Intermembrane Transfer and Assembly of the Type II Secretion System of Gram-Negative Bacteria

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The research programme of our laboratory is concerned with the structure and function of the double membrane envelope of Gram-negative bacteria. We study interactions between the inner and outer membrane as well as the transport of materials across the outer membrane. This transport is a fundamental life process since membranes act as a barrier in protecting the cell, but that barrier must be overcome so that nutrients and proteins required for life can cross them. The major focus of our current research is the structure and function of the type two secretion system (T2SS) that allows Gram negative bacteria to secrete degradative enzymes and toxins. In our previous studies we showed that a complex of two membrane proteins, ExeAB, are required for the transport and assembly of the secretin, the outer membrane channel through which T2SS proteins are secreted. We have demonstrated that ExeA binds to peptidoglycan, a major structural component of the cell envelope, and identified critical amino acids of the peptidoglycan binding site. In our current experiments, a major focus is how interactions between ExeAB and peptidoglycan allow the secretin to assemble into a channel in the outer membrane. We hypothesize that ExeAB itself forms a multimeric scaffold to which the secretin binds in order to assemble. We are using biochemical and genetic approaches to study the multiple factors involved during secretin assembly in E. coli and V. cholerae. In E. coli we hypothesize that the secretins of the two T2SS systems in this bacterium are assembled independently, whereas in V. cholerae we have evidence that there are at least two different mechanisms involved in assembly of the same secretin, and have identified a candidate for the second, as yet unidentified mechanism. These studies will provide new information concerning how Gram negative bacteria secrete enzymes and protein toxins, and lead to new strategies for harnessing bacterial protein synthesis and for interfering with toxin secretion.