal beta-cysteinyl-gamma-glutamyl thiol esters, conformational changes and covalent complex formation. For human alpha 2-macroglobulin, and rat alpha 1-macroglobulin and alpha 2-macroglobulin it is estimated that the overall rate constant of complex formation is greater than 1.10(6) M-1s-1, while it is much lower for human pregnancy zone protein and rat alpha 1-inhibitor 3. More than 95% of the complexed collagenase is covalently bound. The identification of the sites of specific limited proteolysis in the bait regions of the five alpha-macroglobulins shows that cleavage may take place in sequences that are not related to those identified earlier in the collagens. These results greatly expand the repertoire of sequences known to be cleaved by fibroblast collagenase, and suggest that this proteinase has a primary substrate specificity resembling that of the microbial proteinase thermolysin as it preferentially cleaves at the NH2-terminal side of large hydrophobic residues. In addition, the results highlight the unique structure of the flexible alpha-macroglobulin bait region in that it can accommodate a conformation required by the highly restrictive fibroblasts collagenase. It is suggested that alpha-macroglobulins may play an important role in locally controlling the activity of collagenases and perhaps other proteinases of the extracellular matrix.
D000595:Amino Acid Sequence; D000818:Animals; D017364:Collagenases / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology*; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D011485:Protein Binding; D051381:Rats; D016415:Sequence Alignment; D013379:Substrate Specificity; D000511:alpha-Macroglobulins / Q000378:metabolism* / Q000502:physiology
D000511:alpha-Macroglobulins; D017364:Collagenases