A small molecule inhibitor prevents gut bacterial genotoxin production

Volpe, Matthew R.; Velilla, José A.; Daniel-Ivad, Martin; Yao, Jenny J.; Stornetta, Alessia; Villalta, Peter W.; Huang, Hsin-Che; Bachovchin, Daniel A.; Balbo, Silvia; Gaudet, Rachelle; Balskus, Emily P.

doi:10.1038/s41589-022-01147-8

A small molecule inhibitor prevents gut bacterial genotoxin production

Journal Article · 17 October 2022 · Nature Chemical Biology

DOI:https://doi.org/10.1038/s41589-022-01147-8· OSTI ID:2423269

; ; ; Yao, Jenny J.; Stornetta, Alessia; ; Huang, Hsin-Che; ; Balbo, Silvia; ;

Abstract

The human gut bacterial genotoxin colibactin is a possible key driver of colorectal cancer (CRC) development. Understanding colibactin’s biological effects remains difficult owing to the instability of the proposed active species and the complexity of the gut microbiota. Here, we report small molecule boronic acid inhibitors of colibactin biosynthesis. Designed to mimic the biosynthetic precursor precolibactin, these compounds potently inhibit the colibactin-activating peptidase ClbP. Using biochemical assays and crystallography, we show that they engage the ClbP binding pocket, forming a covalent bond with the catalytic serine. These inhibitors reproduce the phenotypes observed in aclbPdeletion mutant and block the genotoxic effects of colibactin on eukaryotic cells. The availability of ClbP inhibitors will allow precise, temporal control over colibactin production, enabling further study of its contributions to CRC. Finally, application of our inhibitors to related peptidase-encoding pathways highlights the power of chemical tools to probe natural product biosynthesis.

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: USDOE

DOE Contract Number:: AC02-06CH11357

OSTI ID:: 2423269

Journal Information:: Nature Chemical Biology, Journal Name: Nature Chemical Biology Journal Issue: 2 Vol. 19; ISSN 1552-4450

Publisher:: Nature Publishing Group

Country of Publication:: United States

Language:: English

References (58)

