TIMP-1 contact sites and perturbations of stromelysin 1 mapped by NMR and a paramagnetic surface probe
TIMP-1 contact sites and perturbations of stromelysin 1 mapped by NMR and a paramagnetic surface probe
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DOI：
10.1021/bi980128h
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发表时间：
1998-07-07
期刊：
影响因子：
2.9
通讯作者：
Van Doren, SR
中科院分区：
文献类型：
--
作者：
Arumugam, S;Hemme, CL;Van Doren, SR
Surfaces of the 173 residue catalytic domain of human matrix metalloproteinase 3 (MMP-3(Delta C)) affected by binding of the N-terminal, 126 residue inhibitory domain of human TIMP-1 (N-TIMP-1) have been investigated using an amide-directed, NMR-based approach. The interface was mapped by a novel method that compares amide proton line broadening by paramagnetic Gd-EDTA in the presence and absence of the binding partner. The results are consistent with the X-ray model of the complex of MMP-3(Delta C) with TIMP-1 (Gomis-Ruth et al. (1997) Nature 389, 77-81). Residues Tyr155, Asn162, Val163, Leu164, His166, Ala167, Ala169, and Phe210 of MMP-3(Delta C) ate protected from broadening by the Gd-EDTA probe by binding to N-TIMP-1. N-TIMP-1-induced exposure of backbone amides of Asp238, Asn240, Gly241, and Ser244 of helix C of MMP-3(Delta C) to Gd-EDTA confirms that the displacement of the N-terminus of MMP-3(Delta C) occurs not only in the crystal but also in solution. These results validate comparative paramagnetic surface probing as a means of mapping protein-protein interfaces. Novel N-TIMP-1-dependent changes in hydrogen bonding near the active site of MMP-3(Delta C) are reported. N-TIMP-1 binding causes the amide of Tyr223 of MMP-3(Delta C;) bound by N-TIMP-1 to exchange with water rapidly, implying a lack of the hydrogen bond observed in the crystal structure. The backbone amide proton of Asn162 becomes protected from rapid exchange upon forming a complex with N-TIMP-1 and could form a hydrogen bond to N-TIMP-1. N-TIMP-1 binding dramatically increases the rate of amide hydrogen exchange of Asp177 of the fifth beta strand of MMP-3(Delta C), disrupting its otherwise stable hydrogen bond.
