Structure Elucidation of Colibactin and Its DNA Cross-Links
Open accessAbstract
Double warhead does DNA damage: Strains of the human gut bacterium Escherichia coli carrying the clb gene cluster produce a secondary metabolite dubbed colibactin and have been provocatively linked to colorectal cancer in some models. Colibactin has been difficult to isolate in full, but pieces of the structure have been worked out, including an electrophilic warhead. Xue et al. found that colibactin contains two conjoined warheads, which is consistent with its ability to alkylate and cross-link DNA. Chemical synthesis and comparison to cell coculture confirm the structure and properties of this unstable and potentially carcinogenic metabolite. Science , this issue p. eaax2685
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Metabolites produced by some intestinal microbiota may be genotoxic and can directly induce DNA damage of host cells or modulate host cell signaling pathways involved in cell proliferation and apoptosis.
“Metabolites produced by some intestinal microbiota may be genotoxic and can directly induce DNA damage of host cells or modulate the basic host cell signaling pathways involved in cell proliferation and apoptosis.”
From The Role of Gut Microbiota in Lung Cancer: From Carcinogenesis to Immunotherapy (2021) · cites this paper as [63]
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Jiang, Y., Stornetta, A., Villalta, P. W., Wilson, M. R., Boudreau, P. D., Zha, L., Balbo, S., & Balskus, E. P. (2019) · Journal of the American Chemical Society · doi:10.1021/jacs.9b02453 · Cited by 66 · see citing works
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“coli promote DDRs by inducing DSBs, probably by DNA alkylation on adenine residues [63,67], SSBs and inter-strand crosslinks in DNA of eukaryotic cells [57,68].”
BackgroundShrestha, E., Coulter, J. B., Guzman, W., Özbek, B., Hess, M. M., Mummert, L. A., Ernst, S. E., Maynard, J. P., Meeker, A. K., Heaphy, C. M., et al. (2021) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.2018976118 · Cited by 58 · see citing works
The relationship between gastrointestinal cancers and the microbiota
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“The structure of colibactin and its DNA cross-links has been solved recently (Xue et al., 2019), and importantly, colibactin DNA-damage signatures have been found in CRC patients (Dziuban ́ ska-Kusibab et al., 2020).”
BackgroundRecent Advances in Mass Spectrometry-Based Structural Elucidation Techniques
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A small molecule inhibitor prevents gut bacterial genotoxin production
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Contribution of pks+ Escherichia coli (E. coli) to Colon Carcinogenesis
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“One would speculate that modulating the oxygen levels in the gut may offer a potential tool for preventing or treating colon cancer associated with colibactin-producing bacteria [130].”
BackgroundUropathogenic E. coli induces DNA damage in the bladder
Chagneau, C. V., Massip, C., Bossuet‐Greif, N., Fremez, C., Motta, J., Shima, A., Besson‐Fournier, C., Faouder, P. L., Cénac, N., Roth, M., et al. (2021) · PLoS Pathogens · doi:10.1371/journal.ppat.1009310 · Cited by 46 · see citing works
Microbiota in cancer: molecular mechanisms and therapeutic interventions
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Self-citationGenotoxins: The Mechanistic Links between Escherichia coli and Colorectal Cancer
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“Active colibactin is composed of two complex biosynthetic intermediates with a nearly symmetrical structure that contains two electrophilic cyclopropane warheads [55].”
BackgroundInfluentialThe microbial genotoxin colibactin exacerbates mismatch repair mutations in colorectal tumors
Dougherty, M. W., Valdés‐Mas, R., Wernke, K. M., Gharaibeh, R. Z., Yang, Y., Brant, J. O., Riva, A., Muehlbauer, M., Elinav, E., Puschhof, J., et al. (2023) · Neoplasia · doi:10.1016/j.neo.2023.100918 · Cited by 45 · see citing works
“Infection of eukaryotic cells with pks + E. coli results in DNA interstrand cross-links (ICLs) [10,11] that are detectable after colonization in vivo [12].”
