Targeting Helicobacter pylori urease activity and maturation: In-cell high-throughput approach for drug discovery

Abstract

BackgroundHelicobacter pylori is a bacterium strongly associated with gastric cancer. It thrives in the acidic environment of the gastric niche of large portions of the human population using a unique adaptive mechanism that involves the catalytic activity of the nickel-dependent enzyme urease. Targeting urease represents a key strategy for drug design and H. pylori eradication. MethodHere, we describe a novel method to screen, directly in the cellular environment, urease inhibitors. A ureolytic Escherichia coli strain was engineered by cloning the entire urease operon in an expression plasmid and used to test in-cell urease inhibition with a high-throughput colorimetric assay. A two-plasmid system was further developed to evaluate the ability of small peptides to block the protein interactions that lead to urease maturation. ResultsThe developed assay is a robust cellular model to test, directly in the cell environment, urease inhibitors. The efficacy of a co-expressed peptide to affect the interaction between UreF and UreD, two accessory proteins necessary for urease activation, was observed. This event involves a process that occurs through folding upon binding, pointing to the importance of intrinsically disordered hot spots in protein interfaces. ConclusionsThe developed system allows the concomitant screening of a large number of drug candidates that interfere with the urease activity both at the level of the enzyme catalysis and maturation. General significanceAs inhibition of urease has the potential of being a global antibacterial strategy for a large number of infections, this work paves the way for the development of new candidates for antibacterial drugs.