[Cloning, sequencing and high expression in Escherichia coli of D-hydantoinase gene from Burkholderia pickettii].
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.
Xu
Zhen
Z
Shanghai Institute of Plant Physiology and Ecology, Shanghai Institute for Biological Science, Chinese Academy of Sciences, Shanghai 200032, China.
Jiang
Wei-Hong
WH
Jiao
Rui-Shen
RS
Yang
Yun-Liu
YL
D000581:Amidohydrolases / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D019117:Burkholderia / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D015342:DNA Probes; D004926:Escherichia coli; D015870:Gene Expression; D008969:Molecular Sequence Data; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D015342:DNA Probes; D000581:Amidohydrolases; C020047:dihydropyrimidinase