Self-citationPark, H. B., Wei, Z., Oh, J., Xu, H., Kim, C., Wang, R., Wyche, T. P., Piizzi, G., Flavell, R., & Crawford, J. (2020) · Nature Microbiology · doi:10.1038/s41564-020-0763-4 · Cited by 39 · see citing works
Self-citationDeciphering Human Microbiota–Host Chemical Interactions
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ClbR Is the Key Transcriptional Activator of Colibactin Gene Expression in Escherichia coli
Wallenstein, A., Rehm, N., Brinkmann, M., Selle, M., Bossuet‐Greif, N., Sauer, D., Bunk, B., Spröer, C., Wami, H. T., Homburg, S., et al. (2020) · mSphere · doi:10.1128/msphere.00591-20 · Cited by 40 · see citing works
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The pks island: a bacterial Swiss army knife? Colibactin: beyond DNA damage and cancer
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Oral and intestinal bacterial exotoxins: Potential linked to carcinogenesis
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Escherichia coli small molecule metabolism at the host–microorganism interface
Gatsios, A., Kim, C. S., & Crawford, J. M. (2021) · Nature Chemical Biology · doi:10.1038/s41589-021-00807-5 · Cited by 39 · see citing works
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“Colibactin is a genotoxin that causes inter-strand cross-links (ICLs) [5] and doublestrand DNA breaks (DSBs), cell cycle arrest, senescence, and chromosomal abnormalities in mammalian cells [6].”
BackgroundThe Role of Microbiota-Derived Metabolites in Colorectal Cancer
Duizer, C., & de Zoete, M. R. (2023) · International Journal of Molecular Sciences · doi:10.3390/ijms24098024 · Cited by 37 · see citing works
“The structure of colibactin allows for its reactivity with DNA as two cyclopropane electrophiles are responsible for DNA alkylation [84,85].”
BackgroundChen, J., Byun, H., Liu, R., Jung, I.-J., Pu, Q., Zhu, C. Y., Tanchoco, E., Alavi, S., Degnan, P. H., Ma, A. T., et al. (2022) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.2121180119 · Cited by 37 · see citing works
“C14-D-Asn is both the hydrolysis product of an intermediate synthesized on ClbN as well as the ClbP-catalyzed cleavage product of precolibactin-1492 (24).”
BackgroundFrontier studies on natural products: moving toward paradigm shifts
Zhao, J., & Yue, J. (2023) · Science China Chemistry · doi:10.1007/s11426-022-1512-0 · Cited by 36 · see citing works
Formation and repair of unavoidable, endogenous interstrand cross-links in cellular DNA
Housh, K., Jha, J. S., Haldar, T., Amin, S. B. M., Islam, T., Wallace, A., Gomina, A., Guo, X., Nel, C., Wyatt, J. W., et al. (2020) · DNA repair · doi:10.1016/j.dnarep.2020.103029 · Cited by 37 · see citing works
Mohite, O. S., Lloyd, C. J., Monk, J. M., Weber, T., & Palsson, B. Ø. (2022) · Synthetic and Systems Biotechnology · doi:10.1016/j.synbio.2022.04.011 · Cited by 34 · see citing works
“Very recently, the structure of the final colibactin product was revealed along with complete biosynthesis, however the export outside the bacteria still remains elusive [13,45].”
BackgroundWami, H. T., Wallenstein, A., Sauer, D., Stoll, M., von Bünau, R., Oswald, É., Müller, R., & Dobrindt, U. (2021) · Microbial Genomics · doi:10.1099/mgen.0.000577 · Cited by 36 · see citing works
“The ability to produce colibactin has been described to increase the pathogenic potential of the producing bacteria and to promote colorectal cancer development [12–17], but has also been related to beneficial effects to the host [3, 18–20].”
