Deletion of a single glycosyltransferase in Caldicellulosiruptor bescii eliminates protein glycosylation and growth on crystalline cellulose
Abstract
Protein glycosylation pathways have been identified in a variety of bacteria and are best understood in pathogens and commensals in which the glycosylation targets are cell surface proteins, such as S layers, pili, and flagella. In contrast, very little is known about the glycosylation of bacterial enzymes, especially those secreted by cellulolytic bacteria. Caldicellulosiruptor bescii secretes several unique synergistic multifunctional biomass-degrading enzymes, notably cellulase A which is largely responsible for this organism's ability to grow on lignocellulosic biomass without the conventional pretreatment. It was recently discovered that extracellular CelA is heavily glycosylated. In this work, we identified an O-glycosyltransferase in the C. bescii chromosome and targeted it for deletion. The resulting mutant was unable to grow on crystalline cellulose and showed no detectable protein glycosylation. Multifunctional biomass-degrading enzymes in this strain were rapidly degraded. Furthermore with the genetic tools available in C. bescii, this system represents a unique opportunity to study the role of bacterial enzyme glycosylation as well an investigation of the pathway for protein glycosylation in a non-pathogen.
- Authors:
- Publication Date:
- Research Org.:
- National Renewable Energy Lab. (NREL), Golden, CO (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1618735
- Alternate Identifier(s):
- OSTI ID: 1476874
- Report Number(s):
- NREL/JA-2700-72558
Journal ID: ISSN 1754-6834; 259; PII: 1266
- Grant/Contract Number:
- CBI; AC36-08GO28308
- Resource Type:
- Published Article
- Journal Name:
- Biotechnology for Biofuels
- Additional Journal Information:
- Journal Name: Biotechnology for Biofuels Journal Volume: 11 Journal Issue: 1; Journal ID: ISSN 1754-6834
- Publisher:
- Springer Science + Business Media
- Country of Publication:
- Netherlands
- Language:
- English
- Subject:
- 09 BIOMASS FUELS; bacterial enzymes; glycosylation; lignocellulosic biomass
Citation Formats
Russell, Jordan, Kim, Sun-Ki, Duma, Justin, Nothaft, Harald, Himmel, Michael E., Bomble, Yannick J., Szymanski, Christine M., and Westpheling, Janet. Deletion of a single glycosyltransferase in Caldicellulosiruptor bescii eliminates protein glycosylation and growth on crystalline cellulose. Netherlands: N. p., 2018.
Web. doi:10.1186/s13068-018-1266-x.
Russell, Jordan, Kim, Sun-Ki, Duma, Justin, Nothaft, Harald, Himmel, Michael E., Bomble, Yannick J., Szymanski, Christine M., & Westpheling, Janet. Deletion of a single glycosyltransferase in Caldicellulosiruptor bescii eliminates protein glycosylation and growth on crystalline cellulose. Netherlands. https://doi.org/10.1186/s13068-018-1266-x
Russell, Jordan, Kim, Sun-Ki, Duma, Justin, Nothaft, Harald, Himmel, Michael E., Bomble, Yannick J., Szymanski, Christine M., and Westpheling, Janet. Mon .
"Deletion of a single glycosyltransferase in Caldicellulosiruptor bescii eliminates protein glycosylation and growth on crystalline cellulose". Netherlands. https://doi.org/10.1186/s13068-018-1266-x.
