Mechanism of class 1 (Glycosylhydrolase family 47) α-mannosidases involved in N-glycan processing and endoplasmic reticulum quality control

被引:92
作者
Karaveg, K
Siriwardena, A
Tempel, W
Liu, ZJ
Glushka, J
Wang, BC
Moremen, KW
机构
[1] Univ Georgia, Complex Carbohydrate Res Ctr, Athens, GA 30602 USA
[2] Univ Georgia, Dept Biochem & Mol Biol, Athens, GA 30602 USA
[3] Univ Mississippi, Dept Chem & Biochem, University, MS 38677 USA
关键词
D O I
10.1074/jbc.M500119200
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
Quality control in the endoplasmic reticulum ( ER) determines the fate of newly synthesized glycoproteins toward either correct folding or disposal by ER-associated degradation. Initiation of the disposal process involves selective trimming of N-glycans attached to misfolded glycoproteins by ER alpha-mannosidase I and subsequent recognition by the ER degradation-enhancing alpha-mannosidase-like protein family of lectins, both members of glycosylhydrolase family 47. The unusual inverting hydrolytic mechanism catalyzed by members of this family is investigated here by a combination of kinetic and binding analyses of wild type and mutant forms of human ER alpha-mannosidase I as well as by structural analysis of a co-complex with an uncleaved thiodisaccharide substrate analog. These data reveal the roles of potential catalytic acid and base residues and the identification of a novel S-3(1) sugar conformation for the bound substrate analog. The co-crystal structure described here, in combination with the C-1(4) conformation of a previously identified co-complex with the glycone mimic, 1-deoxymannojirimycin, indicates that glycoside bond cleavage proceeds through a least motion conformational twist of a properly predisposed substrate in the -1 subsite. A novel H-3(4) conformation is proposed as the exploded transition state.
引用
收藏
页码:16197 / 16207
页数:11
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