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Role of cysteine residues in thermal inactivation of fungal Cel6A cellobiohydrolases

Wu, Indira and Heel, Thomas and Arnold, Frances H. (2013) Role of cysteine residues in thermal inactivation of fungal Cel6A cellobiohydrolases. Biochimica et Biophysica Acta - Proteins and Proteomics, 1834 (8). pp. 1539-1544. ISSN 1570-9639. doi:10.1016/j.bbapap.2013.05.003. https://resolver.caltech.edu/CaltechAUTHORS:20130819-153647713

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Abstract

Numerous protein engineering studies have focused on increasing the thermostability of fungal cellulases to improve production of fuels and chemicals from lignocellulosic feedstocks. However, the engineered enzymes still undergo thermal inactivation at temperatures well below the inactivation temperatures of hyperthermophilic cellulases. In this report, we investigated the role of free cysteines in the thermal inactivation of wild-type and engineered fungal family 6 cellobiohydrolases (Cel6A). The mechanism of thermal inactivation of Cel6A is consistent with disulfide bond degradation and thiol–disulfide exchange. Circular dichroism spectroscopy revealed that a thermostable variant lacking free cysteines refolds to a native-like structure and retains activity after heat treatment over the pH range 5–9. Whereas conserved disulfide bonds are essential for retaining activity after heat treatment, free cysteines contribute to irreversible thermal inactivation in engineered thermostable Cel6A as well as Cel6A from Hypocrea jecorina and Humicola insolens.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1016/j.bbapap.2013.05.003 DOIArticle
ORCID:
AuthorORCID
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2013 Elsevier B.V. Received 9 March 2013; Received in revised form 1 May 2013; Accepted 6 May 2013; Available online 12 May 2013. This work is funded by the Division of Molecular and Cellular Biosciences through grant MCB-0903817 from the U.S. National Science Foundation. T. Heel is funded by the FWF Schroedinger Fellowship J3327-B21.
Funders:
Funding AgencyGrant Number
NSFMCB-0903817
FWF Der WissenschaftsfondsJ3327-B21
Subject Keywords:Cellulase; Cel6A; Thermal inactivation; Disulfide bond degradation; Thiol disulfide exchange
Issue or Number:8
DOI:10.1016/j.bbapap.2013.05.003
Record Number:CaltechAUTHORS:20130819-153647713
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20130819-153647713
Official Citation:Indira Wu, Thomas Heel, Frances H. Arnold, Role of cysteine residues in thermal inactivation of fungal Cel6A cellobiohydrolases, Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Volume 1834, Issue 8, August 2013, Pages 1539-1544, ISSN 1570-9639, http://dx.doi.org/10.1016/j.bbapap.2013.05.003.
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:40725
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:19 Aug 2013 22:59
Last Modified:10 Nov 2021 00:07

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