Extraordinary enantiospecificity of lipase catalysis in organic media induced by purification and catalyst engineering

Tsai, Shau‐Wei; Dordick, Jonathan S.

doi:10.1002/(sici)1097-0290(19961020)52:2<296::aid-bit9>3.0.co;2-l

Cited by 36 publications

(23 citation statements)

References 21 publications

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“…Lipases have been widely investigated for various nonaqueous biotransformations (Therisod and Klibanov, 1987;Klibanov, 1990;Tsai and Dordick, 1996;Ducret et al, 1998;Dong et al, 1999;Kiran and Divakar, 2001). …”

Section: Lipases In Bioconversions In Organic Mediamentioning

confidence: 99%

“…The purified lipase preparation was less active than the crude enzyme in dry n-heptane, whereas the presence of a small concentration of water dramatically activated the purified enzyme but not the crude enzyme in the esterification of racemic 2-(4-chlorophenoxy) propanoic acid with n-butanol (Tsai and Dordick, 1996).…”

Section: Lipases In Resolution Of Racemic Acids and Alcoholsmentioning

confidence: 99%

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Production, purification, characterization, and applications of lipases

Sharma

Chisti

Banerjee

2001

Biotechnology Advances

1,275

712

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Lipases (triacylglycerol acylhydrolases, EC 3.1.1.3) catalyze the hydrolysis and the synthesis of esters formed from glycerol and long-chain fatty acids. Lipases occur widely in nature, but only microbial lipases are commercially significant. The many applications of lipases include speciality organic syntheses, hydrolysis of fats and oils, modification of fats, flavor enhancement in food processing, resolution of racemic mixtures, and chemical analyses. This article discusses the production, recovery, and use of microbial lipases. Issues of enzyme kinetics, thermostability, and bioactivity are addressed. Production of recombinant lipases is detailed. Immobilized preparations of lipases are discussed. In view of the increasing understanding of lipases and their many applications in high-value syntheses and as bulk enzymes, these enzymes are having an increasing impact on bioprocessing. D

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Section: Lipases In Bioconversions In Organic Mediamentioning

confidence: 99%

Section: Lipases In Resolution Of Racemic Acids and Alcoholsmentioning

confidence: 99%

Production, purification, characterization, and applications of lipases

Sharma

Chisti

Banerjee

2001

Biotechnology Advances

1,275

712

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“…However, the enzyme enantioselectivity (i.e., V S /V R ) greatly increases in a range from 2.8fold of 2 to 18.3-fold of 6 after the immobilization. Obviously, the enantioselectivity enhancement can be attributed to the multipoint attachment of SNSM-87 to the support, but not to the diffusion limitation inside the support, as the latter will decrease the apparent enantioselectivity (Tsai and Dordick, 1996).…”

Section: Effects Of Covalent Immobilizationmentioning

confidence: 99%

Improvements of enzyme activity and enantioselectivity via combined substrate engineering and covalent immobilization

Wang

Tsai

Chen

2008

Biotech & Bioengineering

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Esterases, lipases, and serine proteases have been applied as versatile biocatalysts for preparing a variety of chiral compounds in industry via the kinetic resolution of their racemates. In order to meet this requirement, three approaches of enzyme engineering, medium engineering, and substrate engineering are exploited to improve the enzyme activity and enantioselectivity. With the hydrolysis of (R,S)-mandelates in biphasic media consisting of isooctane and pH 6 buffer at 55 degrees C as the model system, the strategy of combined substrate engineering and covalent immobilization leads to an increase of enzyme activity and enantioselectivity from V(S)/(E(t)) = 1.62 mmol/h g and V(S)/V(R) = 43.6 of (R,S)-ethyl mandelate (1) for a Klebsiella oxytoca esterase (named as SNSM-87 from the producer) to 16.7 mmol/h g and 867 of (R,S)-2-methoxyethyl mandelate (4) for the enzyme immobilized on Eupergit C 250L. The analysis is then extended to other (R,S)-2-hydroxycarboxylic acid esters, giving improvements of the enzyme performance from V(S)/(E(t)) = 1.56 mmol/h g and V(S)/V(R) = 41.9 of (R,S)-ethyl 3-chloromandelate (9) for the free esterase to 39.4 mmol/h g and 401 of (R,S)-2-methoxyethyl 3-chloromandelate (16) for the immobilized enzyme, V(S)/(E(t)) = 5.46 mmol/h g and V(S)/V(R) = 8.27 of (R,S)-ethyl 4-chloromandelate (10) for free SNSM-87 to 33.5 mmol/h g and 123 of (R,S)-methyl 4-chloromandelate (14) for the immobilized enzyme, as well as V(S)/(E(t)) = 3.0 mmol/h g and V(S)/V(R) = 7.94 of (R,S)-ethyl 3-phenyllactate (11) for the free esterase to 40.7 mmol/h g and 158 of (R,S)-2-methoxyethyl 3-phenyllactate (18) for the immobilized enzyme. The great enantioselectivty enhancement is rationalized from the alteration of ionization constants of imidazolium moiety of catalytic histidine for both enantiomers and conformation distortion of active site after the covalent immobilization, as well as the selection of leaving alcohol moiety via substrate engineering approach.

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“…Obviously, this can be attributed to the multipoint attachment between the enzyme and support, but not to the diffusion limitation in the support by considering the enantioselectivity enhancement. 22 The former might cause minute conformation changes to the active site and slow down the kinetic process of hydrolyzing (S)-ethyl mandelate, and especially (R)ethyl mandelate. Further experiments by coupling with the kinetic analysis for serine-type hydrolases are being carried out in order to elucidate the interesting behavior of enantioselectivity improvement.…”

Section: Effects Of Solventmentioning

confidence: 99%