The prevalence of antibiotic resistance among pathogenic bacteria has become a major health concern and has spurred the search for novel antibiotic targets. A particularly promising target is the superfamily of bacterial ATP-binding cassette (ABC) transporters, which couple the hydrolysis of ATP to the transport of a wide variety of solutes across the cell membrane. Bacterial ABC transporters work in conjunction with a high affinity solute binding protein (SBP) that specifically binds substrate and delivers it to the transporter. In Salmonella enterica and Streptococcus pneumoniae among others, disruption of genes encoding ABC transporters and SBPs specific for Zn dramatically attenuates virulence in animal models, highlighting these systems as potent drug targets. We have identified two Zn-specific ABC transporter operons in Paracoccus denitrificans, ZnuABC and AztABCD, and have characterized a hitherto hypothetical protein (AztD) that acts as a Zn chaperone, directly transferring Zn to the SBP of that system (AztC). This project will utilize P. dentrificans as a model for highly homologous systems in human pathogens belonging to the carbapenem-resistant Enterobacteriacaea (CRE). These organisms are associated with broad- spectrum antimicrobial resistance and are the causative agents of potentially deadly nosocomial infections. We will determine the precise mechanism of metal binding and transfer for AztC and AztD proteins from P. denitrificans and the CRE pathogen Citrobacter koseri using structural and biophysical techniques. The physiological roles of the Azt and Znu systems will be determined by making genetic knockouts of these genes in P. denitrficans and characterizing growth deficient phenotypes in Zn-limited medium. High-resolution structural information combined with in vivo functionality will yield new insight into the mechanisms of transition metal import in bacteria and potentially provide a basis for the rational design of metal uptake inhibitors as antibiotics for multi- drug resistant pathogens. !
The widespread emergence of antibiotic resistance in bacteria has placed a renewed emphasis on the identification of more effective antibiotics. ABC transporter systems, which enable pathogenic bacteria to acquire metals and other essential nutrients from the host, represent a new class of potential antibiotic target. This project aims to characterize how Zn-specific ABC transporters enable bacteria to acquire this essential metal from the environment, providing structural and functional information that may eventually lead to the production of effective remedies for antibiotic resistant infections.
|Neupane, Durga P; Kumar, Santosh; Yukl, Erik T (2018) Two ABC Transporters and a Periplasmic Metallochaperone Participate in Zinc Acquisition in Paracoccus denitrificans. Biochemistry :|