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

Kim, Sun -Ki; Chung, Daehwan; Himmel, Michael E.; Bomble, Yannick J.; Westpheling, Janet

doi:10.1002/bit.26242

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

Journal Article · Mon Dec 26 00:00:00 EST 2016 · Biotechnology and Bioengineering

DOI:https://doi.org/10.1002/bit.26242· OSTI ID:1349022

Kim, Sun -Ki ^[1]; Chung, Daehwan ^[2]; Himmel, Michael E. ^[2]; Bomble, Yannick J. ^[2]; Westpheling, Janet ^[1]

Univ. of Georgia, Athens, GA (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); National Renewable Energy Lab. (NREL), Golden, CO (United States)

Here, 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.

View Accepted Manuscript (DOE)

Research Organization:: NREL (National Renewable Energy Laboratory (NREL), Golden, CO (United States))

Sponsoring Organization:: USDOE; USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)

Grant/Contract Number:: AC36-08GO28308

OSTI ID:: 1349022

Alternate ID(s):: OSTI ID: 1401885

Report Number(s):: NREL/JA--2700-67747

Journal Information:: Biotechnology and Bioengineering, Journal Name: Biotechnology and Bioengineering Journal Issue: 5 Vol. 114; ISSN 0006-3592

Publisher:: WileyCopyright Statement

Country of Publication:: United States

Language:: English

References (26)

