KnowVolution of a GH5 cellulase from Penicillium verruculosum to improve thermal stability for biomass degradation
Contreras, F., Thiele, M. J., Pramanik, S., Rozhkova, A. M., Dotsenko, A. S., Zorov, I. N., Sinitsyn, A. P., Davari, M. D. & Schwaneberg, U. ACS Sustainable Chemistry & Engineering. 2020, 8, 12388–12399. DOI: 10.1021/acssuschemeng.0c02465
Molecular knowledge on cellulases engineering for increased thermostability offers a high potential for sustainable biomass degradation.
Cellulases can be applied in lignocellulosic biomass degradation, feedstock, and pulp and paper production. In different applications, cellulases need to be active in high temperatures and harsh conditions. Cellulases are an expensive component in the processes; therefore, an increase in the cellulase lifetime will decrease the production costs and achieve more sustainable production. Nowadays, protein engineering has emerged as an important tool to improve enzyme properties. In this paper, we report a KnowVolution campaign of the cellulase PvCel5A from Penicillium verruculosum towards increased thermostability. The PvCel5A evolution shows that the KnowVolution engineering strategy, which combines experimental and computation work to minimize experimental efforts, was highly successful and generated a molecular understanding of the C-terminus’ role in improving thermal resistance. When compared to the wild type PvCel5A, variant PvCel5A-R17 presented an improvement of 5.5-fold in its half-life at 75 °C (from 32 to 175 min), and an increase in the melting temperature (Tm) by 7.7 °C (from 70.8 to 78.5 °C). Interestingly, the specific activity of PvCel5A was not reduced through the improvement of the melting temperature (high specific activity requires enzyme flexibility; thermal resistance requires strong interactions/rigid structures). Glycoside hydrolases from family 5 (GH 5) shares a (β/α)8-barrel, and the C-terminal region is a common feature within all enzymes in this family. Therefore, the stabilization concept can likely be transferred to other GH5 glycoside hydrolases. We believe the molecular knowledge of how to tailor cellulases for increased thermostability offers a great potential for enzymatic degradation and full exploitation of lignocellulosic biomass.
Computational biology division.
This work was funded from the Bundesministerium für Bildung und Forschung (BMBF) project EnzyBioDeg (FKZ: 031B0506) Bioeconomy international. MD simulations were performed with computing resources granted by JARA-HPC from RWTH Aachen University under the project jara0187.Copyright: © ACS Sustainable Chem. Eng.