The group A Streptococcus (GAS) is a cause of significant morbidity throughout the world and must be considered one of the major human bacterial pathogens. The spectrum of GAS disease includes pharyngitis, skin infections, life-threatening soft tissue infections, and the unique postinfectious complications of acute rheumatic fever and glomerulonephritis. One factor that appears to be important in modulating the type of infection is the polysaccharide capsule surrounding the bacteria. Abundant amounts of capsule are associated with more severe infection. The mechanism and regulation of capsule expression in GAS are not well understood. Preliminary evidence from this laboratory suggests that the capsule may be synthesized in the cytoplasm and transported across the cell membrane by means of an ATP binding cassette (ABC) transport system. The initial reports on ABC transporters described the uptake of nutrients by gram-negative bacteria. In the past several years it has become apparent that this family of transporters is ubiquitous and is involved in such critical but diverse activities as the regulation of sporulation in Bacillus subtilis and the export of chemotherapeutic agents in human cells resulting in a multidrug-resistance phenotype. The proposed research is designed to characterize an ABC transport system in GAS. In phase I of the study, the specific aims are to identify and clone the genes of the GAS transporter, to develop an overexpression system for the transporter, and to purify the transporter proteins. These studies will be done under the supervision of Dr. Michael Wessels. During this period the Principal Investigator will take a series of courses on protein and membrane biochemistry and bacterial pathogenesis in order to gain through a combination-of benchwork and focused instruction the necessary experience to proceed to the second phase of the project. In phase II the biochemistry of the GAS transporter will be investigated. The specific goal is to reconstitute transport activity using the purified proteins in membrane and proteoliposome vesicles. In this manner the function of the transporter will be established. These studies will be conducted under the guidance of Dr. Guido Guidotti. A detailed genetic and biochemical description of the GAS transporter will provide important information on the biology of GAS and the function of ABC transporters. In the process of accomplishing these goals, the Principal Investigator will be trained as an independent researcher in the field of bacterial pathogenesis.
