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Updated: May 22, 2026

Sample Preparation using a Lipid Monolayer Method for Electron Crystallographic Studies
Published on: November 20, 2021
The 1.8 Å cholix toxin crystal structure in complex with NAD+ and evidence for a new kinetic model
Robert J Fieldhouse1, René Jørgensen, Miguel R Lugo
1Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
Abstract:
Certain Vibrio cholerae strains produce cholix, a potent protein toxin that has diphthamide-specific ADP-ribosyltransferase activity against eukaryotic elongation factor 2. Here we present a 1.8 Å crystal structure of cholix in complex with its natural substrate, nicotinamide adenine dinucleotide (NAD(+)). We also substituted hallmark catalytic residues by site-directed mutagenesis and analyzed both NAD(+) binding and ADP-ribosyltransferase activity using a fluorescence-based assay. These data are the basis for a new kinetic model of cholix toxin activity. Further, the new structural data serve as a reference for continuing inhibitor development for this toxin class.
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