Site-directed mutagenesis of cysteine to threonine in Proteus vulgaris urease active site increases enzyme activity and stability

被引：4

作者：

Jose, J

Lauter, S

Stein, MA

机构：

[1] Univ Saarland, D-66041 Saarbrucken, Germany

[2] Lion Biosci AG, D-69123 Heidelberg, Germany

来源：

BIOTECHNOLOGY LETTERS | 2001年 / 23卷 / 15期

关键词：

active site; Proteus vulgaris; site-directed mutagenesis; Staphylococcus xylosus; urease;

D O I：

10.1023/A:1010541513179

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Cysteine-319 belongs to the flexible flap at the active site of Proteus vulgaris urease. Replacing this cysteine by threonine resulted in a 20-fold increase of specific activity. Temperature stability increased, susceptibility to inhibition by dipyridyl disulfide decreased, and pH optimum shifted from 8 to 6.9. K-m (35 to 12 mM) and V-max (47.4 to 1.8 mu mol min(-1)) were substancially altered. Both variants of the enzyme were irreversibly inhibited by phenylmethanesulfonyl fluoride.

引用

页码：1263 / 1267

页数：5

共 17 条

[1] A new proposal for urease mechanism based on the crystal structures of the native and inhibited enzyme from Bacillus pasteurii:: why urea hydrolysis casts two nickels [J].