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Title: Engineering the N-terminal end of CelA results in improved performance and growth of Caldicellulosiruptor bescii on crystalline cellulose

Abstract

ABSTRACT CelA is the most abundant enzyme secreted by Caldicellulosiruptor bescii and has been shown to outperform mixtures of commercially available exo‐ and endoglucanases in vitro. CelA contains both a glycoside hydrolase family 9 endoglucanase and a glycoside hydrolase family 48 exoglucanase known to be synergistic in their activity, connected by three cellulose‐binding domains via linker peptides. Here, repeated aspartate residues were introduced into the N ‐terminal ends of CelA GH9 and GH48 domains to improve secretion efficiency and/or catalytic efficiency of CelA. Among several constructs, the highest activity on carboxymethylcellulose (CMC), 0.81 ± 0.03 mg/mL was observed for the C. bescii strain containing CelA with 5‐aspartate tag at the N ‐terminal end of GH9 domain—an 82% increase over wild type CelA. In addition, expression of CelA with N ‐terminal repeated aspartate residues in C. bescii results in a dramatic increase in its ability to grow on Avicel. Biotechnol. Bioeng. 2017;114: 945–950. © 2016 Wiley Periodicals, Inc.

Authors:
 [1];  [2];  [2];  [2];  [1]
+ Show Author Affiliations
  1. Univ. of Georgia, Athens, GA (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); National Renewable Energy Lab. (NREL), Golden, CO (United States)
Publication Date:
Research Org.:
National Renewable Energy Laboratory (NREL), Golden, CO (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER)
OSTI Identifier:
1349022
Alternate Identifier(s):
OSTI ID: 1401885
Report Number(s):
NREL/JA-2700-67747
Journal ID: ISSN 0006-3592
Grant/Contract Number:  
AC36-08GO28308
Resource Type:
Accepted Manuscript
Journal Name:
Biotechnology and Bioengineering
Additional Journal Information:
Journal Volume: 114; Journal Issue: 5; Journal ID: ISSN 0006-3592
Publisher:
Wiley
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; biomass deconstruction; CelA; repeated aspartate residues; Caldicellulosiruptior

Citation Formats

Kim, Sun -Ki, Chung, Daehwan, Himmel, Michael E., Bomble, Yannick J., and Westpheling, Janet. Engineering the N-terminal end of CelA results in improved performance and growth of Caldicellulosiruptor bescii on crystalline cellulose. United States: N. p., 2016. Web. doi:10.1002/bit.26242.
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Kim, Sun -Ki, Chung, Daehwan, Himmel, Michael E., Bomble, Yannick J., & Westpheling, Janet. Engineering the N-terminal end of CelA results in improved performance and growth of Caldicellulosiruptor bescii on crystalline cellulose. United States. https://doi.org/10.1002/bit.26242
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Kim, Sun -Ki, Chung, Daehwan, Himmel, Michael E., Bomble, Yannick J., and Westpheling, Janet. Mon . "Engineering the N-terminal end of CelA results in improved performance and growth of Caldicellulosiruptor bescii on crystalline cellulose". United States. https://doi.org/10.1002/bit.26242. https://www.osti.gov/servlets/purl/1349022.
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@article{osti_1349022,
title = {Engineering the N-terminal end of CelA results in improved performance and growth of Caldicellulosiruptor bescii on crystalline cellulose},
author = {Kim, Sun -Ki and Chung, Daehwan and Himmel, Michael E. and Bomble, Yannick J. and Westpheling, Janet},
abstractNote = {ABSTRACT CelA is the most abundant enzyme secreted by Caldicellulosiruptor bescii and has been shown to outperform mixtures of commercially available exo‐ and endoglucanases in vitro. CelA contains both a glycoside hydrolase family 9 endoglucanase and a glycoside hydrolase family 48 exoglucanase known to be synergistic in their activity, connected by three cellulose‐binding domains via linker peptides. Here, repeated aspartate residues were introduced into the N ‐terminal ends of CelA GH9 and GH48 domains to improve secretion efficiency and/or catalytic efficiency of CelA. Among several constructs, the highest activity on carboxymethylcellulose (CMC), 0.81 ± 0.03 mg/mL was observed for the C. bescii strain containing CelA with 5‐aspartate tag at the N ‐terminal end of GH9 domain—an 82% increase over wild type CelA. In addition, expression of CelA with N ‐terminal repeated aspartate residues in C. bescii results in a dramatic increase in its ability to grow on Avicel. Biotechnol. Bioeng. 2017;114: 945–950. © 2016 Wiley Periodicals, Inc.},
doi = {10.1002/bit.26242},
journal = {Biotechnology and Bioengineering},
number = 5,
volume = 114,
place = {United States},
year = {Mon Dec 26 00:00:00 EST 2016},
month = {Mon Dec 26 00:00:00 EST 2016}
}
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https://doi.org/10.1002/bit.26242
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    Works referencing / citing this record:

    Engineering cellulolytic bacterium Clostridium thermocellum to co-ferment cellulose- and hemicellulose-derived sugars simultaneously
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    • Xiong, Wei; Reyes, Luis H.; Michener, William E.
    • Biotechnology and Bioengineering, Vol. 115, Issue 7
    • DOI: 10.1002/bit.26590

    High-level constitutive expression of leech hyaluronidase with combined strategies in recombinant Pichia pastoris
    journal, January 2020

    Genomic and physiological analyses reveal that extremely thermophilic Caldicellulosiruptor changbaiensis deploys uncommon cellulose attachment mechanisms
    journal, August 2019

    • Khan, Asma M. A. M.; Mendoza, Carl; Hauk, Valerie J.
    • Journal of Industrial Microbiology & Biotechnology, Vol. 46, Issue 9-10
    • DOI: 10.1007/s10295-019-02222-1

    Genetically Engineered Proteins to Improve Biomass Conversion: New Advances and Challenges for Tailoring Biocatalysts
    journal, August 2019

    • Ribeiro, Lucas Ferreira; Amarelle, Vanesa; Alves, Luana de Fátima
    • Molecules, Vol. 24, Issue 16
    • DOI: 10.3390/molecules24162879

    Progress in understanding and overcoming biomass recalcitrance: a BioEnergy Science Center (BESC) perspective
    journal, November 2017

    Genetically Engineered Proteins to Improve Biomass Conversion: New Advances and Challenges for Tailoring Biocatalysts
    journal, August 2019

    • Ribeiro, Lucas Ferreira; Amarelle, Vanesa; Alves, Luana de Fátima
    • Molecules, Vol. 24, Issue 16
    • DOI: 10.3390/molecules24162879
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