Production of Recombinant Trichoderma reesei Cellobiohydrolase II in a New Expression System Based on Wickerhamomyces anomalus.

Dennis J Díaz-Rincón,Claudia M Parra-Giraldo,Carlos J Alméciga-Díaz,Alexander Rodríguez-López,Angela Espejo-Mojica,Raúl A Poutou-Piñales,Ivonne Duque,Balkys Quevedo-Hidalgo,Erika Osorio

doi:10.1155/2017/6980565

Dennis J Díaz-Rincón, Claudia M Parra-Giraldo + Show 7 more

Open Access

https://doi.org/10.1155/2017/6980565

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Journal: Enzyme research	Publication Date: Aug 30, 2017
Citations: 5	License type: CC BY 4.0

Affiliation: Pontificia Universidad Javeriana

Abstract

Cellulase is a family of at least three groups of enzymes that participate in the sequential hydrolysis of cellulose. Recombinant expression of cellulases might allow reducing their production times and increasing the low proteins concentrations obtained with filamentous fungi. In this study, we describe the production of Trichoderma reesei cellobiohydrolase II (CBHII) in a native strain of Wickerhamomyces anomalus. Recombinant CBHII was expressed in W. anomalus 54-A reaching enzyme activity values of up to 14.5 U L−1. The enzyme extract showed optimum pH and temperature of 5.0–6.0 and 40°C, respectively. Enzyme kinetic parameters (KM of 2.73 mM and Vmax of 23.1 µM min−1) were between the ranges of values reported for other CBHII enzymes. Finally, the results showed that an enzymatic extract of W. anomalus 54-A carrying the recombinant T. reesei CBHII allows production of reducing sugars similar to that of a crude extract from cellulolytic fungi. These results show the first report on the use of W. anomalus as a host to produce recombinant proteins. In addition, recombinant T. reesei CBHII enzyme could potentially be used in the degradation of lignocellulosic residues to produce bioethanol, based on its pH and temperature activity profile.

Highlights

Cellulases are enzymes from the glycoside hydrolase family (EC 3.2.1.-) that are expressed by a broad spectrum of bacteria and fungi strains [1, 2]
We describe the production of recombinant cellobiohydrolase II (CBHII, EC 3.2.1.91) from T. reesei using the native strain W. anomalus 54-A
The results showed the potential of W. anomalus 54A as host to produce recombinant proteins and showed that recombinant T. reesei CBHII has similar characteristics to those reported for the wild-type enzyme

Summary

Introduction

Cellulases are enzymes from the glycoside hydrolase family (EC 3.2.1.-) that are expressed by a broad spectrum of bacteria and fungi strains [1, 2]. Extracellular CBHII activity was only detected in W. anomalus 54-A Celo-3.2 clone, reaching a final enzyme activity of 5.7 U L−1 after 96 h of induction (Figure 1(a)).

Results

Conclusion