Role of a cell wall assembly factor in enterococcal antimicrobial resistance

Medical College Of Wisconsin, Milwaukee WI

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Abstract

PROJECT SUMMARY Nearly all Gram-positive bacteria synthesize a transmembrane, Ser/Thr kinase containing 3-5 extracellular PASTA-domains (?PASTA kinase?) that controls critical processes including antibiotic resistance, toxin production, virulence, or cell division; in some bacteria this kinase is essential for viability. As such, these kinases represent attractive targets for new therapeutics. However, a basic understanding of the mechanisms by which kinases in this family function to perceive environmental stimuli in vivo and process that information to coordinate adaptive biological responses is lacking. Such information is critical to inform development of new therapeutic approaches. The research proposed here seeks to help address this gap by elucidating fundamental aspects of function for a representative kinase in this family, the IreK kinase in Enterococcus faecalis, which we have shown is required for intrinsic resistance of E. faecalis to cell-wall-active antibiotics and to detergents present in bile, such as cholate. This research is designed to elucidate new insights into the function of the IreK kinase by exploring its physical and functional relationships with other cell wall assembly factors in E. faecalis. In particular, we will define the composition of a protein complex thought to be involved in cell wall assembly, assess its role in intrinsic antimicrobial resistance, and examine its interactions both physical and functional with IreK to understand how IreK carries out its duties. This work has the potential to establish a new model for transmembrane signaling by PASTA kinases in bacteria and provide new insights into antimicrobial resistance, coordination of cell wall integrity, virulence and other processes in Gram-positive bacteria.