In vivo screening of haloalkane dehalogenase mutants

被引：4

作者：

Chang, CH

Schindler, JF

Unkefer, CJ

Vanderberg, LA

Brainard, JR

Terwilliger, TC

机构：

[1] Univ Calif Los Alamos Natl Lab, Struct Biol Grp, Los Alamos, NM 87545 USA

[2] Univ Calif Los Alamos Natl Lab, Chem Sci & Technol Div, Los Alamos, NM 87545 USA

来源：

BIOORGANIC & MEDICINAL CHEMISTRY | 1999年 / 7卷 / 10期

关键词：

D O I：

10.1016/S0968-0896(99)00148-0

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：

Haloalkane dehalogenase (DhlA) from Xanthobacter autotrophicus GJ10 catalyzes the dehalogenation of short chain primary alkyl halides. Due to the high K-m and low turnover, wild type DhlA is not optimal for applications in bioremediation. We have developed an in vivo screen, based on a colorimetric pH indicator, to identify DhlA mutant with improved catalytic activity. after screening 50,000 colonies, we identified a DhlA mutant with a lower pH optimum. Sequence analysis of the mutant revealed a single substitution, alanine 149 to threonine, which is located close to the active site of DhlA. Replacement of alanine 149 via site-directed mutagenesis with threonine, serine or cysteine retained the mutant phenotype. Other substitutions at position 149 show little or no activity. (C) 1999 Elsevier Science Ltd. All rights reserved.

引用

页码：2175 / 2181

页数：7

共 31 条

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