Collaboration gets the most out of software Morin, Andrew; Eisenbraun, Ben; Key, Jason eLife, Vol. 2 https://doi.org/10.7554/eLife.01456	journal	September 2013
Characterization of the Complete Zwittermicin A Biosynthesis Gene Cluster fromBacillus cereus Kevany, Brian M.; Rasko, David A.; Thomas, Michael G. Applied and Environmental Microbiology, Vol. 75, Issue 4 https://doi.org/10.1128/AEM.02518-08	journal	February 2009
Features and development of Coot Emsley, P.; Lohkamp, B.; Scott, W. G. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4 https://doi.org/10.1107/S0907444910007493	journal	March 2010
MZmine 2: Modular framework for processing, visualizing, and analyzing mass spectrometry-based molecular profile data Pluskal, Tomáš; Castillo, Sandra; Villar-Briones, Alejandro BMC Bioinformatics, Vol. 11, Issue 1 https://doi.org/10.1186/1471-2105-11-395	journal	July 2010
Structural basis of colibactin activation by the ClbP peptidase Velilla, José A.; Volpe, Matthew R.; Kenney, Grace E. Nature Chemical Biology, Vol. 19, Issue 2 https://doi.org/10.1038/s41589-022-01142-z	journal	October 2022
A pragmatic approach to the analysis of DNA histograms with a definable G1 peak Watson, James V.; Chambers, Stephen H.; Smith, Paul J. Cytometry, Vol. 8, Issue 1 https://doi.org/10.1002/cyto.990080101	journal	January 1987
Molecular Basis of Gut Microbiome-Associated Colorectal Cancer: A Synthetic Perspective Healy, Alan R.; Herzon, Seth B. Journal of the American Chemical Society, Vol. 139, Issue 42 https://doi.org/10.1021/jacs.7b07807	journal	October 2017
Accessing Bioactive Natural Products from the Human Microbiome Milshteyn, Aleksandr; Colosimo, Dominic A.; Brady, Sean F. Cell Host & Microbe, Vol. 23, Issue 6 https://doi.org/10.1016/j.chom.2018.05.013	journal	June 2018
Isolation of a Metabolite from the pks Island Provides Insights into Colibactin Biosynthesis and Activity Brotherton, Carolyn A.; Wilson, Matthew; Byrd, Gary Organic Letters, Vol. 17, Issue 6 https://doi.org/10.1021/acs.orglett.5b00432	journal	March 2015
Resistance to nonribosomal peptide antibiotics mediated by d-stereospecific peptidases Li, Yong-Xin; Zhong, Zheng; Hou, Peng Nature Chemical Biology, Vol. 14, Issue 4 https://doi.org/10.1038/s41589-018-0009-4	journal	February 2018
Intestinal Inflammation Targets Cancer-Inducing Activity of the Microbiota Arthur, Janelle C.; Perez-Chanona, Ernesto; Mühlbauer, Marcus Science, Vol. 338, Issue 6103 https://doi.org/10.1126/science.1224820	journal	August 2012
Functional Interaction of Monoubiquitinated FANCD2 and BRCA2/FANCD1 in Chromatin Wang, XiaoZhe; Andreassen, Paul R.; D'Andrea, Alan D. Molecular and Cellular Biology, Vol. 24, Issue 13 https://doi.org/10.1128/MCB.24.13.5850-5862.2004	journal	July 2004
High Prevalence of Mucosa-Associated E. coli Producing Cyclomodulin and Genotoxin in Colon Cancer Buc, Emmanuel; Dubois, Damien; Sauvanet, Pierre PLoS ONE, Vol. 8, Issue 2 https://doi.org/10.1371/journal.pone.0056964	journal	February 2013
Reactivity of an Unusual Amidase May Explain Colibactin’s DNA Cross-Linking Activity Jiang, Yindi; Stornetta, Alessia; Villalta, Peter W. Journal of the American Chemical Society, Vol. 141, Issue 29 https://doi.org/10.1021/jacs.9b02453	journal	June 2019
The human gut bacterial genotoxin colibactin alkylates DNA Wilson, Matthew R.; Jiang, Yindi; Villalta, Peter W. Science, Vol. 363, Issue 6428 https://doi.org/10.1126/science.aar7785	journal	February 2019
Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances Wiegand, Irith; Hilpert, Kai; Hancock, Robert E. W. Nature Protocols, Vol. 3, Issue 2 https://doi.org/10.1038/nprot.2007.521	journal	January 2008
