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Type: Artigo de periódico
Title: A novel and enantioselective epoxide hydrolase from Aspergillus brasiliensis CCT 1435: Purification and characterization
Author: Beloti, LL
Costa, BZ
Toledo, MAS
Santos, CA
Crucello, A
Favaro, MTP
Santiago, AS
Mendes, JS
Marsaioli, AJ
Souza, AP
Abstract: A novel epoxide hydrolase from Aspergillus brasiliensis CCT1435 (AbEH) was cloned and overexpressed in Escherichia call cells with a 6xHis-tag and purified by nickel affinity chromatography. Gel filtration analysis and circular dichroism measurements indicated that this novel AbEH is a homodimer in aqueous solution and contains the typical secondary structure of an alpha/beta hydrolase fold. The activity of AbEH was initially assessed using the fluorogenic probe O-(3,4-epoxybutyl) umbelliferone and was active in a broad range of pH (6-9) and temperature (25-45 degrees C); showing optimum performance at pH 6.0 and 30 degrees C. The Michaelis constant (K-M) and maximum rate (V-max) values were 495 mu M and 0.24 mu M/s, respectively. Racemic styrene oxide (SO) was used as a substrate to assess the AbEH activity and enantioselectivity, and 66% of the SO was hydrolyzed after only 5 min of reaction, with the remaining (S)-SO ee exceeding 99% in a typical kinetic resolution behavior. The AbEH-catalyzed hydrolysis of SO was also evaluated in a biphasic system of water:isooctane; (R)-diol in 84% ee and unreacted (S)-SO in 36% ee were produced, with 43% conversion in 24 h, indicating a discrete enantioconvergent behavior for AbEH. This novel epoxide hydrolase has biotechnological potential for the preparation of enantiopure epoxides or vicinal diols. (C) 2013 Elsevier Inc. All right S reserved.
Country: EUA
Editor: Academic Press Inc Elsevier Science
Citation: Protein Expression And Purification. Academic Press Inc Elsevier Science, v. 91, n. 2, n. 175, n. 183, 2013.
Rights: fechado
Identifier DOI: 10.1016/j.pep.2013.08.001
Date Issue: 2013
Appears in Collections:Unicamp - Artigos e Outros Documentos

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