3up4 | pdb_00003up4
Crystal Structure of OTEMO complex with FAD and NADP (form 3)
Structural highlights
FunctionOTEMO_PSEPU Involved in the degradation of (+)-camphor. Catalyzes the lactonization of 2-oxo-delta(3)-4,5, 5-trimethylcyclopentenylacetyl-CoA (OT-CoA), a key intermediate in the metabolism of camphor. 2-Oxocyclopentyl ethyl acetate is also a good substrate, as is 2-oxocyclohexyl ethyl acetate and methyl-substituted cyclohexanones, but free acid is a poor substrate.[1] [2] [3] Publication Abstract from PubMedA dimeric Baeyer-Villiger monooxygenase (BVMO) catalyzing the lactonization of 2-oxo-Delta(3)-4,5,5-trimethylcyclopentenylacetyl-CoA, a key intermediate in the metabolism of camphor by Pseudomonas putida ATCC 17453 had been initially characterized in 1983 by Trudgill and co-workers (H.J. Ougham, D.G. Taylor, and P.W. Trudgill, J. Bacteriol. 153:140-152, 1983). Here we have cloned and overexpressed the 2-oxo-Delta(3)-4,5,5-trimethylcyclopentenylacetyl-CoA monooxygenase (OTEMO) in Escherichia coli, and determined its three-dimensional structure with bound FAD at 1.95 A resolution as well as with bound FAD and NADP(+) at 2.0 A resolution. OTEMO represents the first homodimeric type 1 BVMO structure bound to FAD/NADP(+). Comparison of several crystal forms of OTEMO bound to FAD and NADP(+) revealed conformational plasticity of several loop regions, some of which have been implicated as contributing to the substrate specificity profile of structurally-related BVMOs. Substrate specificity studies confirmed that the 2-oxo-Delta(3)-4,5,5-trimethylcyclopentenylacetic acid coenzyme A ester is preferred over the free acid. However, the catalytic efficiency (k(cat)/K(m)) favors 2-n-hexyl cyclopentanone (4.3 x 10(5) M(-1)s(-1)) as a substrate, although its affinity (K(m) = 32 muM) was lower than that of the CoA-activated substrate (18 muM). In whole cell biotransformation experiments, OTEMO showed a unique enantiocomplementarity to the action of the prototypical cyclohexanone monooxygenase (CHMO), and appeared to be particularly useful for the oxidation of 4-substituted cyclohexanones. Overall, this work expands our understanding of the molecular structure and mechanistic complexity of the type 1 family of BVMOs as well as expanding the catalytic repertoire of one of its original members. Camphor Pathway 2-Oxo-{Delta}3-4,5,5-trimethylcyclopentenylacetyl-CoA Monooxygenase of Pseudomonas putida ATCC 17453: Cloning, Baeyer-Villiger Biooxidations, and Structures.,Leisch H, Shi R, Grosse S, Morley K, Bergeron H, Cygler M, Iwaki H, Hasegawa Y, Lau PC Appl Environ Microbiol. 2012 Jan 20. PMID:22267661[4] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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