Wang, J., Li, D., Chen, L., Cao, W., Kong, L., Zhang, W., Croll, T. I., Deng, Z., Liang, J., & Wang, Z. (2022) · Nature Communications · doi:10.1038/s41467-022-28284-x · Cited by 35 · see citing works
Self-citationBiosynthesis and bioactivities of microbial genotoxin colibactins
Tang, J., Liu, X., Ye, W., Li, Z., & Qian, P. (2022) · Natural Product Reports · doi:10.1039/d1np00050k · Cited by 35 · see citing works
Physiological drivers of pks+ E. coli in colorectal cancer
Jans, M., & Vereecke, L. (2025) · Trends in Microbiology · doi:10.1016/j.tim.2025.04.010 · Cited by 33 · see citing works
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Chemical Mechanisms of Colonization Resistance by the Gut Microbial Metabolome
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Microbial metabolites: cause or consequence in gastrointestinal disease?
Fobofou, S. A. T., & Savidge, T. (2022) · American Journal of Physiology-Gastrointestinal and Liver Physiology · doi:10.1152/ajpgi.00008.2022 · Cited by 33 · see citing works
“The structural elucidation of colibactin, a long-standing chemistry challenge that remained for almost a decade because of colibactin instability and small quantities, was recently solved (97).”
Wong, J. J., Ho, F. K., Choo, P. Y., Chong, K. K. L., Ho, C. M. B., Neelakandan, R., Keogh, D., Barkham, T., Chen, J. L., Liu, C., et al. (2022) · PLoS Pathogens · doi:10.1371/journal.ppat.1010766 · Cited by 33 · see citing works
Depurination of Colibactin-Derived Interstrand Cross-Links
Xue, M., Wernke, K. M., & Herzon, S. B. (2020) · Biochemistry · doi:10.1021/acs.biochem.9b01070 · Cited by 32 · see citing works
Self-citationBaddock, H. T., Yosaatmadja, Y., Newman, J. A., Schofield, C. J., Gileadi, O., & McHugh, P. J. (2020) · DNA repair · doi:10.1016/j.dnarep.2020.102941 · Cited by 34 · see citing works
Queen, J., Shaikh, F. Y., & Sears, C. L. (2023) · Nature Cancer · doi:10.1038/s43018-023-00602-2 · Cited by 33 · see citing works
“Single-center, double-blind, placebo-controlled, double-blind randomized trial Patients with metastatic gastroesophageal cancer and cachexia (24) Donor (healthy obese individuals) versus autologous FMT via nasoduodenal tube Donor FMT did not improve cachexia measures (primary outcome), but higher DCR and longer OS and…”
BackgroundMicrobiota and colorectal cancer: colibactin makes its mark
Arthur, J. C. (2020) · Nature Reviews Gastroenterology & Hepatology · doi:10.1038/s41575-020-0303-y · Cited by 30 · see citing works
Tumor-resident microbes: the new kids on the microenvironment block
Li, L., Chandra, V., & McAllister, F. (2024) · Trends in Cancer · doi:10.1016/j.trecan.2023.12.002 · Cited by 30 · see citing works
“A pathogenic genomic island in E. coli, termed pks, encodes a genotoxic peptide, colibactin, that promotes CRC in experimental mouse models by increasing adherence to epithelial cells [15] and by forming DNA adducts in adenine-rich regions [16,17].”
BackgroundFanconi anemia-independent DNA inter-strand crosslink repair in eukaryotes
Rogers, C. M., Simmons, R. H., III, Thornburg, G. E. F., Buehler, N. J., & Bochman, M. L. (2020) · Progress in Biophysics and Molecular Biology · doi:10.1016/j.pbiomolbio.2020.08.005 · Cited by 28 · see citing works
“In their model, two “precolibactin” structures that each contain a reactive aminocyclopropane group heterodimerize and are each capable of alkylating adenine bases in AT-rich DNA regions, with a second alkylation creating an ICL (Xue et al., 2019; Dziubanska-Kusibab et al., 2020).”
