Characterization of Natural Colibactin–Nucleobase Adducts by Tandem Mass Spectrometry and Isotopic Labeling. Support for DNA Alkylation by Cyclopropane Ring Opening
- 26 October 2018
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 57 (45), 6391-6394
- https://doi.org/10.1021/acs.biochem.8b01023
Abstract
Colibactins are genotoxic secondary metabolites whose biosynthesis is encoded in the clb gene cluster harbored by certain strains of gut commensal Escherichia coli. Using synthetic colibactin analogues, we previously provided evidence that colibactins alkylate DNA by addition of a nucleotide to an electrophilic cyclopropane intermediate. However, natural colibactin–nucleobase adducts have not been identified, to the best of our knowledge. Here we present the first identification of such adducts, derived from treatment of pUC19 DNA with clb+E. coli. Previous biosynthetic studies established cysteine and methionine as building blocks in colibactin biosynthesis; accordingly, we used cysteine (ΔcysE) and methionine (ΔmetA) auxotrophic strains cultured in media supplemented with l-[U-13C]Cys or l-[U-13C]Met to facilitate the identification of nucleobases bound to colibactins. Using MS2 and MS3 analysis, in conjunction with the known oxidative instability of colibactin cyclopropane-opened products, we were able to characterize adenine adducts derived from cyclopropane ring opening. This study provides the first reported detection of nucleobase adducts derived from clb+E. coli and lends support to our earlier model suggesting DNA alkylation by addition of a nucleotide to an electrophilic cyclopropane.Keywords
Funding Information
- National Cancer Institute (1R01CA215553)
- NIH Office of the Director (1S10OD019967, 1S10ODOD018034)
This publication has 18 references indexed in Scilit:
- Colibactin: More Than a New Bacterial ToxinToxins, 2018
- ClbS Is a Cyclopropane Hydrolase That Confers Colibactin ResistanceJournal of the American Chemical Society, 2017
- Molecular Basis of Gut Microbiome-Associated Colorectal Cancer: A Synthetic PerspectiveJournal of the American Chemical Society, 2017
- The Enterobacterial Genotoxins: Cytolethal Distending Toxin and ColibactinEcoSal Plus, 2016
- A Mechanistic Model for Colibactin-Induced GenotoxicityJournal of the American Chemical Society, 2016
- Linking Biosynthetic Gene Clusters to their Metabolites via Pathway- Targeted Molecular NetworkingCurrent Topics in Medicinal Chemistry, 2016
- Escherichia coli ClbS is a colibactin resistance proteinMolecular Microbiology, 2015
- Colibactin: understanding an elusive gut bacterial genotoxinNatural Product Reports, 2015
- Yatakemycin: total synthesis, DNA alkylation, and biological propertiesNatural Product Reports, 2007
- Shape-Dependent Catalysis: Insights into the Source of Catalysis for the CC-1065 and Duocarmycin DNA Alkylation ReactionAccounts of Chemical Research, 1999