Targeting capsule synthesis in Klebsiella pneumoniae for novel antibiotic discovery: Purification of capsule regulatory proteins

Authors

• AH Abeykoon
• C Mohammadi
• AM Stanley
• N Majdalani
• S Gottesman
• S Buchanan

Abstract

There is an urgent need for novel antibiotics to treat carbapenem-resistant Klebsiella pneumoniae, a nosocomial infection resistant to the majority of available antibiotics. K. pneumoniae secrete a polysaccharide capsule, which provides protection from the immune system and decreases antibiotic penetrance. The highly conserved Rcs signaling cascade is responsible for the regulation of capsule synthesis. Thus, the regulatory proteins involved in this pathway are strong candidates for therapeutic intervention and small molecules that perturb this pathway are of particular interest. This project, conducted in collaboration with Susan Gottesman’s lab (NCI), examines two important proteins (IgaA and RcsF) involved in the regulation of capsule synthesis, for the purpose of novel antibiotic development. The two goals of this project include high-resolution structural analysis of these proteins and a high-throughput screen (HTS) to identify small molecule modulators of this pathway. Several homologs of IgaA, a negative regulator of the RcsF phosphorelay, were expressed in E. coli and purified on the large scale in multiple detergents. Additionally, RcsF, a periplasmic protein, was purified and, upon interaction with IgaA, resulted in a stable complex. Crystallization conditions are currently being established for both of these proteins

Scientific Focus Area: Structural Biology

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