Epoxide hydrolase activity of Chryseomonas luteola for the asymmetric hydrolysis of aliphatic mono-substituted epoxides A.L. Botes 1 , J.A. Steenkamp 2 , M.Z. Letloenyane 2 and M.S. van Dyk 1 * 1 Department of Microbiology and Biochemistry, University of the Orange Free State, P.O. Box 339, Bloemfontein, South Africa; E-mail: smitms@micro.nw.uovs.ac.za 2 Department of Chemistry, University of the Orange Free State, P.O. Box 339, Bloemfontein, South Africa. Asymmetric hydrolysis of a homologous range of straight chain 1,2-epoxyalkanes was achieved using whole cells of Chryseomonas luteola. Depending on the chain length, hydrolyses of the racemic epoxides afforded optically active epoxides and diols with varying degrees of optical purity. In the case of 1,2-epoxyoctane, the enantiomeric excess of the remaining (S)-epoxide and formed (R)-diol was excellent (ee s  98% and ee p = 86%). This is the first report of a bacterial epoxide hydrolase with such unusual enantioselectivity for terminal mono-substituted epoxides bearing no directing group on the chiral C-2 carbon. Benzyl glycidyl ether and the 2,2-disubstituted epoxide, 2-methyl- 1,2-epoxyheptane, were hydrolysed, but no enantioselectivity was observed. Introduction Our recent finding of novel yeast epoxide hydrolases which are highly enantioselective for straight chain 1,2- epoxyalkanes (Weijers et al., 1997; Botes et al., 1998), prompted us to examine an unclassified, yellow pigmented bacterium from our collection, for similar enantioselectiv- ity. No suitable biocatalysts for the biocatalytic resolution of straight-chain epoxides were previously found amongst bacteria. Enantiomeric excess (e.e,) reported for residual (S)-1,2-epoxyoctane varied between 26–57%, and for the formed (R)-diol between 3–54%. The best values reported were for Nocardia, (Ospiran et al., 1997) which can utilize alkenes via an alkene monooxygenase. All bacteria for which epoxide hydrolase activity had been reported, i.e. Rhodococcus, Mycobacterium paraffinicum and Nocardia (Hecht- berger et al., 1993; Mischitz et al., 1995; Faber et al., 1996; Ospiran et al., 1997), produce red or yellow carotinoids as secondary metabolites. Since the bacterium in our collec- tion, which was subsequently identified as a strain of Chryseomonas luteola, was also yellow pigmented, we were optimistic that epoxide hydrolase activity might be pres- ent. We had previously noted the possible relationship between pigment production by microorganisms and epoxide hydrolase activity (Botes et al., 1998). A 2,2-disubstituted epoxide, 2-methyl-1,2-epoxyheptane, for which excellent enantioselectivity was obtained using Rho- dococcus and Nocardia (Ospiran et al., 1997) was included, for comparison. Benzyl glycidyl ether, a potentially useful compound in the synthesis of chiral amino alcohols (Kamal et al., 1992) and bioactive compounds such as -blockers (Kloosterman et al., 1988), were also subjected to hydrol- ysis. Materials and methods General Reactions were monitored and optical purities were ana- lyzed by GLC on fused silica cyclodextrin capillary col- umns (-DEX 120 and -DEX 225, 30 m  0.25 mm, 0.25m film) using N 2 as carrier gas. Absolute configura- tions of the formed aliphatic 1,2-diols were established by GLC analysis after deracemisation of the racemic 1,2-diols to the S-enantiomer by Candida parapsilosis CBS 0604 (Hasegawa et al., 1990). Absolute configurations of the epoxides were deduced from the established elution order on the -DEX 120 and -DEX 225 columns (Weijers et al., 1997). Concentrations of the epoxides and diols were derived from calibration curves. Preparative chroma- tography was performed on silica gel (Merck 60; 40–63 m). 1 H-NMR spectra were recorded in CDCl 3 or acetone- d 6 on a Bruker MSL 300 (300MHz) spectrometer. Chem- ical shifts () are reported in p.p.m. relative to TMS. Epoxides 1 to 8 ()1,2-Epoxypentane (1), ()1,2-Epoxyhexane (2), ()1,2-epoxyoctane (4), and ()1,2-epoxydodecane (6) were commercially available from Fluka. ()1,2- Biotechnology Letters, Vol 20, No 4, April 1998, pp. 427–430 © 1998 Chapman & Hall Biotechnology Letters ⋅ Vol 20 ⋅ No 4 ⋅ 1998 427