The bacterial toxin colibactin triggers prophage induction Silpe, Justin E.; Wong, Joel W. H.; Owen, Siân V. Nature, Vol. 603, Issue 7900 https://doi.org/10.1038/s41586-022-04444-3	journal	February 2022
Patients with familial adenomatous polyposis harbor colonic biofilms containing tumorigenic bacteria Dejea, Christine M.; Fathi, Payam; Craig, John M. Science, Vol. 359, Issue 6375 https://doi.org/10.1126/science.aah3648	journal	February 2018
Host–microbiota maladaptation in colorectal cancer Janney, Alina; Powrie, Fiona; Mann, Elizabeth H. Nature, Vol. 585, Issue 7826 https://doi.org/10.1038/s41586-020-2729-3	journal	September 2020
How good are my data and what is the resolution? Evans, Philip R.; Murshudov, Garib N. Acta Crystallographica Section D Biological Crystallography, Vol. 69, Issue 7 https://doi.org/10.1107/S0907444913000061	journal	June 2013
Self Resistance to the Atypical Cationic Antimicrobial Peptide Edeine of Brevibacillus brevis Vm4 by the N-Acetyltransferase EdeQ Westman, Erin L.; Yan, Marie; Waglechner, Nicholas Chemistry & Biology, Vol. 20, Issue 8 https://doi.org/10.1016/j.chembiol.2013.06.010	journal	August 2013
Development of a covalent inhibitor of gut bacterial bile salt hydrolases Adhikari, Arijit A.; Seegar, Tom C. M.; Ficarro, Scott B. Nature Chemical Biology, Vol. 16, Issue 3 https://doi.org/10.1038/s41589-020-0467-3	journal	February 2020
Locoregional Effects of Microbiota in a Preclinical Model of Colon Carcinogenesis Tomkovich, Sarah; Yang, Ye; Winglee, Kathryn Cancer Research, Vol. 77, Issue 10 https://doi.org/10.1158/0008-5472.CAN-16-3472	journal	April 2017
Small-molecule inhibitors prevent the genotoxic and protumoural effects induced by colibactin-producing bacteria Cougnoux, Antony; Delmas, Julien; Gibold, Lucie Gut, Vol. 65, Issue 2 https://doi.org/10.1136/gutjnl-2014-307241	journal	January 2015
Colibactin DNA-damage signature indicates mutational impact in colorectal cancer Dziubańska-Kusibab, Paulina J.; Berger, Hilmar; Battistini, Federica Nature Medicine, Vol. 26, Issue 7 https://doi.org/10.1038/s41591-020-0908-2	journal	June 2020
Iron Homeostasis Regulates the Genotoxicity of Escherichia coli That Produces Colibactin Tronnet, Sophie; Garcie, Christophe; Rehm, Nadine Infection and Immunity, Vol. 84, Issue 12 https://doi.org/10.1128/IAI.00659-16	journal	September 2016
Copper-Catalyzed Enantioselective Synthesis of β-Boron β-Amino Esters López, Aurora; Clark, Timothy B.; Parra, Alejandro Organic Letters, Vol. 19, Issue 23 https://doi.org/10.1021/acs.orglett.7b02784	journal	November 2017
ClbP Is a Prototype of a Peptidase Subgroup Involved in Biosynthesis of Nonribosomal Peptides Dubois, Damien; Baron, Olivier; Cougnoux, Antony Journal of Biological Chemistry, Vol. 286, Issue 41 https://doi.org/10.1074/jbc.M111.221960	journal	October 2011
Colibactin: understanding an elusive gut bacterial genotoxin Balskus, Emily P. Natural Product Reports, Vol. 32, Issue 11 https://doi.org/10.1039/C5NP00091B	journal	January 2015
The synergistic triad between microcin, colibactin, and salmochelin gene clusters in uropathogenic Escherichia coli Massip, Clémence; Chagneau, Camille V.; Boury, Michèle Microbes and Infection, Vol. 22, Issue 3 https://doi.org/10.1016/j.micinf.2020.01.001	journal	April 2020
The Colibactin Genotoxin Generates DNA Interstrand Cross-Links in Infected Cells Bossuet-Greif, Nadège; Vignard, Julien; Taieb, Frédéric mBio, Vol. 9, Issue 2 https://doi.org/10.1128/mBio.02393-17	journal	March 2018
