Analysis of beta-ketothiolase and acetoacetyl-CoA reductase genes of a methylotrophic bacterium, Paracoccus denitrificans, and their expression in Escherichia coli.
The beta-ketothiolase gene (phaA) and acetoacetyl-CoA reductase gene (phaB) were isolated from Paracoccus denitrificans. Nucleotide sequence analysis showed that they encoded proteins of 391 amino acids with a molecular mass of 40,744 Da and of 242 amino acids with a molecular mass of 25,614 Da, respectively. The predicted gene products exhibited high amino acid identities with those from other bacteria: 64.4-74.0% for the phaA gene product and 47.6-80.6% for the phaB gene product, respectively. Both genes were co-transcribed in a recombinant Escherichia coli. In addition, promoter activity was detected upstream of the phaA gene. Hence, the two genes are organized as an operon, phaA-phaB, in P. denitrificans. NADH was preferred to NADPH as a cofactor of acetoacetyl-CoA reductase.
Yabutani
T
T
Department of Applied Biological Sciences, Faculty of Agriculture, Nagoya University, Japan.
Maehara
A
A
Ueda
S
S
Yamane
T
T
D000102:Acetyl-CoA C-Acyltransferase / Q000235:genetics*; D000429:Alcohol Oxidoreductases / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D015247:Deoxyribonuclease HindIII / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D015870:Gene Expression / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D010231:Paracoccus denitrificans / Q000201:enzymology / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D017422:Sequence Analysis, DNA
D000429:Alcohol Oxidoreductases; C023744:acetoacetyl-CoA reductase; D000102:Acetyl-CoA C-Acyltransferase; D015247:Deoxyribonuclease HindIII