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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, colloid 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. [ABSTRACT FROM AUTHOR]

Intracellular pathogens include all viruses, many bacteria and parasites capable of invading and surviving within host cells. Key to survival is the subversion of host cell pathways by the pathogen for the purpose of propagation and evading the immune system. The intracellular bacterium Shigella flexneri, the causative agent of bacillary dysentery, invades host cells in a vacuole that is subsequently ruptured to allow growth of the pathogen within the host cytoplasm. S. flexneri invasion has been classically described as a macropinocytosis-like process, however the underlying details and the role of macropinosomes in the intracellular bacterial lifestyle have remained elusive. We applied dynamic imaging and advanced large volume correlative light electron microscopy (CLEM) to study the highly transient events of S. flexneri’s early invasion into host epithelial cells and elucidate some of its fundamental features. First, we demonstrate a clear distinction between two compartments formed during the first step of invasion: the bacterial containing vacuole and surrounding macropinosomes, often considered identical. Next, we report a functional link between macropinosomes and the process of vacuolar rupture, demonstrating that rupture timing is dependent on the availability of macropinosomes as well as the activity of the small GTPase Rab11 recruited directly to macropinosomes. We go on to reveal that the bacterial containing vacuole and macropinosomes come into direct contact at the onset of vacuolar rupture. Finally, we demonstrate that S. flexneri does not subvert pre-existing host endocytic vesicles during the invasion steps leading to vacuolar rupture, and propose that macropinosomes are the major compartment involved in these events. These results provide the basis for a new model of the early steps of S. flexneri epithelial cell invasion, establishing a different view of the enigmatic process of cytoplasmic access by invasive bacterial pathogens. [ABSTRACT FROM AUTHOR]