Isoeugenol monooxygenase and its putative regulatory gene are located in the eugenol metabolic gene cluster in Pseudomonas nitroreducens Jin1.
The plant-derived phenylpropanoids eugenol and isoeugenol have been proposed as useful precursors for the production of natural vanillin. Genes involved in the metabolism of eugenol and isoeugenol were clustered in region of about a 30 kb of Pseudomonas nitroreducens Jin1. Two of the 23 ORFs in this region, ORFs 26 (iemR) and 27 (iem), were predicted to be involved in the conversion of isoeugenol to vanillin. The deduced amino acid sequence of isoeugenol monooxygenase (Iem) of strain Jin1 had 81.4% identity to isoeugenol monooxygenase from Pseudomonas putida IE27, which also transforms isoeugenol to vanillin. Iem was expressed in E. coli BL21(DE3) and was found to lead to isoeugenol to vanillin transformation. Deletion and cloning analyses indicated that the gene iemR, located upstream of iem, is required for expression of iem in the presence of isoeugenol, suggesting it to be the iem regulatory gene. Reverse transcription, real-time PCR analyses indicated that the genes involved in the metabolism of eugenol and isoeugenol were differently induced by isoeugenol, eugenol, and vanillin.
Ryu
Ji-Young
JY
Department of Environmental Science and Engineering, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
Seo
Jiyoung
J
Unno
Tatsuya
T
Ahn
Joong-Hoon
JH
Yan
Tao
T
Sadowsky
Michael J
MJ
Hur
Hor-Gil
HG
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001547:Benzaldehydes / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D005054:Eugenol / Q000031:analogs & derivatives* / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics*; D016366:Open Reading Frames / Q000235:genetics; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D001426:Bacterial Proteins; D001547:Benzaldehydes; D005054:Eugenol; C036643:isoeugenol; C100058:vanillin; D006899:Mixed Function Oxygenases