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Comparison of Family 9 Cellulases from Mesophilic and Thermophilic Bacteria

Mingardon, Florence and Bagert, John D. and Maisonnier, Cyprien and Trudeau, Devin L. and Arnold, Frances H. (2011) Comparison of Family 9 Cellulases from Mesophilic and Thermophilic Bacteria. Applied and Environmental Microbiology, 77 (4). pp. 1436-1442. ISSN 0099-2240. PMCID PMC3067227. https://resolver.caltech.edu/CaltechAUTHORS:20110214-092102779

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Abstract

Cellulases containing a family 9 catalytic domain and a family 3c cellulose binding module (CBM3c) are important components of bacterial cellulolytic systems. We measured the temperature dependence of the activities of three homologs: Clostridium cellulolyticum Cel9G, Thermobifida fusca Cel9A, and C. thermocellum Cel9I. To directly compare their catalytic activities, we constructed six new versions of the enzymes in which the three GH9-CBM3c domains were fused to a dockerin both with and without a T. fusca fibronectin type 3 homology module (Fn3). We studied the activities of these enzymes on crystalline cellulose alone and in complex with a miniscaffoldin containing a cohesin and a CBM3a. The presence of Fn3 had no measurable effect on thermostability or cellulase activity. The GH9-CBM3c domains of Cel9A and Cel9I, however, were more active than the wild type when fused to a dockerin complexed to scaffoldin. The three cellulases in complex have similar activities on crystalline cellulose up to 60°C, but C. thermocellum Cel9I, the most thermostable of the three, remains highly active up to 80°C, where its activity is 1.9 times higher than at 60°C. We also compared the temperature-dependent activities of different versions of Cel9I (wild type or in complex with a miniscaffoldin) and found that the thermostable CBM is necessary for activity on crystalline cellulose at high temperatures. These results illustrate the significant benefits of working with thermostable enzymes at high temperatures, as well as the importance of retaining the stability of all modules involved in cellulose degradation.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1128/AEM.01802-10DOIArticle
http://aem.asm.org/cgi/content/abstract/77/4/1436PublisherArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3067227/PubMed CentralArticle
ORCID:
AuthorORCID
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2011 American Society for Microbiology. Received 28 July 2010. Accepted 4 December 2010. Published ahead of print on 17 December 2010. We thank Pete Heinzelman for providing the plasmid encoding H. jecorina CBHII, Indira Wu for producing and purifying this cellulase, and Wendy Higashide, from the James C. Liao research group (UCLA), for providing C. cellulolyticum genomic DNA. This work was supported by the U.S. National Science Foundation (award MCB-0903817) and a grant from the U.S. Department of the Interior—DARPA (award N10AP20004).
Funders:
Funding AgencyGrant Number
NSFMCB-0903817
Department of the InteriorN10AP20004
Issue or Number:4
PubMed Central ID:PMC3067227
Record Number:CaltechAUTHORS:20110214-092102779
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20110214-092102779
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:22158
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:10 Mar 2011 00:11
Last Modified:03 Oct 2019 02:35

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