| Full text | |
| Author(s): |
Beloti, Lilian L.
[1]
;
Costa, Bruna Z.
[2]
;
Toledo, Marcelo A. S.
[1]
;
Santos, Clelton A.
[1]
;
Crucello, Aline
[1]
;
Favaro, Marianna T. P.
[1]
;
Santiago, Andre S.
[1]
;
Mendes, Juliano S.
[1]
;
Marsaioli, Anita J.
[2]
;
Souza, Anete P.
[1, 3]
Total Authors: 10
|
| Affiliation: | [1] Univ Estadual Campinas, Ctr Biol Mol & Engn Genet, BR-13083875 Campinas, SP - Brazil
[2] Univ Estadual Campinas, Inst Quim, Sao Paulo - Brazil
[3] Univ Estadual Campinas, Dept Biol Vegetal, Inst Biol, Sao Paulo - Brazil
Total Affiliations: 3
|
| Document type: | Journal article |
| Source: | Protein Expression and Purification; v. 91, n. 2, p. 175-183, OCT 2013. |
| Web of Science Citations: | 10 |
| 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. (AU) | |
| FAPESP's process: | 11/10378-5 - Purification and functional characterization of copper mine microbial oxidoreductases enzymes |
| Grantee: | Bruna Zucoloto da Costa |
| Support Opportunities: | Scholarships in Brazil - Doctorate |