DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Clostridium difficile toxin glucosyltransferase domains in complex with a non-hydrolyzable UDP-glucose analogue

Abstract

Clostridium difficile is the leading cause of hospital-acquired diarrhea and pseudomembranous colitis worldwide. The organism produces two homologous toxins, TcdA and TcdB, which enter and disrupt host cell function by glucosylating and thereby inactivating key signalling molecules within the host. As a toxin-mediated disease, there has been a significant interest in identifying small molecule inhibitors of the toxins’ glucosyltransferase activities. This study was initiated as part of an effort to identify the mode of inhibition for a small molecule inhibitor of glucosyltransferase activity called apigenin. In the course of trying to get co-crystals with this inhibitor, we determined five different structures of the TcdA and TcdB glucosyltransferase domains and made use of a non-hydrolyzable UDP-glucose substrate. While we were able to visualize apigenin bound in one of our structures, the site was a crystal packing interface and not likely to explain the mode of inhibition. Nevertheless, the structure allowed us to capture an apo-state (one without the sugar nucleotide substrate) of the TcdB glycosyltransferase domain that had not been previously observed. Additionally, comparison of this structure with structures obtained in the presence of a non-hydrolyzable UDP-glucose analogue have allowed us to document multiple conformations of a C-terminal loop important formore » catalysis. We present our analysis of these five new structures with the hope that it will advance inhibitor design efforts for this important class of biological toxins.« less

Authors:
 [1]; ORCiD logo [2]
  1. Vanderbilt Univ., Nashville, TN (United States)
  2. Vanderbilt Univ., Nashville, TN (United States); Vanderbilt Univ. Medical Center, Nashville, TN (United States); The Veterans Affairs Tennessee Valley Healthcare System, Nashville, TN (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
National Institutes of Health (NIH); US Department of Veterans Affairs (VA)
OSTI Identifier:
1438879
Grant/Contract Number:  
AI095755; T32 GM008320; BX002943
Resource Type:
Accepted Manuscript
Journal Name:
Journal of Structural Biology
Additional Journal Information:
Journal Volume: 198; Journal Issue: 3; Journal ID: ISSN 1047-8477
Publisher:
Elsevier
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; Glucosyltransferase; Bacterial toxin; UDP-glucose; Clostridium difficile; Crystallography

Citation Formats

Alvin, Joseph W., and Lacy, D. Borden. Clostridium difficile toxin glucosyltransferase domains in complex with a non-hydrolyzable UDP-glucose analogue. United States: N. p., 2017. Web. doi:10.1016/j.jsb.2017.04.006.
Alvin, Joseph W., & Lacy, D. Borden. Clostridium difficile toxin glucosyltransferase domains in complex with a non-hydrolyzable UDP-glucose analogue. United States. https://doi.org/10.1016/j.jsb.2017.04.006
Alvin, Joseph W., and Lacy, D. Borden. Wed . "Clostridium difficile toxin glucosyltransferase domains in complex with a non-hydrolyzable UDP-glucose analogue". United States. https://doi.org/10.1016/j.jsb.2017.04.006. https://www.osti.gov/servlets/purl/1438879.
@article{osti_1438879,
title = {Clostridium difficile toxin glucosyltransferase domains in complex with a non-hydrolyzable UDP-glucose analogue},
author = {Alvin, Joseph W. and Lacy, D. Borden},
abstractNote = {Clostridium difficile is the leading cause of hospital-acquired diarrhea and pseudomembranous colitis worldwide. The organism produces two homologous toxins, TcdA and TcdB, which enter and disrupt host cell function by glucosylating and thereby inactivating key signalling molecules within the host. As a toxin-mediated disease, there has been a significant interest in identifying small molecule inhibitors of the toxins’ glucosyltransferase activities. This study was initiated as part of an effort to identify the mode of inhibition for a small molecule inhibitor of glucosyltransferase activity called apigenin. In the course of trying to get co-crystals with this inhibitor, we determined five different structures of the TcdA and TcdB glucosyltransferase domains and made use of a non-hydrolyzable UDP-glucose substrate. While we were able to visualize apigenin bound in one of our structures, the site was a crystal packing interface and not likely to explain the mode of inhibition. Nevertheless, the structure allowed us to capture an apo-state (one without the sugar nucleotide substrate) of the TcdB glycosyltransferase domain that had not been previously observed. Additionally, comparison of this structure with structures obtained in the presence of a non-hydrolyzable UDP-glucose analogue have allowed us to document multiple conformations of a C-terminal loop important for catalysis. We present our analysis of these five new structures with the hope that it will advance inhibitor design efforts for this important class of biological toxins.},
doi = {10.1016/j.jsb.2017.04.006},
journal = {Journal of Structural Biology},
number = 3,
volume = 198,
place = {United States},
year = {Wed Apr 19 00:00:00 EDT 2017},
month = {Wed Apr 19 00:00:00 EDT 2017}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record