BackgroundCellular Stress Upregulates Indole Signaling Metabolites in Escherichia coli
Kim, C. S., Li, J., Barco, B., Park, H. B., Gatsios, A., Damania, A., Wang, R., Wyche, T. P., Piizzi, G., Clay, N. K., et al. (2020) · Cell chemical biology · doi:10.1016/j.chembiol.2020.03.003 · Cited by 27 · see citing works
Self-citationPiciocchi, A., Germinario, E. A. P., García‐Etxebarria, K., Rossi, S., Sanchez-Mete, L., Porowska, B., Stigliano, V., Trentino, P., Oddi, A., Accarpio, F., et al. (2021) · Toxins · doi:10.3390/toxins13080569 · Cited by 27 · see citing works
Structural basis of colibactin activation by the ClbP peptidase
Velilla, J. A., Volpe, M. R., Kenney, G. E., Walsh, R. M., Balskus, E. P., & Gaudet, R. (2022) · Nature Chemical Biology · doi:10.1038/s41589-022-01142-z · Cited by 27 · see citing works
Structure Elucidation of Secondary Metabolites: Current Frontiers and Lingering Pitfalls
DiBello, M., Healy, A. R., Nikolayevskiy, H., Xu, Z. F., & Herzon, S. B. (2023) · Accounts of Chemical Research · doi:10.1021/acs.accounts.3c00183 · Cited by 26 · see citing works
Self-citationCurrent understandings of colibactin regulation
Addington, E., Sandalli, S., & Roe, A. J. (2024) · Microbiology · doi:10.1099/mic.0.001427 · Cited by 26 · see citing works
“These have revealed that colibactin has a nearly symmetrical structure which is characterised by two electrophilic cyclopropane warheads capable of forming adducts with and alkylating DNA [22, 23, 26, 37–39].”
BackgroundGarofalo, M., Payros, D., Oswald, É., Nougayrède, J., & Oswald, I. P. (2022) · The Science of The Total Environment · doi:10.1016/j.scitotenv.2022.153280 · Cited by 26 · see citing works
“We recently observed that DON increases genotoxicity induced by colibactin, a bacterial toxin that causes peculiar DNA-interstrand crosslink lesions (Payros et al. , 2017; Bossuet-Greif et al. , 2018; Xue et al. , 2019).”
Unraveling the genomic landscape of colorectal cancer through mutational signatures
Díaz‐Gay, M., & Alexandrov, L. B. (2021) · Advances in cancer research · doi:10.1016/bs.acr.2021.03.003 · Cited by 25 · see citing works
Microbe-Driven Genotoxicity in Gastrointestinal Carcinogenesis
Hartl, K., & Sigal, M. J. (2020) · International Journal of Molecular Sciences · doi:10.3390/ijms21207439 · Cited by 24 · see citing works
Zhou, T., Hirayama, Y., Tsunematsu, Y., Suzuki, N., Tanaka, S., Uchiyama, N., Goda, Y., Yoshikawa, Y., Iwashita, Y., Sato, M., et al. (2021) · Journal of the American Chemical Society · doi:10.1021/jacs.1c01495 · Cited by 24 · see citing works
A tale of two bacteria – Bacteroides fragilis, Escherichia coli, and colorectal cancer
Lichtenstern, C. R., & Lamichhane-Khadka, R. (2023) · Frontiers in Bacteriology · doi:10.3389/fbrio.2023.1229077 · Cited by 23 · see citing works
“The structure of colibactin has recently been con fi rmed as containing an a - dicarbonyl group and two electrophilic spirocyclopropyldihydro-2-pyrrolone groups that can undergo ring-opening to attach to DNA (Xue et al., 2019).”
InfluentialCancer microbiota: a focus on tumor-resident bacteria
Vella, G., & Rescigno, M. (2025) · EMBO Reports · doi:10.1038/s44319-025-00482-w · Cited by 25 · see citing works
“This toxin induces DNA adduct formation and double-strand breaks, thereby contributing to genomic instability (Nougayrède et al, 2006; Wilson et al, 2019; Xue et al, 2019).”
Robust performance of a live bacterial therapeutic chassis lacking the colibactin gene cluster
Kalantari, A., James, M. J., Renaud, L., Perreault, M., Monahan, C. E., McDonald, M. N., Hava, D. L., & Isabella, V. M. (2023) · PLoS ONE · doi:10.1371/journal.pone.0280499 · Cited by 21 · see citing works
“The structure of colibactin was only recently solved using a combination of approaches including multiple modifications to the pks genomic island, a number of analytical techniques, and chemical synthesis of proposed and later confirmed structures found within the colibactin biosynthetic pathway [24].”
