A structural view of the action of Escherichia coli (lacZ) β-galactosidase

被引:189
作者
Juers, DH
Heightman, TD
Vasella, A
McCarter, JD
Mackenzie, L
Withers, SG
Matthews, BW [1 ]
机构
[1] Univ Oregon, Howard Hughes Med Inst, Inst Mol Biol, Eugene, OR 97403 USA
[2] Univ Oregon, Dept Phys, Eugene, OR 97403 USA
[3] ETH Zentrum, Organ Chem Lab, CH-8092 Zurich, Switzerland
[4] Univ British Columbia, Dept Chem, Vancouver, BC V6T 1Z1, Canada
关键词
D O I
10.1021/bi011727i
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
The structures of a series of complexes designed to mimic intermediates along the reaction coordinate for beta -galactosidase are presented. These complexes clarify and enhance previous proposals regarding the catalytic mechanism. The nucleophile, Glu537, is seen to covalently bind to the galactosyl moiety. Of the two potential acids, Mg2+ and Glu461, the latter is in better position to directly assist in leaving group departure, suggesting that the metal ion acts in a secondary role. A sodium ion plays, a part in substrate binding by directly ligating the galactosyl 6-hydroxyl. The proposed reaction coordinate involves the movement of the galactosyl moiety deep into the active site pocket. For those ligands that do bind dee ly there is an associated conformational change in which residues within loop 794-804 move up to 10 Angstrom closer to the site of binding. In some cases this can be inhibited by the binding of additional Ligands. The resulting restricted access to the intermediate helps to explain why allolactose, the natural inducer for the lac operon, is the preferred product of transglycosylation.
引用
收藏
页码:14781 / 14794
页数:14
相关论文
共 78 条
[1]  
[Anonymous], 1972, The Enzymes
[2]  
Beckwith J. R., 1970, LACTOSE OPERON
[3]   The crystal structures of Sinapis alba myrosinase and a covalent glycosyl-enzyme intermediate provide insights into the substrate recognition and active-site machinery of an S-glycosidase [J].
Burmeister, WP ;
Cottaz, S ;
Driguez, H ;
Iori, R ;
Palmieri, S ;
Henrissat, B .
STRUCTURE, 1997, 5 (05) :663-675
[4]   *PURIFICATION ET PROPRIETES DE LA BETA-GALACTOSIDASE (LACTASE) DESCHERICHIA-COLI [J].
COHN, M ;
MONOD, J .
BIOCHIMICA ET BIOPHYSICA ACTA, 1951, 7 (01) :153-174
[5]  
CUPPLES CG, 1990, J BIOL CHEM, V265, P5512
[6]  
DAVIES G, 1998, COMPREHENSIVE BIOL C, P136
[7]  
DESCHAVANNE PJ, 1978, J BIOL CHEM, V253, P833
[8]   Collaborative computational project, number 4: Providing programs for protein crystallography [J].
Dodson, EJ ;
Winn, M ;
Ralph, A .
MACROMOLECULAR CRYSTALLOGRAPHY, PT B, 1997, 277 :620-633
[9]   SITE SPECIFIC MUTANTS OF BETA-GALACTOSIDASE SHOW THAT TYR-503 IS UNIMPORTANT IN MG-2+ BINDING BUT THAT GLU-461 IS VERY IMPORTANT AND MAY BE A LIGAND TO MG-2+ [J].
EDWARDS, RA ;
CUPPLES, CG ;
HUBER, RE .
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 1990, 171 (01) :33-37
[10]   Escherichia coli β-galactosidase recognizes a high-energy conformation of C-lactose, a nonhydrolizable substrate analogue.: NMR and modeling studies of the molecular complex [J].
Espinosa, JF ;
Montero, E ;
Vian, A ;
García, JL ;
Dietrich, H ;
Schmidt, RR ;
Martín-Lomas, M ;
Imberty, A ;
Cañada, FJ ;
Jiménez-Barbero, J .
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 1998, 120 (06) :1309-1318