Solution Structure of the Catalytic Domain of Human Stromelysin-1 Complexed to a Potent, Nonpeptidic Inhibitor

Abstract

The full three-dimensional structure of the catalytic domain of human stromelysin-1 (SCD) complexed to a novel and potent, nonpeptidic inhibitor has been determined by nuclear magnetic resonance spectroscopy (NMR). To accurately mimic assay conditions, the structure was obtained in Tris buffer at pH 6.8 and without the presence of organic solvent. The results showed that the major site of enzyme-inhibitor interaction occurs in the S1' pocket whereas portions of the inhibitor that occupy the shallow S2' and S1 pockets remained primarily solvent exposed. Because this relatively small inhibitor could not deeply penetrate stromelysin's long narrow hydrophobic S1' pocket, the enzyme was found to adopt a dramatic fold in the loop region spanning residues 221-231, allowing occupation of the solvent-accessible S1' channel by the enzyme itself. This remarkable conformational fold at the enzyme binding site resulted in constriction of the S1' loop region about the inhibitor. Examination of the tertiary structure of the stromelysin-inhibitor complex revealed few hydrogen-bonding or hydrophobic interactions between the inhibitor and enzyme that can contribute to overall binding energy; hence the resultant compact structure may in part account for the relatively high potency exhibited by this inhibitor.

The Design, Structure, and Therapeutic Application of Matrix Metalloproteinase Inhibitors

“…The hydroxamic acid group, found to be the most potent, coordinates the catalytic zinc in a bidentate manner adopting a trigonal bipyrimidal geometry [57,58]. The hydroxamate nitrogen is protonated in the bound state and forms a hydrogen bond with the carbonyl oxygen of the enzyme's conserved alanine (Ala 165 ) [59]. These features establish a near perfect coordination geometry of the hydroxamic acid with the catalytic zinc.…”

“…The non-peptidic inhibitors, whose structures were determined in complex with various MMPs, were either sulfonamides, phosphinamides or sulfones with hydroxamic acid or carboxylate zinc-binding functions [59,68,69,78,79]. The first reported structure of a non-peptidic sulfonamide hydoxamic acid inhibitor was CGS 27023A (17; see Table 8 below) in complex with MMP-3 [59,93], Fig. (6).…”

“…A para-(biphenyl)ethyl moiety is positioned to allow deep penetration into the S 1ṕ ocket. The mode of binding of CGS 27023A (17) to MMP-3 as well as the full 3-dimensional solution structure of the catalytic domain of MMP-3 complexed with CGS 27023A have been determined by NMR spectroscopy [59,93,140,141]. The para-methoxyphenyl substituent occupies, but does not fill, the S 1´ specificity pocket while the isopropyl and pyridylmethyl substituents occupy the S 1 and S 2ś ubsites, respectively.…”

Resonance assignments for stromelysin complexed with a β-sulfonyl hydroxamate inhibitor

“…Among these sets, AN 4173 (Li et al 1998) lists only 1 H and 15 N Fig. 1 Two-dimensional 1 H-15 N HSQC spectrum of *1 mM uniformly 13 C, 15 N labeled stromelysin complexed with a b-sulfonyl hydroxamate inhibitor in 10 mM 2 H 4 -imidazole, pH 6.6, 2.5 mM CaCl 2 , 5 lM ZnCl 2 and 0.02% NaN 3 at 27°C.…”

Effect of Species Differences on Stromelysin-1 (MMP-3) Inhibitor Potency

“…The catalytic domain can be expressed independently and functions as a competent protease with substrate and inhibitor specificities typical of the full-length parent enzyme (11)(12)(13)(14). Several structures of MMP catalytic domains (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32) and one full-length enzyme (33) have been reported and describe a common protein fold for the MMP active-site domain. This consists of an open-face ␣-␤ sandwich made up of three ␣-helices packed against a 5-stranded ␤-sheet.…”