8682
J . Org. Chem. 2001, 66, 8682-8684
reported previously.4 The glucose-6-phosphate (G6P)/
High En a n tioselectivity a n d Br oa d
Su bstr a te Sp ecificity of a Ca r bon yl
Red u cta se: Tow a r d a Ver sa tile Bioca ta lyst
glucose-6-phosphate dehydrogenase (G6PDH) system8
was used to regenerate a catalytic amount of NADPH in
the enzymatic reductions. Under the reaction conditions
employed, the enzyme and NADPH make approximately
30 000 and 100 turnovers at 100% conversion, respec-
tively.4 The results of the asymmetric reductions using
the purified enzyme or the BY whole cells are sum-
marized in Table 1, where the previously reported results
are also shown for comparison.
Tadashi Ema,* Hiroyuki Moriya, Toru Kofukuda,
Tomomasa Ishida, Kentaro Maehara,
Masanori Utaka, and Takashi Sakai*
Department of Applied Chemistry, Faculty of Engineering,
Okayama University, Tsushima, Okayama 700-8530, J apan
ema@cc.okayama-u.ac.jp
Received August 13, 2001
The enzymatic reductions showed higher enantio-
selectivity than the corresponding whole-cell reductions
in most cases, and 13 out of 20 alcohols obtained in the
former had the enantiomeric purity of more than 98%
enantiomeric excess (ee) (Table 1). The selectivity was
even inverted by using the purified enzyme as compared
with the corresponding whole-cell reductions (entries 12-
16 and 19). The isolated yields for some ester-containing
alcohols were higher in the enzymatic reductions than
in the corresponding whole-cell reductions (entries 5-8,
11, 13, 16-17, and 20), partly because hydrolases that
cleave the ester bond of the substrates and/or the reduced
products were not contained in the enzymatic reductions.
The isolated yields for 32 and 35 in the enzymatic
reductions were very low, which is due to the relatively
low enzymatic activity for 12 and 15.
In tr od u ction
The simultaneous achievement of high enantioselec-
tivity and broad substrate specificity is one of the most
important aspects of chiral catalysts. Recent experimen-
tal results1 and mechanistic studies2 indicate that even
an enzyme can exert high enantioselectivity that is
compatible with broad substrate specificity. A carbonyl
reductase that we have purified from bakers’ yeast (BY,3
Saccharomyces cerevisiae) exhibits high enantioselectivity
for a variety of ketones including R-chloro ketone, R-ac-
etoxy ketone, R- and â-keto esters, and â-diketones.4 This
carbonyl reductase is a monomeric, NADPH-dependent
enzyme with a molecular mass of ca. 37 kDa, although
its natural substrate and function are unknown. BY is a
useful biocatalyst,5 partly because such an enzyme,
capable of showing broad substrate specificity, is con-
tained in the cells. Although many reductases have been
isolated from BY6 and other organisms,7 little is known
about the scope and limitation of substrate specificity of
isolated enzymes. In this paper, we report that the
potential capabilities of this enzyme as a versatile chiral
biocatalyst are promising and even surprising.
Optically active alcohols useful for organic synthesis
were obtained by the enzymatic reductions. Some of them
have been used as chiral building blocks for natural
products and biologically active compounds: e.g., (R)-34
for fluoxetine,9 (S)-35 for dihydrokawain,10 (S)-36 for
xestospongin A,11 and (S)-37 for pyrenophorin.12 The
antipodal enantiomer of (S)-32 has been used in the
syntheses of compactin analogues.13 Compounds (R)-22
and (S)-26 are applicable to the syntheses of chiral host
molecules.14 As shown in Table 1, both aliphatic and
aromatic ketones were successfully transformed. Not only
R- and â-keto esters 9-16 but also γ-keto ester 17 were
converted to the corresponding alcohols in high enantio-
meric excesses. Entries 2, 6, and 12 indicate that olefin
is inert for the enzyme. Previously, we have observed that
the enzymatic reductions of R-chloro ketone 1 and R-ac-
etoxy ketone 5 give (R)-21 and (S)-25, respectively. In
this study, R-chloro ketones 2-4 were reduced to the
corresponding (R)-alcohols, while R-acetoxy ketones 6-8
were reduced to the corresponding (S)-alcohols. Thus, this
single reductase can afford the derivatives of both enan-
tiomers of 1,2-diols in a manner that is dependent on the
R-substituent (Cl or OAc).
