Metallochaperone UreG serves as a new target for design of urease inhibitor: A novel strategy for development of antimicrobials

摘要

Urease as a potential target of antimicrobial drugs has received considerable attention given its versatile roles in microbial infection. Development of effective urease inhibitors, however, is a significant challenge due to the deeply buried active site and highly specific substrate of a bacterial urease. Conventionally, urease inhibitors are designed by either targeting the active site or mimicking substrate of urease, which is not efficient. Up to now, only one effective inhibitor—acetohydroxamic acid (AHA)—is clinically available, but it has adverse side effects. Herein, we demonstrate that a clinically used drug, colloidal bismuth subcitrate, utilizes an unusual way to inhibit urease activity, i.e., disruption of urease maturation process via functional perturbation of a metallochaperone, UreG. Similar phenomena were also observed in various pathogenic bacteria, suggesting that UreG may serve as a general target for design of new types of urease inhibitors. Using Helicobacter pylori UreG as a showcase, by virtual screening combined with experimental validation, we show that two compounds targeting UreG also efficiently inhibited urease activity with inhibitory concentration (IC)_(50)values of micromolar level, resulting in attenuated virulence of the pathogen. We further demonstrate the efficacy of the compounds in a mammalian cell infection model. This study opens up a new opportunity for the design of more effective urease inhibitors and clearly indicates that metallochaperones involved in the maturation of important microbial metalloenzymes serve as new targets for devising a new type of antimicrobial drugs. Author summary Urease, a metalloenzyme that catalyzes the hydrolysis of urea, plays important roles in the survival and virulence of many microbial pathogens, and has long been considered an important drug target for the development of novel antimicrobials. However, its deeply buried active site and highly specific substrate of bacterial urease make it very challenging to design effective urease inhibitors by conventional approaches. In this study, we reveal that a bismuth-based drug (colloidal bismuth subcitrate) inhibits urease activity in an unusual way. This drug binds the urease accessary protein UreG and inhibits its GTPase activity, thus perturbing nickel insertion into the apo-urease, a process called urease maturation. UreG is therefore proposed as an alternative target for the development of urease inhibitors. Using H . pylori UreG as an example, combined with virtual screening and experimental validation, we further show that several small molecules that bind and functionally disrupt UreG could indeed inhibit urease activity in bacteria and in a cell infection model and possess potent antimicrobial activity. In summary, we discovered metallochaperone UreG as a new target for the design of urease inhibitors. Such a strategy should have a broad application in the development of metalloenzyme inhibitors.