PMID- 16597975
OWN - NLM
STAT- MEDLINE
DCOM- 20060628
LR  - 20181224
IS  - 0099-2240 (Print)
IS  - 1098-5336 (Electronic)
IS  - 0099-2240 (Linking)
VI  - 72
IP  - 4
DP  - 2006 Apr
TI  - Pseudomonad cyclopentadecanone monooxygenase displaying an uncommon spectrum of 
      Baeyer-Villiger oxidations of cyclic ketones.
PG  - 2707-20
AB  - Baeyer-Villiger monooxygenases (BVMOs) are biocatalysts that offer the prospect 
      of high chemo-, regio-, and enantioselectivity in the organic synthesis of 
      lactones or esters from a variety of ketones. In this study, we have cloned, 
      sequenced, and overexpressed in Escherichia coli a new BVMO, cyclopentadecanone 
      monooxygenase (CpdB or CPDMO), originally derived from Pseudomonas sp. strain 
      HI-70. The 601-residue primary structure of CpdB revealed only 29% to 50% 
      sequence identity to those of known BVMOs. A new sequence motif, characterized by 
      a cluster of charged residues, was identified in a subset of BVMO sequences that 
      contain an N-terminal extension of approximately 60 to 147 amino acids. The 
      64-kDa CPDMO enzyme was purified to apparent homogeneity, providing a specific 
      activity of 3.94 micromol/min/mg protein and a 20% yield. CPDMO is monomeric and 
      NADPH dependent and contains approximately 1 mol flavin adenine dinucleotide per 
      mole of protein. A deletion mutant suggested the importance of the N-terminal 54 
      amino acids to CPDMO activity. In addition, a Ser261Ala substitution in a 
      Rossmann fold motif resulted in an improved stability and increased affinity of 
      the enzyme towards NADPH compared to the wild-type enzyme (K(m) = 8 microM versus 
      K(m) = 24 microM). Substrate profiling indicated that CPDMO is unusual among 
      known BVMOs in being able to accommodate and oxidize both large and small ring 
      substrates that include C(11) to C(15) ketones, methyl-substituted C(5) and C(6) 
      ketones, and bicyclic ketones, such as decalone and beta-tetralone. CPDMO has the 
      highest affinity (K(m) = 5.8 microM) and the highest catalytic efficiency 
      (k(cat)/K(m) ratio of 7.2 x 10(5) M(-1) s(-1)) toward cyclopentadecanone, hence 
      the Cpd designation. A number of whole-cell biotransformations were carried out, 
      and as a result, CPDMO was found to have an excellent enantioselectivity (E > 
      200) as well as 99% S-selectivity toward 2-methylcyclohexanone for the production 
      of 7-methyl-2-oxepanone, a potentially valuable chiral building block. Although 
      showing a modest selectivity (E = 5.8), macrolactone formation of 
      15-hexadecanolide from the kinetic resolution of 2-methylcyclopentadecanone using 
      CPDMO was also demonstrated.
FAU - Iwaki, Hiroaki
AU  - Iwaki H
AD  - National Research Council Canada, Biotechnology Research Institute, 6100 
      Royalmount Avenue, Montreal, Quebec H4P 2R2, Canada.
FAU - Wang, Shaozhao
AU  - Wang S
FAU - Grosse, Stephan
AU  - Grosse S
FAU - Bergeron, Helene
AU  - Bergeron H
FAU - Nagahashi, Ayako
AU  - Nagahashi A
FAU - Lertvorachon, Jittiwud
AU  - Lertvorachon J
FAU - Yang, Jianzhong
AU  - Yang J
FAU - Konishi, Yasuo
AU  - Konishi Y
FAU - Hasegawa, Yoshie
AU  - Hasegawa Y
FAU - Lau, Peter C K
AU  - Lau PC
LA  - eng
SI  - GENBANK/AB232538
SI  - GENBANK/AB234289
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Appl Environ Microbiol
JT  - Applied and environmental microbiology
JID - 7605801
RN  - 0 (DNA, Bacterial)
RN  - 0 (DNA, Ribosomal)
RN  - 0 (Fatty Acids)
RN  - 0 (Hydrocarbons, Alicyclic)
RN  - 0 (Ketones)
RN  - 0 (RNA, Ribosomal, 16S)
RN  - CCW02D961F (pentadecanoic acid)
RN  - EC 1.- (Mixed Function Oxygenases)
SB  - IM
MH  - Amino Acid Sequence
MH  - Cloning, Molecular
MH  - DNA, Bacterial/analysis
MH  - DNA, Ribosomal/analysis
MH  - Fatty Acids/chemistry/*metabolism
MH  - Hydrocarbons, Alicyclic/metabolism
MH  - Ketones/*metabolism
MH  - *Mixed Function Oxygenases/chemistry/genetics/metabolism
MH  - Molecular Sequence Data
MH  - Oxidation-Reduction
MH  - Pseudomonas/classification/*enzymology/genetics/growth & development
MH  - RNA, Ribosomal, 16S/genetics
MH  - Sequence Analysis, DNA
MH  - Soil Microbiology
MH  - Substrate Specificity
PMC - PMC1449013
EDAT- 2006/04/07 09:00
MHDA- 2006/06/29 09:00
PMCR- 2006/08/01
CRDT- 2006/04/07 09:00
PHST- 2006/04/07 09:00 [pubmed]
PHST- 2006/06/29 09:00 [medline]
PHST- 2006/04/07 09:00 [entrez]
PHST- 2006/08/01 00:00 [pmc-release]
AID - 72/4/2707 [pii]
AID - 2789-05 [pii]
AID - 10.1128/AEM.72.4.2707-2720.2006 [doi]
PST - ppublish
SO  - Appl Environ Microbiol. 2006 Apr;72(4):2707-20. doi: 
      10.1128/AEM.72.4.2707-2720.2006.