F a ir ly Ma r k ed En a n tioselectivity for th e Hyd r olysis of Am in o Acid
Ester s by Ch em ica lly Mod ified En zym es1
Yoshihiro Yano, Kenji Shimada, J iro Okai, Koichi Goto, Yoko Matsumoto, and
Ryuichi Ueoka*
Division of Applied Chemistry, Graduate School of Sojo University (Former Name: Kumamoto Institute of
Technology), 4-22-1 Ikeda, Kumamoto 860-0082, J apan
ueoka@life.sojo-u.ac.jp
Received October 1, 2002
The hydrolysis (deacylation) of enantiomeric substrates by the chemically modified enzymes
decanoyl-R-chymotrypsin and decanoyl-trypsin was studied. Reaction activity for decanoyl-R-
chymotrypsin was lower than that for the native enzyme, although intriguingly the enantioselectivity
was markedly enhanced as compared with the native enzyme. In particular, the apparently complete
enantioselective catalysis was attained for the hydrolytic cleavage of p-nitrophenyl N-dodecanoyl-
D(L)-phenylalaninates. The enhancement of enantioselectivity, however, was not observed for
decanoyl-trypsin. These results suggest that the chemically modified R-chymotrypsin by addition
of hydrophobic groups has promoted enantioselectivity for the hydrolysis of hydrophobic esters.
In tr od u ction
strength4,10-13 and by changing the composition of the
coaggregates.2,5,8-10,14 In particular, almost complete L-
enantioselective catalysis4,10-12,15 can be attributed to
optimization in the conformation of the enzyme model
in the coaggregate systems.2,10,16 On the other hand, Okai
and co-workers have developed a chemical modification
method to introduce a modifying group to an amino
moiety in R-chymotrypsin.17
In this study, we investigated the enantioselective
hydrolysis of enantiomeric substrates (Z-D(L)-Phe-PNA,
Z-D(L)-Phe-PNP, C12-D(L)-Phe-PNP, and Z-D(L)-Lys-PNP‚
HCl) catalyzed by native enzymes (R-chymotrypsin (Csin);
trypsin (Tsin)) and the chemically modified enzymes
(decanoyl-R-chymotrypsin (Dec-Csin); decanoyltrypsin
Enzymes are attractive catalysts because of their
exquisite chemo-, regio-, and stereospecificity and their
impressive catalytic efficiencies. For example, R-chymo-
trypsin catalyzes the hydrolytic cleavage of peptide bonds
at the carboxyl side of either Phe, Tyr, or Trp residues
in proteins. Developing artificial enzymes with enzyme-
like specificity is often carried out using biomimetic
chemistry. Enzyme model studies have been the subject
of continued interest in such areas as the development
of stereoselective reaction sites for the hydrolysis of
enantiomeric substrates and in aiding the understanding
of the origins of stereoselectivity in the proteolytic
enzyme. In particular, micelles are often used as an
enzyme model, because the structure and properties
provide a remarkably close analogy to those of globular
proteins, including enzymes. The analogy between mi-
celles and proteins may be extended for certain reactions
since micelles exhibit a catalytic activity that has several
characteristics of enzyme catalysis.
From this viewpoint, stereoselective cleavages of N-
protected amino acid and peptide p-nitrophenyl esters
in various surfactant aggregate systems have been used
as models to probe the origins of proteolytic enzymes. In
the course of the study on the stereoselective hydrolysis
of amino acid esters in the coaggregate systems, Ueoka
and others emphasized that the stereochemical control
is attained by regulating temperature2-10 and ionic
(3) Ueoka, R.; Matsumoto, Y. J . Org. Chem. 1984, 49, 3774.
(4) Ueoka, R.; Matsumoto, Y.; Yoshino, T.; Hirose, T.; Kikuchi, J .;
Murakami, Y. Chem. Lett. 1986, 127.
(5) Ueoka, R.; Moss, R. A.; Swarup, S.; Matsumoto, Y.; Strauss, G.;
Murakami, Y. J . Am. Chem. Soc. 1985, 107, 2185.
(6) Matsumoto, Y.; Ueoka, R. J . Org. Chem. 1990, 55, 5797.
(7) Ueoka, R.; Yamada, E.; Yamashita, O.; Matsumoto, Y.; Kato, Y.
Tetrahedron Lett. 1991, 32, 6597.
(8) Ueoka, R.; Mori, S.; Moss, R. A. Langmuir 1994, 10, 2892.
(9) Goto, K.; Matsumoto, Y.; Ueoka, R. J . Org. Chem. 1995, 60, 3342.
(10) Tanoue, O.; Baba, M.; Tokunaga, Y.; Goto, K.; Matsumoto, Y.;
Ueoka, R. Tetrahedron Lett. 1999, 40, 2129.
(11) Ueoka, R.; Matsumoto, Y.; Yoshino, T.; Watanabe, N.; Omura,
K.; Murakami, Y. Chem. Lett. 1986, 1743.
(12) Ueoka, R.; Cho, M.; Matsumoto, Y.; Goto, K.; Kato, Y.; Harada,
K.; Sugii, A. Tetrahedron Lett. 1990, 36, 5335.
(13) Goto, K.; Imamura, C.; Yamamoto, S.; Matsumoto, Y.; Ueoka,
R. Chem. Lett. 1994, 2081.
* To whom correspondence should be addressed. Phone: +81-96-
326-3111. Fax: +81-96-326-3000.
(1) Preliminary communication: Ueoka, R.; Okai, J .; Shimada, K.;
Segawa, D.; Nakata, T.; Okai, H. Chem. Lett. 1994, 2261.
(2) Ueoka, R.; Matsumoto, Y.; Moss, R. A.; Swarup, S.; Sugii, A.;
Harada, K.; Kikuchi, J .; Murakami, Y. J . Am. Chem. Soc. 1988, 110,
1588.
(14) Ueoka, R.; Dozono, H.; Matsumoto, Y.; Moss, R. A.; Cho, M.;
Kitahara, K.; Kato, Y. Chem. Pharm. Bull. 1990, 38, 219.
(15) Ihara, Y.; Igata, K.; Okubo, Y.; Nango, M. J . Chem. Soc., Chem.
Commun. 1989, 1900.
(16) Lehn, J . M. Angew. Chem., Int. Ed. Engl. 1988, 27, 89.
(17) Kawasaki, Y.; Murakami, M.; Dosako, S.; Azuse, I.; Nakamura,
T.; Okai, H. Biosci. Biotech. Biochem. 1992, 56, 441.
10.1021/jo0265075 CCC: $25.00 © 2003 American Chemical Society
Published on Web 01/18/2003
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J . Org. Chem. 2003, 68, 1314-1318