High-Resolution Solution Structure of the Inhibitor-Free Catalytic Fragment of Human Fibroblast Collagenase Determined by Multidimensional NMR

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• 作者：Franklin J. Moy ; Pranab K. Chanda ; Scott Cosmi ; Michael R. Pisano ; Charlotte Urbano ; Jim Wilhelm ; and Robert Powers
• 刊名：Biochemistry
• 出版年：1998
• 出版时间：February 10, 1998
• 年：1998
• 卷：37
• 期：6
• 页码：1495 - 1504
• 全文大小：163K
• 年卷期：v.37,no.6(February 10, 1998)
• ISSN：1520-4995

文摘

The high-resolution solution structure of the inhibitor-freecatalytic fragment of human fibroblastcollagenase (MMP-1), a protein of 18.7 kDa, which is a member of thematrix metalloproteinase family,has been determined using three-dimensional heteronuclear NMRspectroscopy. A total of 30 structureswere calculated by means of hybrid distance geometry-simulatedannealing using a total of 3333experimental NMR restraints, consisting of 2409 approximate interprotondistance restraints, 84 distancerestraints for 42 backbone hydrogen bonds, 426 torsion anglerestraints, 125 ³J_{NHages/gifchars/alpha.gif" BORDER=0>}restraints, 153 Cages/gifchars/alpha.gif" BORDER=0>restraints, and 136 Cages/gifchars/beta2.gif" BORDER=0 ALIGN="middle"> restraints. The atomic rms distributionabout the mean coordinate positions forthe 30 structures for residues 7-137 and 145-163 is 0.42 ± 0.04Å for the backbone atoms, 0.80 ± 0.04Å for all atoms, and 0.50 ± 0.03 Å for all atoms excludingdisordered side chains. The overall structureof MMP-1 is composed of a ages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-sheet consisting of five ages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-strands in amixed parallel and anti-parallelarrangement and three ages/gifchars/alpha.gif" BORDER=0>-helices. A best-fit superposition of theNMR structure of inhibitor-free MMP-1with the 1.56 Å resolution X-ray structure by Spurlino et al.[Spurlino, J. C., Smallwood, A. M., Carlton,D. D., Banks, T. M., Vavra, K. J., Johnson, J. S., Cook, E. R., Falvo,J., and Wahl, R. C., et al. (1994)Proteins: Struct., Funct., Genet. 19, 98-109] complexedwith a hydroxamate inhibitor yields a backboneatomic rms difference of 1.22 Å. The majority of differencesbetween the NMR and X-ray structureoccur in the vicinity of the active site for MMP-1. This includesan increase in mobility for residues138-144 and a displacement for the Ca²⁺-loop (residues74-80). Distinct differences were observed forside-chain torsion angles, in particular, the ages/gifchars/chi.gif" BORDER=0 >₁ for N80is -60ages/entities/deg.gif"> in the NMR structure compared to 180ages/entities/deg.gif">in the X-ray. This results in the side chain of N80 occupying andpartially blocking access to the activesite of MMP-1.