Cloning, sequencing and high expression in Escherichia coli of D-hydantoinase gene from Burkholderia pickettii.
- Author:
Zhen XU
1
;
Wei-Hong JIANG
;
Rui-Shen JIAO
;
Yun-Liu YANG
Author Information
1. Shanghai Institute of Plant Physiology and Ecology, Shanghai Institute for Biological Science, Chinese Academy of Sciences, Shanghai 200032, China.
- Publication Type:Journal Article
- MeSH:
Amidohydrolases;
genetics;
metabolism;
Amino Acid Sequence;
Animals;
Burkholderia;
classification;
enzymology;
genetics;
Cloning, Molecular;
DNA Probes;
Escherichia coli;
Gene Expression;
Molecular Sequence Data;
Recombination, Genetic;
Sequence Analysis, DNA;
Sequence Homology, Amino Acid
- From:
Chinese Journal of Biotechnology
2002;18(2):149-154
- CountryChina
- Language:Chinese
-
Abstract:
A strain, MMR003, used for D-p-HPG production in industry was classified as Burkholderia pickettii by morphological observation and biochemical characterization. The gene encoding the D-hydantoinase enzyme was cloned, sequenced and expressed in Escherichia coli. The nucleotide sequence of the 5.0 kb insert of subclone pXZ-total was determined. One open reading frame of 1374 bp was found and predicted to encode a polypeptide consisting of 458 amino acids in size of 50 kD. The amino acid sequence alignment of D-hydantoinase from Burkholderia pickettii shows the 85% homologous with the corresponding enzyme from Agrobacterium radiobacter NRRL B11291. The D-hydantoinase gene (dha) harboured in the plasmid pXZPH2 in E. coli BL21(DE3) was highly expressed by IPTG induction. The D-hydantoinase activity for D, L-p-hydroxyphenylhydantion is 0.66 u/mL broth, which is 2-fold increase compared to the parent strain Burkholderia pickettii.