Simple amino acid tags improve both expression and secretion of Candida antarctica lipase B in recombinant Escherichia coli : Simple Tags for CalB Secretion In Kim, Sun-Ki; Park, Yong-Cheol; Lee, Hyung Ho Biotechnology and Bioengineering, Vol. 112, Issue 2 https://doi.org/10.1002/bit.25361	journal	November 2014
Export incompatibility of N-terminal basic residues in a mature polypeptide of Eschericbia coli can be alleviated by optimising the signal peptide Maclntyre, Sheila; Eschbach, Marie-Luise; Mutschler, Bettina Molecular and General Genetics MGG, Vol. 221, Issue 3 https://doi.org/10.1007/BF00259413	journal	May 1990
Identification and characterization of CbeI, a novel thermostable restriction enzyme from Caldicellulosiruptor bescii DSM 6725 and a member of a new subfamily of HaeIII-like enzymes Chung, Dae-Hwan; Huddleston, Jennifer R.; Farkas, Joel Journal of Industrial Microbiology & Biotechnology, Vol. 38, Issue 11 https://doi.org/10.1007/s10295-011-0976-x	journal	May 2011
Improved growth media and culture techniques for genetic analysis and assessment of biomass utilization by Caldicellulosiruptor bescii Farkas, Joel; Chung, Daehwan; Cha, Minseok Journal of Industrial Microbiology & Biotechnology, Vol. 40, Issue 1, p. 41-49 https://doi.org/10.1007/s10295-012-1202-1	journal	November 2012
Can delignification decrease cellulose digestibility in acid pretreated corn stover? Ishizawa, Claudia I.; Jeoh, Tina; Adney, William S. Cellulose, Vol. 16, Issue 4 https://doi.org/10.1007/s10570-009-9313-1	journal	June 2009
The Sec protein-translocation pathway Mori, Hiroyuki; Ito, Koreaki Trends in Microbiology, Vol. 9, Issue 10 https://doi.org/10.1016/S0966-842X(01)02174-6	journal	October 2001
Recombinant protein secretion in Escherichia coli Mergulhão, F. J. M.; Summers, D. K.; Monteiro, G. A. Biotechnology Advances, Vol. 23, Issue 3 https://doi.org/10.1016/j.biotechadv.2004.11.003	journal	May 2005
Protein secretion pathways in Bacillus subtilis: Implication for optimization of heterologous protein secretion No authors listed Biotechnology Advances, Vol. 25, Issue 1 https://doi.org/10.1016/j.biotechadv.2006.08.002	journal	January 2007
Application of repeated aspartate tags to improving extracellular production of Escherichia coli l-asparaginase isozyme II Kim, Sun-Ki; Min, Won-Ki; Park, Yong-Cheol Enzyme and Microbial Technology, Vol. 79-80 https://doi.org/10.1016/j.enzmictec.2015.06.017	journal	November 2015
Degradation of microcrystalline cellulose and non-pretreated plant biomass by a cell-free extracellular cellulase/hemicellulase system from the extreme thermophilic bacterium Caldicellulosiruptor bescii Kanafusa-Shinkai, Sumiyo; Wakayama, Jun'ichi; Tsukamoto, Kazumi Journal of Bioscience and Bioengineering, Vol. 115, Issue 1 https://doi.org/10.1016/j.jbiosc.2012.07.019	journal	January 2013
Enhanced production of 2′-fucosyllactose in engineered Escherichia coli BL21star(DE3) by modulation of lactose metabolism and fucosyltransferase Chin, Young-Wook; Kim, Ji-Yeong; Lee, Won-Heong Journal of Biotechnology, Vol. 210 https://doi.org/10.1016/j.jbiotec.2015.06.431	journal	September 2015
Cellulase Assays Adney, William S.; Ehrman, Christine I.; Baker, John O. ACS Symposium Series https://doi.org/10.1021/bk-1994-0566.ch010	book	October 1994
The twin-arginine translocation (Tat) protein export pathway Palmer, Tracy; Berks, Ben C. Nature Reviews Microbiology, Vol. 10, Issue 7 https://doi.org/10.1038/nrmicro2814	journal	June 2012
Alteration of the amino terminus of the mature sequence of a periplasmic protein can severely affect protein export in Escherichia coli. Li, P.; Beckwith, J.; Inouye, H. Proceedings of the National Academy of Sciences, Vol. 85, Issue 20 https://doi.org/10.1073/pnas.85.20.7685	journal	October 1988
Revealing Nature's Cellulase Diversity The Digestion Mechanism of Caldicellulosiruptor bescii CelA Brunecky, R.; Alahuhta, M.; Xu, Q. Science, Vol. 342, Issue 6165, p. 1513-1516 https://doi.org/10.1126/science.1244273	journal	December 2013
Efficient Degradation of Lignocellulosic Plant Biomass, without Pretreatment, by the Thermophilic Anaerobe "Anaerocellum thermophilum" DSM 6725 Yang, Sung-Jae; Kataeva, Irina; Hamilton-Brehm, Scott D. Applied and Environmental Microbiology, Vol. 75, Issue 14, p. 4762-4769 https://doi.org/10.1128/AEM.00236-09	journal	May 2009
Use of Label-Free Quantitative Proteomics To Distinguish the Secreted Cellulolytic Systems of Caldicellulosiruptor bescii and Caldicellulosiruptor obsidiansis Lochner, Adriane; Giannone, Richard J.; Rodriguez, Miguel Applied and Environmental Microbiology, Vol. 77, Issue 12 https://doi.org/10.1128/AEM.02811-10	journal	April 2011
Genome Sequence of the Anaerobic, Thermophilic, and Cellulolytic Bacterium "Anaerocellum thermophilum" DSM 6725 Kataeva, I. A.; Yang, S. -J.; Dam, P. Journal of Bacteriology, Vol. 191, Issue 11 https://doi.org/10.1128/JB.00256-09	journal	April 2009
A Nine-Residue Synthetic Propeptide Enhances Secretion Efficiency of Heterologous Proteins inLactococcus lactis Le Loir, Y.; Gruss, A.; Ehrlich, S. D. Journal of Bacteriology, Vol. 180, Issue 7 https://doi.org/10.1128/JB.180.7.1895-1903.1998	journal	January 1998
Implications of cellobiohydrolase glycosylation for use in biomass conversion Jeoh, Tina; Michener, William; Himmel, Michael E. Biotechnology for Biofuels, Vol. 1, Issue 1 https://doi.org/10.1186/1754-6834-1-10	journal	January 2008
Cellulose crystallinity index: measurement techniques and their impact on interpreting cellulase performance Park, Sunkyu; Baker, John O.; Himmel, Michael E. Biotechnology for Biofuels, Vol. 3, Issue 1 https://doi.org/10.1186/1754-6834-3-10	journal	January 2010
Heterologous complementation of a pyrF deletion in Caldicellulosiruptor hydrothermalisgenerates a new host for the analysis of biomass deconstruction Groom, Joseph; Chung, Daehwan; Young, Jenna Biotechnology for Biofuels, Vol. 7, Issue 1 https://doi.org/10.1186/s13068-014-0132-8	journal	September 2014
Deletion of Caldicellulosiruptor besciiCelA reveals its crucial role in the deconstruction of lignocellulosic biomass Young, Jenna; Chung, Daehwan; Bomble, Yannick J. Biotechnology for Biofuels, Vol. 7, Issue 1 https://doi.org/10.1186/s13068-014-0142-6	journal	October 2014
Expression of the Acidothermus cellulolyticus E1 endoglucanase in Caldicellulosiruptor bescii enhances its ability to deconstruct crystalline cellulose Chung, Daehwan; Young, Jenna; Cha, Minseok Biotechnology for Biofuels, Vol. 8, Issue 1 https://doi.org/10.1186/s13068-015-0296-x	journal	August 2015
Heterologous expression of family 10 xylanases from Acidothermus cellulolyticus enhances the exoproteome of Caldicellulosiruptor bescii and growth on xylan substrates Kim, Sun-Ki; Chung, Daehwan; Himmel, Michael E. Biotechnology for Biofuels, Vol. 9, Issue 1 https://doi.org/10.1186/s13068-016-0588-9	journal	August 2016
Molecular and Biochemical Analyses of CbCel9A/Cel48A, a Highly Secreted Multi-Modular Cellulase by Caldicellulosiruptor bescii during Growth on Crystalline Cellulose Yi, Zhuolin; Su, Xiaoyun; Revindran, Vanessa PLoS ONE, Vol. 8, Issue 12 https://doi.org/10.1371/journal.pone.0084172	journal	December 2013

