SOLUTION STRUCTURE OF THE CATALYTIC DOMAIN OF HUMAN STROMELYSIN COMPLEXED WITH A HYDROPHOBIC INHIBITOR

Stromelysin, a representative matrix metalloproteinase and target of d rug development efforts, plays a prominent role in the pathological pr oteolysis associated with arthritis and secondarily in that of cancer metastasis and invasion. To provide a structural template to aid the d evelopment of therapeutic inhibitors, we have determined a medium-reso lution structure of a 20-kDa complex of human stromelysin's catalytic domain with a hydrophobic peptidic inhibitor using multinuclear, multi dimensional NMR spectroscopy. This domain of this zinc hydrolase conta ins a mixed beta-sheet comprising one antiparallel strand and four par allel strands, three helices, and a methionine-containing turn near th e catalytic center. The ensemble of 20 structures was calculated using , on average, 8 interresidue NOE restraints per residue for the 166-re sidue protein fragment complexed with a 4-residue substrate analogue. The mean RMS deviation (RMSD) to the average structure for backbone he avy atoms is 0.91 Angstrom and for all heavy atoms is 1.42 Angstrom. T he structure has good stereochemical properties, including its backbon e torsion angles. The beta-sheet and alpha-helices of the catalytic do mains of human stromelysin (NMR model) and human fibroblast collagenas e (X-ray crystallographic model of Lovejoy B et al., 1994b, Biochemist ry 33:8207-8217) superimpose well, having a pairwise RMSD for backbone heavy atoms of 2.28 Angstrom when three loop segments are disregarded . The hydroxamate-substituted inhibitor binds across the hydrophobic a ctive site of stromelysin in an extended conformation. The first hydro phobic side chain is deeply buried in the principal S'(1) subsite, the second hydrophobic side chain is located on the opposite side of the inhibitor backbone in the hydrophobic S'(2) surface subsite, and a thi rd hydrophobic side chain (P'(3)) lies at the surface.