Enzymatic Resolution of Racemic Phenyloxirane by a Novel Epoxide Hydrolase from Aspergillus Niger SQ-6 and Its Fed-batch Fermentation

Overview

Journal J Ind Microbiol Biotechnol

Publisher Oxford University Press

Specialty Biotechnology

Date 2005 Dec 2

PMID 16320035

Authors

Yanbin Liu

Qian Sha

Sheng Wu

Jianjun Wang

Liu Yang

Wanru Sun

Affiliations

Soon will be listed here.

Abstract

A microorganism with the ability to catalyze the resolution of racemic phenyloxirane was isolated and identified as Aspergillus niger SQ-6. Chiral capillary electrophoresis was successfully applied to separate both phenyloxirane and phenylethanediol. The epoxide hydrolase (EH) involved in this resolution process was (R)-stereospecific and constitutively expressed. When whole cells were used during the biotransformation process, the optimum temperature and pH for stereospecific vicinal diol production were 35 degrees C and 7.0, respectively. After a 24-h conversion, the enantiomer excess of (R)-phenylethanediol produced was found to be >99%, with a conversion rate of 56%. In fed-batch fermentations at 30 degrees C for 44 h, glycerol (20 g L(-1)) and corn steep liquor (CSL) (30 g L(-1)) were chosen as the best initial carbon and nitrogen sources, and EH production was markedly improved by pulsed feeding of sucrose (2 g L(-1) h(-1)) and continuous feeding of CSL (1 g L(-1) h(-1)) at a fermentation time of 28 h. After optimization, the maximum dry cell weight achieved was 24.5+/-0.8 g L(-1); maximum EH production was 351.2+/-13.1 U L(-1) with a specific activity of 14.3+/-0.5 U g(-1). Partially purified EH exhibited a temperature optimum at 37 degrees C and pH optimum at 7.5 in 0.1 M phosphate buffer. This study presents the first evidence for the existence of a predicted epoxide racemase, which might be important in the synthesis of epoxide intermediates.

References

Cagnon J, PORTO A, Marsaioli A, Manfio G, Eguchi S . First evaluation of the Brazilian microorganisms biocatalytic potential. Chemosphere. 1999; 38(10):2237-42. DOI: 10.1016/s0045-6535(98)00442-1. View

Orru R, Faber K . Stereoselectivities of microbial epoxide hydrolases. Curr Opin Chem Biol. 1999; 3(1):16-21. DOI: 10.1016/s1367-5931(99)80004-0. View

Jin H, Li Z . Enantioselective hydrolysis of o-nitrostyrene oxide by whole cells of Aspergillus niger CGMCC 0496. Biosci Biotechnol Biochem. 2002; 66(5):1123-5. DOI: 10.1271/bbb.66.1123. View

Kotik M, Brichac J, Kyslik P . Novel microbial epoxide hydrolases for biohydrolysis of glycidyl derivatives. J Biotechnol. 2005; 120(4):364-75. DOI: 10.1016/j.jbiotec.2005.06.011. View

Archelas A, Furstoss R . Epoxide hydrolases: new tools for the synthesis of fine organic chemicals. Trends Biotechnol. 1998; 16(3):108-16. DOI: 10.1016/S0167-7799(97)01161-X. View

Fretland A, Omiecinski C . Epoxide hydrolases: biochemistry and molecular biology. Chem Biol Interact. 2001; 129(1-2):41-59. DOI: 10.1016/s0009-2797(00)00197-6. View

Nellaiah H, Morisseau C, Archelas A, Furstoss R, Baratti J . Enantioselective hydrolysis of p-nitrostyrene oxide by an epoxide hydrolase preparation from Aspergillus niger. Biotechnol Bioeng. 1996; 49(1):70-7. DOI: 10.1002/(SICI)1097-0290(19960105)49:1<70::AID-BIT9>3.0.CO;2-Q. View

Genzel Y, Archelas A, Broxterman Q, Schulze B, Furstoss R . Microbiological transformations. 47. A step toward a green chemistry preparation of enantiopure (S)-2-, -3-, and -4-pyridyloxirane via an epoxide hydrolase catalyzed kinetic resolution. J Org Chem. 2001; 66(2):538-43. DOI: 10.1021/jo001406x. View

Bhargava S, Wenger K, Marten M . Pulsed addition of limiting-carbon during Aspergillus oryzae fermentation leads to improved productivity of a recombinant enzyme. Biotechnol Bioeng. 2003; 82(1):111-7. DOI: 10.1002/bit.10548. View

10.

Meijer J, Depierre J . Cytosolic epoxide hydrolase. Chem Biol Interact. 1988; 64(3):207-49. DOI: 10.1016/0009-2797(88)90100-7. View

11.

Archelas A, Furstoss R . Synthesis of enantiopure epoxides through biocatalytic approaches. Annu Rev Microbiol. 1997; 51:491-525. DOI: 10.1146/annurev.micro.51.1.491. View

12.

Choi W, Lee E, Yoon S, Yang S, Choi C . Biocatalytic production of chiral epichlorohydrin in organic solvents. J Biosci Bioeng. 2005; 88(3):339-41. DOI: 10.1016/s1389-1723(00)80022-5. View

13.

Bhatnagar T, Manoj K, Baratti J . A spectrophotometric method to assay epoxide hydrolase activity. J Biochem Biophys Methods. 2001; 50(1):1-13. DOI: 10.1016/s0165-022x(01)00162-2. View

14.

Bhargava S, Nandakumar M, Roy A, Wenger K, Marten M . Pulsed feeding during fed-batch fungal fermentation leads to reduced viscosity without detrimentally affecting protein expression. Biotechnol Bioeng. 2002; 81(3):341-7. DOI: 10.1002/bit.10481. View

15.

Wang J, Zheng G, Yang L, Sun W . Enantioselective separation of epoxides by capillary electrophoresis employing sulfated beta-cyclodextrin as chiral selector. Analyst. 2001; 126(4):438-40. DOI: 10.1039/b100205h. View

16.

Morisseau C, Archelas A, Guitton C, FAUCHER D, Furstoss R, Baratti J . Purification and characterization of a highly enantioselective epoxide hydrolase from Aspergillus niger. Eur J Biochem. 1999; 263(2):386-95. DOI: 10.1046/j.1432-1327.1999.00519.x. View

17.

Arand M, Hemmer H, Durk H, Baratti J, Archelas A, Furstoss R . Cloning and molecular characterization of a soluble epoxide hydrolase from Aspergillus niger that is related to mammalian microsomal epoxide hydrolase. Biochem J. 1999; 344 Pt 1:273-80. PMC: 1220641. DOI: 10.1042/0264-6021:3440273. View

18.

Seidegard J, Depierre J . Microsomal epoxide hydrolase. Properties, regulation and function. Biochim Biophys Acta. 1983; 695(3-4):251-70. DOI: 10.1016/0304-419x(83)90014-8. View

19.

Olsvik E, Kristiansen B . Rheology of filamentous fermentations. Biotechnol Adv. 1994; 12(1):1-39. DOI: 10.1016/0734-9750(94)90288-7. View

20.

Orru R, Archelas A, Furstoss R, Faber K . Epoxide hydrolases and their synthetic applications. Adv Biochem Eng Biotechnol. 1999; 63:145-67. DOI: 10.1007/3-540-69791-8_7. View