Comparative Metabolomics and Structural Characterizations Illuminate Colibactin Pathway-Dependent Small Molecules Vizcaino, Maria I.; Engel, Philipp; Trautman, Eric Journal of the American Chemical Society, Vol. 136, Issue 26 https://doi.org/10.1021/ja503450q	journal	June 2014
Slow-Binding Inhibition of γ-Glutamyl Transpeptidase by γ-boroGlu Stein, Ross L.; DeCicco, Carl; Nelson, David Biochemistry, Vol. 40, Issue 19 https://doi.org/10.1021/bi010147i	journal	April 2001
Structure elucidation of colibactin and its DNA cross-links Xue, Mengzhao; Kim, Chung Sub; Healy, Alan R. Science, Vol. 365, Issue 6457 https://doi.org/10.1126/science.aax2685	journal	August 2019
DPP9’s Enzymatic Activity and Not Its Binding to CARD8 Inhibits Inflammasome Activation Griswold, Andrew R.; Ball, Daniel P.; Bhattacharjee, Abir ACS Chemical Biology, Vol. 14, Issue 11 https://doi.org/10.1021/acschembio.9b00462	journal	September 2019
Bacterial genotoxin colibactin promotes colon tumour growth by inducing a senescence-associated secretory phenotype Cougnoux, Antony; Dalmasso, Guillaume; Martinez, Ruben Gut, Vol. 63, Issue 12 https://doi.org/10.1136/gutjnl-2013-305257	journal	March 2014
A high-throughput, multiplexed assay for superfamily-wide profiling of enzyme activity Bachovchin, Daniel A.; Koblan, Luke W.; Wu, Wengen Nature Chemical Biology, Vol. 10, Issue 8 https://doi.org/10.1038/nchembio.1578	journal	July 2014
Gut microbiota in colorectal cancer: mechanisms of action and clinical applications Wong, Sunny H.; Yu, Jun Nature Reviews Gastroenterology & Hepatology, Vol. 16, Issue 11 https://doi.org/10.1038/s41575-019-0209-8	journal	September 2019
Alleviating Cancer Drug Toxicity by Inhibiting a Bacterial Enzyme Wallace, B. D.; Wang, H.; Lane, K. T. Science, Vol. 330, Issue 6005 https://doi.org/10.1126/science.1191175	journal	November 2010
Genotoxicity of Escherichia coli Nissle 1917 strain cannot be dissociated from its probiotic activity Olier, Maïwenn; Marcq, Ingrid; Salvador-Cartier, Christel Gut Microbes, Vol. 3, Issue 6 https://doi.org/10.4161/gmic.21737	journal	November 2012
Escherichia coli Induces DNA Double-Strand Breaks in Eukaryotic Cells Nougayrede, J. -P. Science, Vol. 313, Issue 5788 https://doi.org/10.1126/science.1127059	journal	August 2006
Deciphering the interplay between the genotoxic and probiotic activities of Escherichia coli Nissle 1917 Massip, Clémence; Branchu, Priscilla; Bossuet-Greif, Nadège PLOS Pathogens, Vol. 15, Issue 9 https://doi.org/10.1371/journal.ppat.1008029	journal	September 2019
Genomic aberrations after short-term exposure to colibactin-producing E. coli transform primary colon epithelial cells Iftekhar, Amina; Berger, Hilmar; Bouznad, Nassim Nature Communications, Vol. 12, Issue 1 https://doi.org/10.1038/s41467-021-21162-y	journal	February 2021
Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix Liebschner, Dorothee; Afonine, Pavel V.; Baker, Matthew L. Acta Crystallographica Section D Structural Biology, Vol. 75, Issue 10 https://doi.org/10.1107/S2059798319011471	journal	October 2019
Critical Intermediates Reveal New Biosynthetic Events in the Enigmatic Colibactin Pathway Li, Zhong-Rui; Li, Yongxin; Lai, Jennifer Y. H. ChemBioChem, Vol. 16, Issue 12 https://doi.org/10.1002/cbic.201500239	journal	June 2015
Overview of the CCP 4 suite and current developments Winn, Martyn D.; Ballard, Charles C.; Cowtan, Kevin D. Acta Crystallographica Section D Biological Crystallography, Vol. 67, Issue 4 https://doi.org/10.1107/S0907444910045749	journal	March 2011
Genetic Structure and Distribution of the Colibactin Genomic Island among Members of the Family Enterobacteriaceae Putze, J.; Hennequin, C.; Nougayrede, J. -P. Infection and Immunity, Vol. 77, Issue 11 https://doi.org/10.1128/IAI.00522-09	journal	August 2009
