INVOLVEMENT OF A REGION NEAR VALINE-69 OF TISSUE INHIBITOR OF METALLOPROTEINASES (TIMP)-1 IN THE INTERACTION WITH MATRIX METALLOPROTEINASE-3 (STROMELYSIN-1)

Tissue inhibitors of metalloproteinases (TIMPs) inhibit matrix metallo proteinases (MMPs) by forming a 1:1 stoichiometric complex, but the in hibition mechanism of these inhibitors is not known. Here we have inve stigated the reactive site of TIMP-1 by its proteinase susceptibility before and after forming a complex with MMP-3 (stromelysin 1). When TI MP-1 was allowed to react with human neutrophil elastase, its inhibito ry activity was destroyed. This resulted from cleavage of the Val(69)- Cys(70) bond. However, cleavage of this bond by neutrophil elastase wa s prevented when TIMP-1 formed a complex with the catalytic domain of MMP-3, and full TIMP-1 activity was restored after dissociation of the complex at pH 3.0 in the presence of EDTA. These results indicate tha t the region around Va(169) closely associates with an active MMP; The three-dimensional structure of the N-terminal domain of TIMP-2 elucid ated by NMR studies [Williamson, Martorell, Carr, Murphy, Docherty, Fr eedman and Feeney (1994) Biochemistry 33, 11745-11759] reveals that. V a(169) and Cys(70) form part of an extended ridge that also includes t he N-terminal section of the inhibitor. This region is probably involv ed in the interaction with the catalytic domains of MMPs.