Engineering of Selective TIMPs

Abstract

Differences in proteinase susceptibility between free TIMP‐1 and the TIMP‐1‐MMP‐3 complex and mutagenesis studies suggested that the residues around the disulfide bond between Cys1 and Cys70 in TIMP‐1 may interact with MMPs. The crystal structure of the complex between TIMP‐1 and the catalytic domain of MMP‐3 has revealed that the α‐amino group of Cys1 bidentately chelates the catalytic zinc of MMP‐3 and the Thr2 side chain occupies the S₁′ pocket. Generation of the N‐terminal domain of TIMP‐1 (N‐TIMP‐1) variants with 15 different amino acid substitutions for Thr2 has indicated that the nature of the side chain of residue 2 has a major effect on the affinity of N‐TIMP‐1 for three different MMPs (MMPs‐1, ‐2 and ‐3). The results also demonstrate that the mode of binding of N‐TIMP‐1 residue 2 differs from the binding of the P₁′ residue of a peptide substrate.