@article{osti_1618735,
title = {Deletion of a single glycosyltransferase in Caldicellulosiruptor bescii eliminates protein glycosylation and growth on crystalline cellulose},
author = {Russell, Jordan and Kim, Sun-Ki and Duma, Justin and Nothaft, Harald and Himmel, Michael E. and Bomble, Yannick J. and Szymanski, Christine M. and Westpheling, Janet},
abstractNote = {Protein glycosylation pathways have been identified in a variety of bacteria and are best understood in pathogens and commensals in which the glycosylation targets are cell surface proteins, such as S layers, pili, and flagella. In contrast, very little is known about the glycosylation of bacterial enzymes, especially those secreted by cellulolytic bacteria. Caldicellulosiruptor bescii secretes several unique synergistic multifunctional biomass-degrading enzymes, notably cellulase A which is largely responsible for this organism's ability to grow on lignocellulosic biomass without the conventional pretreatment. It was recently discovered that extracellular CelA is heavily glycosylated. In this work, we identified an O-glycosyltransferase in the C. bescii chromosome and targeted it for deletion. The resulting mutant was unable to grow on crystalline cellulose and showed no detectable protein glycosylation. Multifunctional biomass-degrading enzymes in this strain were rapidly degraded. Furthermore with the genetic tools available in C. bescii, this system represents a unique opportunity to study the role of bacterial enzyme glycosylation as well an investigation of the pathway for protein glycosylation in a non-pathogen.},
doi = {10.1186/s13068-018-1266-x},
journal = {Biotechnology for Biofuels},
number = 1,
volume = 11,
place = {Netherlands},
year = {Mon Sep 24 00:00:00 EDT 2018},
month = {Mon Sep 24 00:00:00 EDT 2018}
}
https://doi.org/10.1186/s13068-018-1266-x
Web of Science
Works referenced in this record:
Bacterial Protein N -Glycosylation: New Perspectives and Applications
journal, January 2013
- Nothaft, Harald; Szymanski, Christine M.
- Journal of Biological Chemistry, Vol. 288, Issue 10
An Evolving Hierarchical Family Classification for Glycosyltransferases
journal, April 2003
- Coutinho, Pedro M.; Deleury, Emeline; Davies, Gideon J.
- Journal of Molecular Biology, Vol. 328, Issue 2
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
journal, May 2011
- Chung, Dae-Hwan; Huddleston, Jennifer R.; Farkas, Joel
- Journal of Industrial Microbiology & Biotechnology, Vol. 38, Issue 11
High activity CAZyme cassette for improving biomass degradation in thermophiles
journal, February 2018
- Brunecky, Roman; Chung, Daehwan; Sarai, Nicholas S.
- Biotechnology for Biofuels, Vol. 11, Issue 1
Identification of Campylobacter jejuni genes involved in commensal colonization of the chick gastrointestinal tract: C. jejuni commensal colonization
journal, March 2004
- Hendrixson, David R.; DiRita, Victor J.
- Molecular Microbiology, Vol. 52, Issue 2
Protein O-mannosylation: What we have learned from baker's yeast
journal, November 2013
- Loibl, Martin; Strahl, Sabine
- Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, Vol. 1833, Issue 11
Deletion of Caldicellulosiruptor besciiCelA reveals its crucial role in the deconstruction of lignocellulosic biomass
journal, October 2014
- Young, Jenna; Chung, Daehwan; Bomble, Yannick J.
- Biotechnology for Biofuels, Vol. 7, Issue 1
The carbohydrate-active enzymes database (CAZy) in 2013
journal, November 2013
- Lombard, Vincent; Golaconda Ramulu, Hemalatha; Drula, Elodie
- Nucleic Acids Research, Vol. 42, Issue D1
Isolation and bioinformatic analysis of a novel transposable element, ISCbe4, from the hyperthermophilic bacterium, Caldicellulosiruptor bescii
journal, October 2013
- Cha, Minseok; Wang, Hao; Chung, Daehwan
- Journal of Industrial Microbiology & Biotechnology, Vol. 40, Issue 12
Direct conversion of plant biomass to ethanol by engineered Caldicellulosiruptor bescii
journal, June 2014
- Chung, D.; Cha, M.; Guss, A. M.
- Proceedings of the National Academy of Sciences, Vol. 111, Issue 24, p. 8931-8936
Biosynthesis of the N-Linked Glycan in Campylobacter jejuni and Addition onto Protein through Block Transfer
journal, March 2006
- Kelly, J.; Jarrell, H.; Millar, L.
- Journal of Bacteriology, Vol. 188, Issue 7
Harnessing glycosylation to improve cellulase activity
journal, June 2012
- Beckham, Gregg T.; Dai, Ziyu; Matthews, James F.
- Current Opinion in Biotechnology, Vol. 23, Issue 3
Caldicellulosiruptor Core and Pangenomes Reveal Determinants for Noncellulosomal Thermophilic Deconstruction of Plant Biomass
journal, May 2012
- Blumer-Schuette, S. E.; Giannone, R. J.; Zurawski, J. V.
