Role of his-224 in the anomalous pH dependence of human stromelysin-1

被引:29
作者
Holman, CM
Kan, CC
Gehring, MR
Van Wart, HE
机构
[1] Roche Biosci, Inflammatory Dis Unit, Palo Alto, CA 94304 USA
[2] Agouron Pharmaceut Inc, San Diego, CA 92121 USA
关键词
D O I
10.1021/bi9822170
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
A plot of the pH dependence of k(cat)/K-M for human stromelysin-1 (HS) exhibits a narrow range of maximal activity extending from pH 5.75 to 6.25 and a broad shoulder in the pH range of 7.5-8.5. In contrast, the pH profiles that have been reported for other members of the matrix metalloproteinase (MMP) family are bell-shaped and exhibit neutral pH optima. We hypothesized that the anomalous pH dependence of HS reflects the ionization of His-224, a residue located in a flexible loop that contributes to the S-1' binding pocket of the enzyme. HS is the only known MMP that has a histidine in this position. To test this hypothesis, the H224Q mutant of the short form (lacking the C-terminal hemopexin-like domain) of HS (sHS) has been prepared and studied. The pH profile of H224Q sHS is bell-shaped and similar to those reported for other MMPs. Although H224Q and wild-type sHS possess similar activities at pH <6, the k(cat)/K-M of H224Q sHS is more than 5-fold greater than that of the wild-type enzyme at pH >7. These data strongly suggest that the deprotonation of His-224 attenuates the activity of HS, thereby accounting for its low pH optimum and the characteristic shoulder in its pH profile. This attenuation of activity appears to be predominantly a K-M effect, reflecting a decrease in the affinity of the enzyme for the peptide substrate.
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页码:677 / 681
页数:5
相关论文
共 49 条
[1]   Molecular recognition of protein-ligand complexes: Applications to drug design [J].
Babine, RE ;
Bender, SL .
CHEMICAL REVIEWS, 1997, 97 (05) :1359-1472
[2]   STROMELYSIN-1 - 3-DIMENSIONAL STRUCTURE OF THE INHIBITED CATALYTIC DOMAIN AND OF THE C-TRUNCATED PROENZYME [J].
BECKER, JW ;
MARCY, AI ;
ROKOSZ, LL ;
AXEL, MG ;
BURBAUM, JJ ;
FITZGERALD, PMD ;
CAMERON, PM ;
ESSER, CK ;
HAGMANN, WK ;
HERMES, JD ;
SPRINGER, JP .
PROTEIN SCIENCE, 1995, 4 (10) :1966-1976
[3]   MATRIX METALLOPROTEINASES - A REVIEW [J].
BIRKEDALHANSEN, H ;
MOORE, WGI ;
BODDEN, MK ;
WINDSOR, LJ ;
BIRKEDALHANSEN, B ;
DECARLO, A ;
ENGLER, JA .
CRITICAL REVIEWS IN ORAL BIOLOGY & MEDICINE, 1993, 4 (02) :197-250
[4]   THE X-RAY CRYSTAL-STRUCTURE OF THE CATALYTIC DOMAIN OF HUMAN NEUTROPHIL COLLAGENASE INHIBITED BY A SUBSTRATE-ANALOG REVEALS THE ESSENTIALS FOR CATALYSIS AND SPECIFICITY [J].
BODE, W ;
REINEMER, P ;
HUBER, R ;
KLEINE, T ;
SCHNIERER, S ;
TSCHESCHE, H .
EMBO JOURNAL, 1994, 13 (06) :1263-1269
[5]   STRUCTURE OF THE CATALYTIC DOMAIN OF HUMAN FIBROBLAST COLLAGENASE COMPLEXED WITH AN INHIBITOR [J].
BORKAKOTI, N ;
WINKLER, FK ;
WILLIAMS, DH ;
DARCY, A ;
BROADHURST, MJ ;
BROWN, PA ;
JOHNSON, WH ;
MURRAY, EJ .
NATURE STRUCTURAL BIOLOGY, 1994, 1 (02) :106-110
[6]   MATRILYSIN-INHIBITOR COMPLEXES - COMMON THEMES AMONG METALLOPROTEASES [J].
BROWNER, MF ;
SMITH, WW ;
CASTELHANO, AL .
BIOCHEMISTRY, 1995, 34 (20) :6602-6610
[7]   Site-directed mutagenesis of the active site glutamate in human matrilysin: Investigation of its role in catalysis [J].
Cha, J ;
Auld, DS .
BIOCHEMISTRY, 1997, 36 (50) :16019-16024
[8]   Metal and pH dependence of heptapeptide catalysis by human matrilysin [J].
Cha, JH ;
Pedersen, MV ;
Auld, DS .
BIOCHEMISTRY, 1996, 35 (49) :15831-15838
[9]  
Cleland W W, 1977, Adv Enzymol Relat Areas Mol Biol, V45, P273
[10]   BIOCHEMICAL-CHARACTERIZATION OF MATRILYSIN - ACTIVATION CONFORMS TO THE STEPWISE MECHANISMS PROPOSED FOR OTHER MATRIX METALLOPROTEINASES [J].
CRABBE, T ;
WILLENBROCK, F ;
EATON, D ;
HYNDS, P ;
CARNE, AF ;
MURPHY, G ;
DOCHERTY, AJP .
BIOCHEMISTRY, 1992, 31 (36) :8500-8507