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Biochem J. 2005 Nov 15;392(Pt 1):127-34.

Functional basis for the overlap in ligand interactions and substrate specificities of matrix metalloproteinases-9 and -2.

Author information

1
Department of Periodontics, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, MC 7894, San Antonio, TX 78229-3900, USA.

Abstract

The MMPs (matrix metalloproteinases) MMP-9 and -2 each possess a unique CBD (collagen-binding domain) containing three fibronectin type II-like modules. The present experiments investigated whether the contributions to ligand interactions and enzymatic activities by the CBD of MMP-9 (CBD-9) corresponded to those of CBD in MMP-2 (CBD-2). The interactions of recombinant CBD-9 with a series of collagen types and extracellular matrix molecules were characterized by protein-protein binding assays. CBD-9 bound native and denatured type I, II, III, IV and V collagen, as well as Matrigel and laminin, with apparent K(d) values of (0.1-6.8)x10(-7) M, which were similar to the K(d) values for CBD-2 [(0.2-3.7)x10(-7) M]. However, CBD-9 bound neither native nor denatured type VI collagen. We also generated two modified MMPs, MMP-9(E402A) and MMP-2(E404A), by site-specific mutations in the active sites to obtain enzymes with intact ligand binding, but abrogated catalytic properties. In subsequent competitive binding assays, CBD-9 and MMP-9(E402A) inhibited the interactions of MMP-2(E404A) and, conversely, CBD-2 and MMP-2(E404A) competed with MMP-9(E402A) binding to native and denatured type I collagens, pointing to shared binding sites. Importantly, the capacity of CBD-9 to disrupt the MMP-9 and MMP-2 binding of collagen translated to inhibition of the gelatinolytic activity of the enzymes. Collectively, these results emphasize the essential contribution of CBD-9 to MMP-9 substrate binding and gelatinolysis, and demonstrate that the CBDs of MMP-9 and MMP-2 bind the same or closely positioned sites on type I collagen.

PMID:
16008524
PMCID:
PMC1317671
DOI:
10.1042/BJ20050650
[Indexed for MEDLINE]
Free PMC Article

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