Mechanisms investigations of O antigen synthesis in gram negative bacteria

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The opportunistic bacteria Pseudomonas aeruginosa (PA), can cause death due to lung, kidney or wound infections and occur in many hospital-acquired infections. These bacteria protect themselves with an outer coat of carbohydrates that play a major role in the interactions between bacteria and their host environment. PA synthesize two types of outer antigenic carbohydrates (O antigens), the common A band and the serotype-specific B band. The genes responsible for the assembly of these carbohydrates are known but their exact functions have not been established, mainly due to lack of appropriate methods. We have recently developed these methods supported by NSERC, and have established the enzymatic functions of a number of gene products from intestinal bacteria E. coli, Shigella and Salmonella. In a related project, we have developed methods to reduce the adhesion of PA to lung cells, which will help to reduce infections. Our goal for this proposal is to understand the biochemical mechanisms and the functions of the enzymes that assemble the outer carbohydrates (O antigens) in PA. We will use the genes responsible for O antigen synthesis to produce the enzymes which will be purified and studied in detail. The P.I. is an expert in this field. We will collaborate with the microbiologist that has cloned these genes in PA, with several chemists that will help to synthesize new carbohydrates for these studies, and with a protein structure expert that will help to analyze the enzymes. We will therefore develop new methods in order to understand how important pathogenic bacteria function. Our long term goal is to develop non-toxic inhibitors to block O antigen synthesis in specific pathogens, thus reducing their virulence. Four graduate students and several undergraduate students will be trained in the fields of microbiology, enzymology, carbohydrate chemistry and protein biochemistry. These trainees will have a unique expertise that can be applied in the academic field, in biotechnology, medical engineering and drug development. This expertise and knowledge will be valuable for Universities, research laboratories and industry of Canada.