Heterologous expression of the cytochrome P450cam hydroxylase operon and the repressor gene of Pseudomonas putida in Escherichia coli.
The cytochrome P450cam hydroxylase operon (camDCAB) of Pseudomonas putida is negatively regulated by a repressor, CamR, which also represses its own gene. The expression in P. putida of both camR and camDCAB is derepressed in the presence of D-camphor. We examined the expression in Escherichia coli of camR and camDCAB by monitoring the enzyme activity of the camD gene product. In the presence or absence of D-camphor in the cell culture, the expression in E. coli of camD was significant and constitutive, suggesting no expression of camR. This lack of expression was confirmed by monitoring the beta-galactosidase activity of camR-lacZ translational fusions. However, S1 nuclease mapping revealed that synthesis of camR mRNA in E. coli was significant and constitutive, as observed in the case of camDCAB mRNA. Thus, it is likely that the expression of camR in E. coli is limited at the translational level.
Aramaki
H
H
Department of Microbiology, Daiichi College of Pharmaceutical Sciences, Fukuoka, Japan.
Fujita
M
M
Sagara
Y
Y
Amemura
A
A
Horiuchi
T
T
D002164:Camphor / Q000494:pharmacology; D019475:Camphor 5-Monooxygenase; D003577:Cytochrome P-450 Enzyme System / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006899:Mixed Function Oxygenases / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D009876:Operon / Q000235:genetics; D010957:Plasmids; D016958:Pseudomonas putida / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000032:analysis; D001616:beta-Galactosidase / Q000032:analysis
D012333:RNA, Messenger; D002164:Camphor; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D019475:Camphor 5-Monooxygenase; D015719:Single-Strand Specific DNA and RNA Endonucleases; D001616:beta-Galactosidase