Cloning and random mutagenesis of the Erwinia herbicola tyrR gene for high-level expression of tyrosine phenol-lyase.

ABSTRACT: Tyrosine phenol-lyase (Tpl), which can synthesize 3, 4-dihydroxyphenylalanine from pyruvate, ammonia, and catechol, is a tyrosine-inducible enzyme. Previous studies demonstrated that the tpl promoter of Erwinia herbicola is activated by the TyrR protein of Escherichia coli. In an attempt to create a high-Tpl-expressing strain, we cloned the tyrR gene of E. herbicola and then randomly mutagenized it. Mutant TyrR proteins with enhanced ability to activate tpl were screened for by use of the lac reporter system in E. coli. The most increased transcription of tpl was observed for the strain with the mutant tyrR allele involving amino acid substitutions of alanine, cysteine, and glycine for valine-67, tyrosine-72, and glutamate-201, respectively. A tyrR-deficient derivative of E. herbicola was

SUBMITTER: Katayama T 

PROVIDER: S-EPMC92377 | biostudies-literature | 2000 Nov

REPOSITORIES: biostudies-literature