Resu lts a n d Discu ssion
The purification of the enzyme and asymmetric reduc-
tions were carried out according to the procedures
(1) For example: (a) Wong, C.-H.; Whitesides, G. M. Enzymes in
Synthetic Organic Chemistry; Pergamon: Oxford, 1994. (b) Enzyme
Catalysis in Organic Synthesis; Drauz, K., Waldmann, H., Eds.; VCH:
New York, 1995. (c) Faber, K. Biotransformations in Organic Chem-
istry; Springer-Verlag: Berlin, 1995.
(2) Ema, T.; Kobayashi, J .; Maeno, S.; Sakai, T.; Utaka, M. Bull.
Chem. Soc. J pn. 1998, 71, 443.
(3) Abbreviations used in this paper: BY, bakers’ yeast; G6P,
glucose-6-phosphate; G6PDH, glucose-6-phosphate dehydrogenase;
NADP+, â-nicotinamide adenine dinucleotide phosphate; NADPH,
reduced form of NADP+; SDS-PAGE, sodium dodecyl sulfate poly-
acrylamide gel electrophoresis.
(4) Ema, T.; Sugiyama, Y.; Fukumoto, M.; Moriya, H.; Cui, J .-N.;
Sakai, T.; Utaka, M. J . Org. Chem. 1998, 63, 4996.
(5) For example, see: (a) Servi, S. Synthesis 1990, 1. (b) Csuk, R.;
Gla¨nzer, B. I. Chem. Rev. 1991, 91, 49.
(8) Wong, C.-H.; Whitesides, G. M. J . Am. Chem. Soc. 1981, 103,
4890.
(9) Cheˆnevert, R.; Fortier, G.; Rhlid, R. B. Tetrahedron 1992, 48,
6769.
(10) Spino, C.; Mayes, N.; Desfosse´s, H. Tetrahedron Lett. 1996, 37,
6503.
(11) Baldwin, J . E.; Melman, A.; Lee, V.; Firkin, C. R.; Whitehead,
R. C. J . Am. Chem. Soc. 1998, 120, 8559.
(12) Baldwin, J . E.; Adlington, R. M.; Ramcharitar, S. H. Synlett
1992, 875.
(13) (a) Bennett, F.; Knight, D. W. Tetrahedron Lett. 1988, 29, 4865.
(b) Bennett, F.; Knight, D. W.; Fenton, G. J . Chem. Soc., Perkin Trans.
1 1991, 133.
(14) Bradshaw, J . S.; Huszthy, P.; McDaniel, C. W.; Zhu, C. Y.;
Dalley, N. K.; Izatt, R. M.; Lifson, S. J . Org. Chem. 1990, 55,
3129.
(6) (a) Shieh, W.-R.; Gopalan, A. S.; Sih, C. J . J . Am. Chem. Soc.
1985, 107, 2993. (b) Nakamura, K.; Kawai, Y.; Nakajima, N.; Ohno,
A. J . Org. Chem. 1991, 56, 4778. (c) Shieh, W.-R.; Sih, C. J .
Tetrahedron: Asymmetry 1993, 4, 1259. (d) Ishihara, K.; Nakajima,
N.; Tsuboi, S.; Utaka, M. Bull. Chem. Soc. J pn. 1994, 67, 3314. (e)
Nakamura, K.; Kondo, S.; Kawai, Y.; Nakajima, N.; Ohno, A. Biosci.
Biotechnol. Biochem. 1994, 58, 2236. (f) Nakamura, K.; Kondo, S.;
Nakajima, N.; Ohno, A. Tetrahedron 1995, 51, 687.
(7) (a) Kim, M.-J .; Whitesides, G. M. J . Am. Chem. Soc. 1988, 110,
2959. (b) Casy, G. Tetrahedron Lett. 1992, 33, 8159. (c) St. Clair, N.;
Wang, Y.-F.; Margolin, A. L. Angew. Chem., Int. Ed. 2000, 39, 380.
The cell-free extracts are also used as biocatalysts: (d) Bradshaw, C.
W.; Hummel, W.; Wong, C.-H. J . Org. Chem. 1992, 57, 1532.
10.1021/jo0108257 CCC: $20.00 © 2001 American Chemical Society
Published on Web 11/09/2001