Citation Metrics:
Cited by: 10 works
Citation information provided by
Web of Science

Save / Share:

Works referenced in this record:

PHENIX: a comprehensive Python-based system for macromolecular structure solution
journal, January 2010

  • Adams, Paul D.; Afonine, Pavel V.; Bunkóczi, Gábor
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 213-221
  • DOI: 10.1107/S0907444909052925

Low pH-induced Formation of Ion Channels by Clostridium difficile Toxin B in Target Cells
journal, January 2001

  • Barth, Holger; Pfeifer, Gunther; Hofmann, Fred
  • Journal of Biological Chemistry, Vol. 276, Issue 14
  • DOI: 10.1074/jbc.M009445200

Structures and mechanisms of glycosyltransferases
journal, July 2005

  • Breton, Christelle; Šnajdrová, Lenka; Jeanneau, Charlotte
  • Glycobiology, Vol. 16, Issue 2
  • DOI: 10.1093/glycob/cwj016

A Common Motif of Eukaryotic Glycosyltransferases Is Essential for the Enzyme Activity of Large Clostridial Cytotoxins
journal, July 1998

  • Busch, Christian; Hofmann, Fred; Selzer, Jörg
  • Journal of Biological Chemistry, Vol. 273, Issue 31
  • DOI: 10.1074/jbc.273.31.19566

Involvement of a Conserved Tryptophan Residue in the UDP-Glucose Binding of Large Clostridial Cytotoxin Glycosyltransferases
journal, May 2000

  • Busch, Christian; Hofmann, Fred; Gerhard, Ralf
  • Journal of Biological Chemistry, Vol. 275, Issue 18
  • DOI: 10.1074/jbc.275.18.13228

Toxins A and B from Clostridium difficile differ with respect to enzymatic potencies, cellular substrate specificities, and surface binding to cultured cells.
journal, October 1997

  • Chaves-Olarte, E.; Weidmann, M.; Eichel-Streiber, C.
  • Journal of Clinical Investigation, Vol. 100, Issue 7
  • DOI: 10.1172/JCI119698

Crystal structure of Clostridium difficile toxin A
journal, January 2016

  • Chumbler, Nicole M.; Rutherford, Stacey A.; Zhang, Zhifen
  • Nature Microbiology, Vol. 1, Issue 1
  • DOI: 10.1038/nmicrobiol.2015.2

Clostridium difficile Toxins A and B Are Cation-dependent UDP-glucose Hydrolases with Differing Catalytic Activities
journal, June 1998


Features and development of Coot
journal, March 2010

  • Emsley, P.; Lohkamp, B.; Scott, W. G.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4
  • DOI: 10.1107/S0907444910007493

The Donor Subsite of Trehalose-6-phosphate Synthase: BINARY COMPLEXES WITH UDP-GLUCOSE AND UDP-2-DEOXY-2-FLUORO-GLUCOSE AT 2 Å RESOLUTION
journal, October 2003

