Multiple sequence alignment-inspired mutagenesis of marine epoxide hydrolase of Mugil cephalus for enhancing enantioselective hydrolytic activity

Sung Hee Choi; Hee Sook Kim; Eun Yeol Lee

Journal of Industrial and Engineering Chemistry, Vol.18, No.1, 72-76, January, 2012

DOI10.1016/j.jiec.2011.11.085 Export Citation

Multiple sequence alignment-inspired mutagenesis of marine epoxide hydrolase of Mugil cephalus for enhancing enantioselective hydrolytic activity

Sung Hee Choi, Hee Sook Kim, and Eun Yeol Lee^{1, †}

Department of Food Science and Biotechnology, Kyungsung University, Busan 608-736, Republic of Korea
¹Department of Chemical Engineering, Kyung Hee University, Gyeonggi-do 446-701, Republic of Korea

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The marine fish microsomal epoxide hydrolase (mEH) of Mugil cephalus was engineered to enhance the enantioselective hydrolytic activity by multiple sequence alignment-inspired mutagenesis. The amino acid sequences of Aspergillus niger, Rhodotorula glutinis, zebra fish and human mEH were aligned and analyzed for identifying target amino acids. Single-point mutants (Q170K, E186K, E378D) and doublepoint mutants (E378D-Q170K, E378D-Y348F, E378D-Y348H) were developed and their hydrolytic activities were compared. The double-point mutant, E378D-Q170K, exhibited an enhanced hydrolytic activity by 4.6-fold, compared to the wild-type M. cephalus mEH. Enantiopure (S)-styrene oxide could be readily prepared with high enantiopurity more than 99%ee by using the double-point mutant.

Keywords:Epoxide hydrolase;Multiple sequence alignment;Mutagenesis;Mugil cephalus

[References]

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Hwang S, Choi CY, Lee EY, J. Ind. Eng. Chem., 16(1), 1 (2010)
Sylvestre J, Chautard H, Cedrone F, Delcourt M, Org. Process Res. Dev., 10, 562 (2006)
Reetz MT, Adv. Catal., 49, 1 (2006)
Lee EY, Shuler ML, Biotechnol. Bioeng., 98(2), 318 (2007)
Reetz MT, J. Org. Chem., 74, 5767 (2009)
van Loo B, Spelberg JHL, Kingma J, Sonke T, Wubbolts MG, Janssen DB, Chem.Biol., 11, 981 (2004)
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Lee SJ, Kim HS, Kim SJ, Park S, Kim BJ, Shuler ML, Lee EY, Biotechnol. Lett., 29(2), 237 (2007)
Choi SH, Kim HS, Lee EY, Biotechnol. Lett., 31(10), 1617 (2009)
Davidson AR, in: Guerois R, de la Paz ML (Eds.), Protein Design: Methods and Applications, Humana Press, New Jersey, 171 (2006)
Barth S, Fischer M, Schmid RD, Pleiss J, Bioinformatics., 20, 2845 (2004)
Yang X, Safrany ST, Shears SB, J. Biol. Chem., 274, 35434 (1999)
Arand M, Muller F, Mecky A, Hinz W, Urban P, Pompon D, Kellner R, Oesch F, Biochem. J., 337, 37 (1999)
Rink R, Spelberg JHL, Pieters RJ, Kingma J, Nardini M, Kellogg RM, Dijkstra BW, Janssen DB, J. Am. Chem. Soc., 121(32), 7417 (1999)

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[1] Archelas A, Furstoss R, Curr. Opin. Chem. Biol., 5, 112 (2001)

[2] Hwang S, Choi CY, Lee EY, J. Ind. Eng. Chem., 16(1), 1 (2010)

[3] Sylvestre J, Chautard H, Cedrone F, Delcourt M, Org. Process Res. Dev., 10, 562 (2006)

[4] Reetz MT, Adv. Catal., 49, 1 (2006)

[5] Lee EY, Shuler ML, Biotechnol. Bioeng., 98(2), 318 (2007)

[6] Reetz MT, J. Org. Chem., 74, 5767 (2009)

[7] van Loo B, Spelberg JHL, Kingma J, Sonke T, Wubbolts MG, Janssen DB, Chem.Biol., 11, 981 (2004)

[8] Reetz MT, Torre C, Eipper A, Lohmer RM, Hermes M, Brunner B, Maichele A, Bocola M, Arand M, Cronin A, Genzel Y, Archelas A, Furstoss R, Org. Lett., 6, 177 (2004)

[9] Cedrone F, Niel S, Ait-abdelkader N, Torre C, Krumm H, Roca S, Bhatnagar T, Maichele A, Reetz MT, Baratti JC, Biocatal. Biotransform., 21, 357 (2003)

[10] Lee SJ, Kim HS, Kim SJ, Park S, Kim BJ, Shuler ML, Lee EY, Biotechnol. Lett., 29(2), 237 (2007)

[11] Choi SH, Kim HS, Lee EY, Biotechnol. Lett., 31(10), 1617 (2009)

[12] Davidson AR, in: Guerois R, de la Paz ML (Eds.), Protein Design: Methods and Applications, Humana Press, New Jersey, 171 (2006)

[13] Barth S, Fischer M, Schmid RD, Pleiss J, Bioinformatics., 20, 2845 (2004)

[14] Yang X, Safrany ST, Shears SB, J. Biol. Chem., 274, 35434 (1999)

[15] Arand M, Muller F, Mecky A, Hinz W, Urban P, Pompon D, Kellner R, Oesch F, Biochem. J., 337, 37 (1999)

[16] Rink R, Spelberg JHL, Pieters RJ, Kingma J, Nardini M, Kellogg RM, Dijkstra BW, Janssen DB, J. Am. Chem. Soc., 121(32), 7417 (1999)