EFFICIENT CATALYSIS OF DISULFIDE FORMATION DURING PROTEIN-FOLDING WITH A SINGLE ACTIVE-SITE CYSTEINE

被引:71
|
作者
WUNDERLICH, M
OTTO, A
MASKOS, K
MUCKE, M
SECKLER, R
GLOCKSHUBER, R
机构
[1] UNIV REGENSBURG,INST BIOPHYS & PHYS BIOCHEM,D-93040 REGENSBURG,GERMANY
[2] UNIV BAYREUTH,BIOCHEM LAB,D-95440 BAYREUTH,GERMANY
关键词
DSBA PROTEIN; PROTEIN DISULFIDE ISOMERASE (PDI); HIRUDIN; OXIDATIVE PROTEIN FOLDING; DISULFIDE INTERCHANGE REACTIONS;
D O I
10.1006/jmbi.1995.0119
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
Protein disulfide isomerases (PDIs) catalyze disulfide bond formation during protein folding in vivo and are essential for viability in eukaryotic cells. They share the active-site sequence C-X-X-C that forms a catalytic disulfide. The recent finding that the EUG1 protein, a PDI-related yeast protein, with C-X-X-S sequence at its active sites can complement PDI-deficiency raised the general question of whether disulfide-isomerase activity is essential for cell viability or whether PDI variants with single active-site thiol groups can be catalytically active as disulfide isomerases. We investigated the function of the catalytic cysteine residues in DsbA, a PDI-related protein required for disulfide formation in the periplasmic space of Escherichia coli, by replacing C30 and C33 with alanine. While the mutant C30A and the double mutant CC30/33AA are inactive, C33A catalyzes disulfide-interchange reactions and oxidative renaturation of the reduced, unfolded thrombin inhibitor hirudin with close to wild-type efficiency Thus, the single active-site thiol group of C30 is sufficient for disulfide-isomerase activity of the DsbA protein.
引用
收藏
页码:28 / 33
页数:6
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