- Journal of Bacteriology, Vol. 194, Issue 15
Insights into plant biomass conversion from the genome of the anaerobic thermophilic bacterium Caldicellulosiruptor bescii DSM 6725
journal, January 2011
- Dam, Phuongan; Kataeva, Irina; Yang, Sung-Jae
- Nucleic Acids Research, Vol. 39, Issue 8
Extremely thermophilic microorganisms for biomass conversion status and prospects
journal, June 2008
- Blumer-Schuette, Sara E.; Kataeva, Irina; Westpheling, Janet
- Current Opinion in Biotechnology, Vol. 19, Issue 3, p. 210-217
Revealing Nature's Cellulase Diversity The Digestion Mechanism of Caldicellulosiruptor bescii CelA
journal, December 2013
- Brunecky, R.; Alahuhta, M.; Xu, Q.
- Science, Vol. 342, Issue 6165, p. 1513-1516
Biochemical Characterization of the O-Linked Glycosylation Pathway in Neisseria gonorrhoeae Responsible for Biosynthesis of Protein Glycans Containing N , N ′-Diacetylbacillosamine
journal, June 2011
- Hartley, Meredith D.; Morrison, Michael J.; Aas, Finn Erik
- Biochemistry, Vol. 50, Issue 22
N-Linked Protein Glycosylation Is Required for Full Competence in Campylobacter jejuni 81-176
journal, September 2004
- Larsen, J. C.; Szymanski, C.; Guerry, P.
- Journal of Bacteriology, Vol. 186, Issue 19
Substrate Specificity of Cytoplasmic N -Glycosyltransferase
journal, June 2014
- Naegeli, Andreas; Michaud, Gaëlle; Schubert, Mario
- Journal of Biological Chemistry, Vol. 289, Issue 35
Construction of a Stable Replicating Shuttle Vector for Caldicellulosiruptor Species: Use for Extending Genetic Methodologies to Other Members of This Genus
journal, May 2013
- Chung, Daehwan; Cha, Minseok; Farkas, Joel
- PLoS ONE, Vol. 8, Issue 5, Article No. e62881
Molecular and Biochemical Analyses of CbCel9A/Cel48A, a Highly Secreted Multi-Modular Cellulase by Caldicellulosiruptor bescii during Growth on Crystalline Cellulose
journal, December 2013
- Yi, Zhuolin; Su, Xiaoyun; Revindran, Vanessa
- PLoS ONE, Vol. 8, Issue 12
Homologous Expression of the Caldicellulosiruptor bescii CelA Reveals that the Extracellular Protein Is Glycosylated
journal, March 2015
- Chung, Daehwan; Young, Jenna; Bomble, Yannick J.
- PLOS ONE, Vol. 10, Issue 3
N-Glycosylation of Campylobacter jejuni Surface Proteins Promotes Bacterial Fitness
journal, March 2013
- Alemka, Abofu; Nothaft, Harald; Zheng, Jing
- Infection and Immunity, Vol. 81, Issue 5
A common pathway for O -linked protein-glycosylation and synthesis of capsule in Acinetobacter baumannii
journal, July 2013
- Lees-Miller, Robert G.; Iwashkiw, Jeremy A.; Scott, Nichollas E.
- Molecular Microbiology, Vol. 89, Issue 5
Glycosylation of bacterial cellulases prevents proteolytic cleavage between functional domains
journal, December 1987
- Langsford, M. L.; Gilkes, N. R.; Singh, B.
- FEBS Letters, Vol. 225, Issue 1-2
Efficient Degradation of Lignocellulosic Plant Biomass, without Pretreatment, by the Thermophilic Anaerobe "Anaerocellum thermophilum" DSM 6725
journal, May 2009
- Yang, Sung-Jae; Kataeva, Irina; Hamilton-Brehm, Scott D.
- Applied and Environmental Microbiology, Vol. 75, Issue 14, p. 4762-4769
Use of Label-Free Quantitative Proteomics To Distinguish the Secreted Cellulolytic Systems of Caldicellulosiruptor bescii and Caldicellulosiruptor obsidiansis
journal, April 2011
- Lochner, Adriane; Giannone, Richard J.; Rodriguez, Miguel
- Applied and Environmental Microbiology, Vol. 77, Issue 12
Protein glycosylation in bacteria: sweeter than ever
journal, October 2010
- Nothaft, Harald; Szymanski, Christine M.
