Cloning and characterization of the enzyme epoxide hydrolase of the Trichoderma reesei.

Author(s)

Gabriel Stephani de Oliveira

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI)
• Defense date: 2018-05-16

Examining board members

Felipe Santiago Chambergo Alcalde; Cristiane Rodrigues Guzzo Carvalho; Fernanda Marques da Cunha; Viviane Abreu Nunes Cerqueira Dantas; Elen Aquino Perpetuo; Roberto Kopke Salinas

• Advisor: Felipe Santiago Chambergo Alcalde

Abstract

Epoxide hydrolases (EHs) are enzymes that catalyze the hydrolysis of epoxides to their corresponding diols, present a potential biotechnological application (separation of enantiomers for the production of drugs), they are involved in the metabolism of polyunsaturated fatty acids and EH inhibitors are being studied for possible use in the treatment of diseases. An epoxide hydrolase enzyme(TrEH) from the filamentous fungus Trichoderma reesei QM9414 was cloned, expressed, purified and functionally and structurally characterized. The activity of TrEH was determined with the substrate styrene oxide (racemic), showing higher activity at temperatures of 23 to 50 °C, at pH 7.2 at 37 °C, and the catalytic constants Km= 4.6 mM and kcat= 336 s-1. The recombinant enzyme has been shown to be enantioselective, because it preferably hydrolyzes (S)-(-)-styrene oxide, (R)-(-)-epichlorohydrin and (S)-(-)-1,2- epoxybutane. TrEH inhibitors have been identified and some of them inhibit up to 60% growth of T. reesei . The tertiary structure of TrEH (1.7 Å) was determined by crystallography, showing α/ β-hydrolase folding and low homology with any other EH structure. TrEH is a new soluble epoxide hydrolase enzyme whose properties show its potential for use in biotechnological applications. (AU)

• FAPESP's process: 15/03329-9 - Cloning and characterization of the enzymes epoxide hydrolases from Trichoderma reesei
• Grantee: Gabriel Stephani de Oliveira
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)