  • Gibson, Robert P.; Tarling, Chris A.; Roberts, Shirley
  • Journal of Biological Chemistry, Vol. 279, Issue 3
  • DOI: 10.1074/jbc.M307643200

Crystal structure of UDP-glucose:anthocyanidin 3- O -glucosyltransferase from Clitoria ternatea
journal, September 2013

  • Hiromoto, Takeshi; Honjo, Eijiro; Tamada, Taro
  • Journal of Synchrotron Radiation, Vol. 20, Issue 6
  • DOI: 10.1107/S0909049513020712

Sequencing and analysis of the gene encoding the α-toxin of Clostridium novyi proves its homology to toxins A and B of Clostridium difficile
journal, November 1995

  • Hofmann, Fred; Herrmann, Andrea; Habermann, Ernst
  • Molecular and General Genetics MGG, Vol. 247, Issue 6
  • DOI: 10.1007/BF00290398

Structure and mode of action of clostridial glucosylating toxins: the ABCD model
journal, May 2008


Clostridium difficile Glucosyltransferase Toxin B-essential Amino Acids for Substrate Binding
journal, September 2007

  • Jank, Thomas; Giesemann, Torsten; Aktories, Klaus
  • Journal of Biological Chemistry, Vol. 282, Issue 48
  • DOI: 10.1074/jbc.M703138200

Glucosylation of Rho proteins by Clostridium difficile toxin B
journal, June 1995

  • Just, I.; Selzer, J.; Wilm, M.
  • Nature, Vol. 375, Issue 6531
  • DOI: 10.1038/375500a0

XDS
journal, January 2010

  • Kabsch, Wolfgang
  • Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2
  • DOI: 10.1107/S0907444909047337

[20] Processing of X-ray diffraction data collected in oscillation mode
book, January 1997


Structural Determinants of Clostridium difficile Toxin A Glucosyltransferase Activity
journal, January 2012

  • Pruitt, Rory N.; Chumbler, Nicole M.; Rutherford, Stacey A.
  • Journal of Biological Chemistry, Vol. 287, Issue 11
  • DOI: 10.1074/jbc.M111.298414

Autocatalytic cleavage of Clostridium difficile toxin B
journal, March 2007

  • Reineke, Jessica; Tenzer, Stefan; Rupnik, Maja
  • Nature, Vol. 446, Issue 7134
  • DOI: 10.1038/nature05622

Structural Basis for the Function of Clostridium difficile Toxin B
journal, September 2005

  • Reinert, Dirk J.; Jank, Thomas; Aktories, Klaus
  • Journal of Molecular Biology, Vol. 351, Issue 5
  • DOI: 10.1016/j.jmb.2005.06.071

Mutational Analysis of the Enzymatic Domain of Clostridium difficile Toxin B Reveals Novel Inhibitors of the Wild-Type Toxin
journal, June 2003


Small Molecule Inhibitors of Clostridium difficile Toxin B-Induced Cellular Damage
journal, February 2015


Application of Mutated Clostridium difficile Toxin A for Determination of Glucosyltransferase-Dependent Effects
journal, September 2006

  • Teichert, M.; Tatge, H.; Schoentaube, J.
  • Infection and Immunity, Vol. 74, Issue 10
  • DOI: 10.1128/IAI.00545-06

Chimeric Glycosyltransferases for the Generation of Hybrid Glycopeptides
journal, June 2009


Conformational Changes and Reaction of Clostridial Glycosylating Toxins
journal, April 2008

  • Ziegler, Mathias O. P.; Jank, Thomas; Aktories, Klaus
  • Journal of Molecular Biology, Vol. 377, Issue 5
  • DOI: 10.1016/j.jmb.2007.12.065

Works referencing / citing this record:

The role of toxins in Clostridium difficile infection
journal, October 2017

  • Chandrasekaran, Ramyavardhanee; Lacy, D. Borden
  • FEMS Microbiology Reviews, Vol. 41, Issue 6
  • DOI: 10.1093/femsre/fux048