Grant: Discovery Grants Program - Individual

Bacteria use sophisticated protein complexes to assemble structures that provide motility, adhere to multiple surfaces, and transfer DNA or proteins across membranes. Our research aims to understand, at a structural and functional level, how these protein systems assemble and interact to affect infection and virulence, increase genetic diversity, and propagate resistance strategies.    One of the most versatile and complex bacterial secretion systems is the type IV secretion system (T4SS), which can be used to actively transfer DNA or proteins into and out of a cell. The process of horizontal gene transfer, also known as conjugation, of mobile plasmids is a main function of T4SSs that enables the rapid spread of advantageous genes within a bacterial population. Conjugative plasmids, such as the F plasmid of E. coli, encode genes required for T4SS assembly, pilus synthesis, plasmid transfer, and prevention of redundant plasmid transfer.     Our research program is interested in understanding the regulation of F plasmid transfer between cells, and understanding how the proteins of the F plasmid T4SS interact structurally to assemble the conjugative pilus. In the current proposal, we are studying three components of the T4SS of the F plasmid: (1) the pore forming proteins TraW and TrbC; (2) the mating pair stabilization/entry exclusion proteins TraG, TraS and TraN; and (3) the T4SS coordinating protein TraH. We will determine high resolution X-ray crystal structures of these proteins, and characterize relevant protein-protein interactions. We will also explore protein dynamics using hydrogen-deuterium exchange mass spectrometry and small angle X-ray scattering. We will relate these structural observations to biological function using protein mutants and mating assays. These studies will provide a comprehensive picture of the T4SS proteins of the F plasmid, and how they functionally interact to result in DNA transfer from one bacterium to another.     The long term goal of our research program is to understand the complex interactions of proteins within the T4SS assembly machinery during conjugation. This research will lead to more targeted strategies to reduce the spread of DNA through T4SS-mediated conjugation.