- Nature Reviews Microbiology, Vol. 8, Issue 11
Properties and gene structure of a bifunctional cellulolytic enzyme (CelA) from the extreme thermophile 'Anaerocellum thermophilum' with separate glycosyl hydrolase family 9 and 48 catalytic domains
journal, February 1998
- Zverlov, V.; Mahr, S.; Riedel, K.
- Microbiology, Vol. 144, Issue 2, p. 457-465
Campylobacter Protein Glycosylation Affects Host Cell Interactions
journal, April 2002
- Szymanski, C. M.
- Infection and Immunity, Vol. 70, Issue 4
Heterologous complementation of a pyrF deletion in Caldicellulosiruptor hydrothermalisgenerates a new host for the analysis of biomass deconstruction
journal, September 2014
- Groom, Joseph; Chung, Daehwan; Young, Jenna
- Biotechnology for Biofuels, Vol. 7, Issue 1
The Multi Domain Caldicellulosiruptor bescii CelA Cellulase Excels at the Hydrolysis of Crystalline Cellulose
journal, August 2017
- Brunecky, Roman; Donohoe, Bryon S.; Yarbrough, John M.
- Scientific Reports, Vol. 7, Issue 1
GlycoPP: A Webserver for Prediction of N- and O-Glycosites in Prokaryotic Protein Sequences
journal, July 2012
- Chauhan, Jagat S.; Bhat, Adil H.; Raghava, Gajendra P. S.
- PLoS ONE, Vol. 7, Issue 7
Functional Analysis of the Glucan Degradation Locus in Caldicellulosiruptor bescii Reveals Essential Roles of Component Glycoside Hydrolases in Plant Biomass Deconstruction
journal, October 2017
- Conway, Jonathan M.; McKinley, Bennett S.; Seals, Nathaniel L.
- Applied and Environmental Microbiology, Vol. 83, Issue 24
Glycosylated linkers in multimodular lignocellulose-degrading enzymes dynamically bind to cellulose
journal, August 2013
- Payne, Christina M.; Resch, Michael G.; Chen, Liqun
- Proceedings of the National Academy of Sciences, Vol. 110, Issue 36
A Combined Transmembrane Topology and Signal Peptide Prediction Method
journal, May 2004
- Käll, Lukas; Krogh, Anders; Sonnhammer, Erik L. L.
- Journal of Molecular Biology, Vol. 338, Issue 5
Methylation by a Unique α-class N4-Cytosine Methyltransferase Is Required for DNA Transformation of Caldicellulosiruptor bescii DSM6725
journal, August 2012
- Chung, Daehwan; Farkas, Joel; Huddleston, Jennifer R.
- PLoS ONE, Vol. 7, Issue 8
Whole Proteome pI Values Correlate with Subcellular Localizations of Proteins for Organisms within the Three Domains of Life
journal, April 2001
- Schwartz, R.; Ting, C. S.; King, J.
- Genome Research, Vol. 11, Issue 5
Improved growth media and culture techniques for genetic analysis and assessment of biomass utilization by Caldicellulosiruptor bescii
journal, November 2012
- Farkas, Joel; Chung, Daehwan; Cha, Minseok
- Journal of Industrial Microbiology & Biotechnology, Vol. 40, Issue 1, p. 41-49
Metabolic engineering of Caldicellulosiruptor bescii yields increased hydrogen production from lignocellulosic biomass
journal, January 2013
- Cha, Minseok; Chung, Daehwan; Elkins, James G.
- Biotechnology for Biofuels, Vol. 6, Issue 1
Distinct roles of N- and O-glycans in cellulase activity and stability
journal, December 2017
- Amore, Antonella; Knott, Brandon C.; Supekar, Nitin T.
- Proceedings of the National Academy of Sciences, Vol. 114, Issue 52
Glycosylation of the phosphate binding protein, PstS, in Streptomyces coelicolor by a pathway that resembles protein O-mannosylation in eukaryotes
journal, January 2009
- Wehmeier, S.; Varghese, A. S.; Gurcha, S. S.
- Molecular Microbiology, Vol. 71, Issue 2