Cited By (6)

Progress in understanding and overcoming biomass recalcitrance: a BioEnergy Science Center (BESC) perspective Gilna, Paul; Lynd, Lee R.; Mohnen, Debra Biotechnology for Biofuels, Vol. 10, Issue 1 https://doi.org/10.1186/s13068-017-0971-1	journal	November 2017
Genetically Engineered Proteins to Improve Biomass Conversion: New Advances and Challenges for Tailoring Biocatalysts Ribeiro, Lucas Ferreira; Amarelle, Vanesa; Alves, Luana de Fátima Molecules, Vol. 24, Issue 16 https://doi.org/10.3390/molecules24162879	journal	August 2019
Engineering cellulolytic bacterium Clostridium thermocellum to co-ferment cellulose- and hemicellulose-derived sugars simultaneously Xiong, Wei; Reyes, Luis H.; Michener, William E. Biotechnology and Bioengineering, Vol. 115, Issue 7 https://doi.org/10.1002/bit.26590	journal	April 2018
High-level constitutive expression of leech hyaluronidase with combined strategies in recombinant Pichia pastoris Huang, Hao; Liang, Qixing; Wang, Yang Applied Microbiology and Biotechnology, Vol. 104, Issue 4 https://doi.org/10.1007/s00253-019-10282-7	journal	January 2020
Genomic and physiological analyses reveal that extremely thermophilic Caldicellulosiruptor changbaiensis deploys uncommon cellulose attachment mechanisms Khan, Asma M. A. M.; Mendoza, Carl; Hauk, Valerie J. Journal of Industrial Microbiology & Biotechnology, Vol. 46, Issue 9-10 https://doi.org/10.1007/s10295-019-02222-1	journal	August 2019
Genome Stability in Engineered Strains of the Extremely Thermophilic Lignocellulose-Degrading Bacterium Caldicellulosiruptor bescii Williams-Rhaesa, Amanda M.; Poole, Farris L.; Dinsmore, Jessica T. Applied and Environmental Microbiology, Vol. 83, Issue 14 https://doi.org/10.1128/aem.00444-17	journal	May 2017

Similar Records

Deletion of Caldicellulosiruptor bescii CelA reveals its crucial role in the deconstruction of lignocellulosic biomass

Program Document · Thu Oct 09 00:00:00 EDT 2014 · Biotechnology for Biofuels · OSTI ID:1163406

Homologous expression of the Caldicellulosiruptor bescii CelA reveals that the extracellular protein is glycosylated

Journal Article · Sun Mar 22 20:00:00 EDT 2015 · PLoS ONE · OSTI ID:1220729

Related Subjects

09 BIOMASS FUELS
Caldicellulosiruptior
CelA
biomass deconstruction
repeated aspartate residues

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

Citation Formats

References (26)

Cited By (6)

Similar Records

Related Subjects