PMID: 8463259Apr 5, 1993Paper

Role of zinc-binding- and hemopexin domain-encoded sequences in the substrate specificity of collagenase and stromelysin-2 as revealed by chimeric proteins.

The Journal of Biological Chemistry
R Sanchez-LopezZ Werb

Abstract

The relationship of enzyme structure to substrate specificity for the matrix metalloproteinases interstitial collagenase and stromelysin-2 has been investigated by analysis of the cleavage specificity of recombinant human collagenase-stromelysin-2 hybrid proteins and C terminally truncated collagenase and stromelysin-2. Two series of chimeric proteins were devised by progressive substitution of exon-encoded domains. The recombinant proteins were expressed in COS-7 cells as protein A-fusion proteins and purified on an IgG affinity matrix. Treatment with 4-amino-phenylmercuric acetate released active metalloproteinase of the sizes predicted for the chimeric proteins. Active forms of both the chimeric protein series and the short form enzymes expressed both casein- and gelatin-degrading activities. Like stromelysin, the catalytic activity of stromelysin-2 was contained in the N-terminal domain (encoded by exons 1-5) and was apparently independent of the C-terminal domain (encoded by exons 6-10). Only full-length collagenase displayed a triple helicase (collagenolytic) activity; no combination of N- or C-terminal collagenase domains fused with stromelysin-2 domains had such activity. This suggests that the triple helicase activity ...Continue Reading

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