MethodLeveraging Microbial Genomes and Genomic Context for Chemical Discovery
Kountz, D. J., & Balskus, E. P. (2021) · Accounts of Chemical Research · doi:10.1021/acs.accounts.1c00100 · Cited by 21 · see citing works
Adherent-invasive Escherichia coli in Crohn’s disease: the 25th anniversary
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Extension of Diagnostic Fragmentation Filtering for Automated Discovery in DNA Adductomics
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Prevalence and implications of pKs-positive Escherichia coli in colorectal cancer
de Souza, J. B., Campos, L. A. D. A., Palácio, S. B., de Castro, M. C. A. B., & Cavalcanti, I. M. F. (2024) · Life Sciences · doi:10.1016/j.lfs.2024.122462 · Cited by 20 · see citing works
Human microbiota peptides: important roles in human health
Shah, A. B., & Shim, S. H. (2024) · Natural Product Reports · doi:10.1039/d4np00042k · Cited by 19 · see citing works
Zechner, E. L., & Kienesberger, S. (2024) · Gut Microbes · doi:10.1080/19490976.2024.2430423 · Cited by 20 · see citing works
“66 ICLs impede DNA replication and require repair in a replication-coupled manner.”
The microbiome-product colibactin hits unique cellular targets mediating host–microbe interaction
Mousa, W. K. (2022) · Frontiers in Pharmacology · doi:10.3389/fphar.2022.958012 · Cited by 19 · see citing works
Oxygen concentration modulates colibactin production
Bossuet‐Greif, N., Guyonnet, C., Chagneau, C. V., Tang-Fichaux, M., Pénary, M., Loubet, D., Branchu, P., Oswald, É., & Nougayrède, J. (2023) · Gut Microbes · doi:10.1080/19490976.2023.2222437 · Cited by 18 · see citing works
Bacterial small molecule metabolites implicated in gastrointestinal cancer development
Turocy, T., & Crawford, J. M. (2024) · Nature Reviews Microbiology · doi:10.1038/s41579-024-01103-4 · Cited by 20 · see citing works
“80,81).”
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- Authors
- Xue, M., Kim, C. S., Healy, A. R., Wernke, K. M., Wang, Z., Frischling, M. C., Shine, E. E., Wang, W., Herzon, S. B., & Crawford, J. M.
- Affiliations
- Yale University (US)
- Corresponding author
- Xue, Mengzhao
- Kim, Chung Sub
- Herzon, Seth B.
- Crawford, Jason M.
- ORCID
- Venue
- Science
- Published
- 2019-09-06
- Volume
- 365
- Issue
- 6457
- ISSN
- 0036-8075 (ISSN-L)
- 1095-9203
- PMC
- PMC6820679
- OpenAlex
- W2967933500
- Cited by
- 257
- References
- 47
- Citations by year
- 2026: 24 · 2025: 42 · 2024: 40 · 2023: 26 · 2022: 38 · 2021: 43 · 2020: 34 · 2019: 10
- FWCI
- 19.37
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Science (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- Funders
- National Science Foundation
- National Institutes of Health (R01GM110506, 1DP2-CA186575, R01CA215553, T32GM067543)
- Charles H. Revson Foundation
- Topics
- Health Sciences › Medicine › Pharmacology › Microbial Natural Products and Biosynthesis
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biotechnology › Microbial Metabolism and Applications
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Toxicology › Bioactive Compounds and Antitumor Agents
- Keywords
- colibactin, DNA cross-links, structure elucidation, colorectal cancer, gut microbiota, bacterial virulence factors, isotopic labeling, tandem mass spectrometry, chemical synthesis
- MeSH terms
- Escherichia coli, Isotope Labeling, Mutation, Peptide Hydrolases, Peptides, Protein Conformation, DNA Adducts, Escherichia coli Proteins, Tandem Mass Spectrometry, Polyketides, Secondary Metabolism
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