A gene cluster for cyclohexanone oxidation wascloned from Rhodococcus sp. TK6, which is capable of growthon cyclohexanone as the sole carbon source. The 9,185-bpDNA sequence analysis revealed seven potential open readingframes (ORFs), designated as ssd-chnR-chnD-chnC-chnB-chnE-partial pcd. The chnBCDE genes encode enzymes for theby cyclohexanone monooxygenase (ChnB), ε-caprolactonehydrolase (ChnC), 6-hydroxyhexanoate dehydrogenase (ChnD),and 6-oxohexanoate dehydrogenase (ChnE). Furthermore, thepresence of a regulatory element in the downstream region ofthe chnD gene supports the notion that chnR is a putativeregulatory gene. Among them, the activity of ChnB wasconfirmed and characterized, folowing their expression andpurification in Escherichia coli chnBgene (chnB gene with 6 successive codons for His at the 3'terminus).