Amending microbiota by targeting intestinal inflammation with TNF blockade attenuates development of colorectal cancer Yang, Ye; Gharaibeh, Raad Z.; Newsome, Rachel C. Nature Cancer, Vol. 1, Issue 7 https://doi.org/10.1038/s43018-020-0078-7	journal	June 2020
DIALS : implementation and evaluation of a new integration package Winter, Graeme; Waterman, David G.; Parkhurst, James M. Acta Crystallographica Section D Structural Biology, Vol. 74, Issue 2 https://doi.org/10.1107/S2059798317017235	journal	February 2018
A Prodrug Resistance Mechanism Is Involved in Colibactin Biosynthesis and Cytotoxicity Brotherton, Carolyn A.; Balskus, Emily P. Journal of the American Chemical Society, Vol. 135, Issue 9 https://doi.org/10.1021/ja312154m	journal	February 2013
The landscape of somatic mutation in normal colorectal epithelial cells Lee-Six, Henry; Olafsson, Sigurgeir; Ellis, Peter Nature, Vol. 574, Issue 7779 https://doi.org/10.1038/s41586-019-1672-7	journal	October 2019
Boron Containing Compounds as Protease Inhibitors Smoum, Reem; Rubinstein, Abraham; Dembitsky, Valery M. Chemical Reviews, Vol. 112, Issue 7 https://doi.org/10.1021/cr608202m	journal	April 2012
A commensal-encoded genotoxin drives restriction of Vibrio cholerae colonization and host gut microbiome remodeling Chen, Jiandong; Byun, Hyuntae; Liu, Rui Proceedings of the National Academy of Sciences, Vol. 119, Issue 11 https://doi.org/10.1073/pnas.2121180119	journal	March 2022
Mutational signature in colorectal cancer caused by genotoxic pks+ E. coli Pleguezuelos-Manzano, Cayetano; Puschhof, Jens; Rosendahl Huber, Axel Nature, Vol. 580, Issue 7802 https://doi.org/10.1038/s41586-020-2080-8	journal	February 2020
The versatility of boron in biological target engagement Diaz, Diego B.; Yudin, Andrei K. Nature Chemistry, Vol. 9, Issue 8 https://doi.org/10.1038/nchem.2814	journal	July 2017
In Vitro Characterization of the Colibactin-Activating Peptidase ClbP Enables Development of a Fluorogenic Activity Probe Volpe, Matthew R.; Wilson, Matthew R.; Brotherton, Carolyn A. ACS Chemical Biology, Vol. 14, Issue 6 https://doi.org/10.1021/acschembio.9b00069	journal	May 2019
Activity-based protein profiling: The serine hydrolases Liu, Y.; Patricelli, M. P.; Cravatt, B. F. Proceedings of the National Academy of Sciences, Vol. 96, Issue 26 https://doi.org/10.1073/pnas.96.26.14694	journal	December 1999
One Step Forward for Reducing False Positive and False Negative Compound Identifications from Mass Spectrometry Metabolomics Data: New Algorithms for Constructing Extracted Ion Chromatograms and Detecting Chromatographic Peaks Myers, Owen D.; Sumner, Susan J.; Li, Shuzhao Analytical Chemistry, Vol. 89, Issue 17 https://doi.org/10.1021/acs.analchem.7b00947	journal	August 2017

Similar Records

Structural basis of colibactin activation by the ClbP peptidase

Journal Article · 2022 · Nature Chemical Biology · OSTI ID:2423638

Structural basis of the amidase ClbL central to the biosynthesis of the genotoxin colibactin

Journal Article · 2023 · Acta Crystallographica. Section D. Structural Biology · OSTI ID:2423641

ClbM is a versatile, cation-promiscuous MATE transporter found in the colibactin biosynthetic gene cluster

Journal Article · 2017 · Biochemical and Biophysical Research Communications · OSTI ID:22696801

Related Subjects

Biochemistry & Molecular Biology

A small molecule inhibitor prevents gut bacterial genotoxin production

Citation Formats

References (58)

Similar Records

Related Subjects