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Engineered thermostable fungal cellulases exhibit efficient synergistic cellulose hydrolysis at elevated temperatures

Trudeau, Devin L. and Lee, Toni M. and Arnold, Frances H. (2014) Engineered thermostable fungal cellulases exhibit efficient synergistic cellulose hydrolysis at elevated temperatures. Biotechnology and Bioengineering, 111 (12). pp. 2390-2397. ISSN 0006-3592. doi:10.1002/bit.25308.

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A major obstacle to using widely available and low-cost lignocellulosic feedstocks to produce renewable fuels and chemicals is the high cost and low efficiency of the enzyme mixtures used to hydrolyze cellulose to fermentable sugars. One possible solution entails engineering current cellulases to function efficiently at elevated temperatures in order to boost reaction rates and exploit several other advantages of a higher temperature process. Here we describe the creation of the most stable reported fungal endoglucanase, a derivative of Hypocrea jecorina (anamorph Trichoderma reesei) Cel5A, by combining stabilizing mutations identified using consensus design, chimera studies, and structure-based computational methods. The engineered endoglucanase has an optimal temperature that is 17 °C higher than wild type H. jecorina Cel5A, and hydrolyzes 1.5 times as much cellulose over 60 h at its optimum temperature compared to the wild type enzyme at its optimal temperature.This enzyme complements previously-engineered highly-active, thermostable variants of the fungal cellobiohydrolases Cel6A and Cel7A in a thermostable cellulase mixture that hydrolyzes cellulose synergistically at an optimum temperature of 70 °C over 60 h.The thermostable mixture produces three times as much total sugar as the best mixture of the wild type enzymes operating at its optimum temperature of 60 °C, clearly demonstrating the advantage of higher-temperature cellulose hydrolysis.

Item Type:Article
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URLURL TypeDescription Information
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2014 Wiley Periodicals, Inc. Accepted manuscript online: 11 Jun 2014; Manuscript Accepted: 2 Jun 2014; Manuscript Revised: 28 May 2014; Manuscript Received: 6 Mar 2014. Article first published online: 5 Aug 2014. The authors thank Indira Wu, Mathew Smith, Alex Nisthal, and two anonymous reviewers for helpful discussions and suggestions. This work was supported by the Institute for Collaborative Biotechnologies through grant [W911NF-09-D-0001] from the U.S. Army Research and The National Central University, Taiwan, though a Cooperative Agreement for Energy Research Collaboration. DLT was supported by a Canadian National Science and Engineering Research Council post-graduate fellowship [PGSD3-404332-2011]. TML was supported by a National Science Foundation graduate fellowship. A patent application has been submitted for the enzyme variants and mixtures described in this manuscript.
Funding AgencyGrant Number
Army Research Office (ARO)W911NF-09-D-0001
National Central University (Taiwan)UNSPECIFIED
Natural Sciences and Engineering Research Council of Canada (NSERC)PGSD3-404332-2011
NSF Graduate Research FellowshipUNSPECIFIED
Subject Keywords:cellulase; endoglucanase; cellobiohydrolase; thermostability; synergy; biofuels
Issue or Number:12
Record Number:CaltechAUTHORS:20140618-110555181
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Official Citation:Trudeau, D. L., Lee, T. M. and Arnold, F. H. (2014), Engineered thermostable fungal cellulases exhibit efficient synergistic cellulose hydrolysis at elevated temperatures. Biotechnol. Bioeng., 111: 2390–2397. doi: 10.1002/bit.25308
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:46335
Deposited By: Tony Diaz
Deposited On:18 Jun 2014 18:48
Last Modified:10 Nov 2021 17:24

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