A grant has been awarded to Dr. Philip Silverman of the Oklahoma Medical Research Foundation to elucidate the structural biology of conjugal (lateral) DNA transfer between F+ and F- strains of Escherichia coli. This research seeks to define how the 30 or so DNA transfer (Tra) proteins encoded by the F plasmid are organized at the bacterial cell surface and tests the hypothesis that these proteins constitute a molecular machine that mediates conjugal DNA transfer. The first specific aim is to characterize the interactions among the Tra proteins with the goal of understanding what the Tra protein machine looks like from a biochemical perspective. The second specific aim of the research is to apply indirect immunofluorescence microscopy to determine how many Tra machines there are on each bacterial cell and how they are related spatially to each other, to F DNA, and to the bacterial cell surface as a whole. This research is the first step towards a detailed structure/function analysis of conjugal DNA transfer mediated by the F plasmid. To broaden the impact of our research to science education in Oklahoma's underserved rural schools, a third specific aim of the project is to survey antibiotic resistance plasmids in bacteria from the intestinal flora of livestock throughout Oklahoma. These R plasmids will be isolated and partially characterized by rural high school science students in conjunction with The Oklahoma Science Project, which Dr. Silverman founded and directs. Several considerations make this project both timely and important. First, conjugal DNA transfer is connected to the dissemination of antibiotic resistance, an increasingly alarming public health problem. Second, it is one of several mechanisms by which bacterial cells secrete macromolecules across their surface barriers; these mechanisms are responsible for the pathogenicity of many bacteria. Third, conjugal DNA transfer has applications in biotechnology; a better understanding of the process will increase the number and scope of such applications. Finally, the project will create more opportunities for Oklahoma's rural students, including its sizable Native American cohort, to enter technology career paths.
