Sequence-based screening for self-sufficient P450 monooxygenase from a metagenome library.
OBJECTIVE
Cytochrome P450 monooxygenases (CYPs) are useful catalysts for oxidation reactions. Self-sufficient CYPs harbour a reductive domain covalently connected to a P450 domain and are known for their robust catalytic activity with great potential as biocatalysts. In an effort to expand genetic sources of self-sufficient CYPs, we devised a sequence-based screening system to identify them in a soil metagenome.


RESULTS
We constructed a soil metagenome library and performed sequence-based screening for self-sufficient CYP genes. A new CYP gene, syk181, was identified from the metagenome library. Phylogenetic analysis revealed that SYK181 formed a distinct phylogenic line with 46% amino-acid-sequence identity to CYP102A1 which has been extensively studied as a fatty acid hydroxylase. The heterologously expressed SYK181 showed significant hydroxylase activity towards naphthalene and phenanthrene as well as towards fatty acids.


CONCLUSIONS
Sequence-based screening of metagenome libraries is expected to be a useful approach for searching self-sufficient CYP genes. The translated product of syk181 shows self-sufficient hydroxylase activity towards fatty acids and aromatic compounds.


CONCLUSIONS
SYK181 is the first self-sufficient CYP obtained directly from a metagenome library. The genetic and biochemical information on SYK181 are expected to be helpful for engineering self-sufficient CYPs with broader catalytic activities towards various substrates, which would be useful for bioconversion of natural products and biodegradation of organic chemicals.
Kim
B S
BS
College of Life and Environmental Sciences, Korea University, seongbuk-Gu, Seoul, Korea. biskim@korea.ac.kr
Kim
S Y
SY
Park
J
J
Park
W
W
Hwang
K Y
KY
Yoon
Y J
YJ
Oh
W K
WK
Kim
B Y
BY
Ahn
J S
JS
D003577:Cytochrome P-450 Enzyme System / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D030541:Databases, Genetic; D005227:Fatty Acids / Q000378:metabolism; D015870:Gene Expression; D015698:Genomic Library; D010802:Phylogeny; D012988:Soil Microbiology
D005227:Fatty Acids; D003577:Cytochrome P-450 Enzyme System