Project 1

Researcher: Dr. Chris Colbert
Project Title:  Understanding Outer Membrane Signaling in Gram-Negative Bacteria

Pathogenicity and the ability to form biofilms of Gram-negative bacteria are dependent on the acquisition of iron. Iron is imported from the environment via specific, high affinity, iron chelating molecules, called siderophores that bind to a set of tightly-regulated, high affinity, Ton-B dependent outer membrane transporters (TBDTs). The transcription of TBDTs is tightly regulated by multiple mechanisms, one of which up-regulates TBDT transcription in response to signals induced by extracellular iron-laden siderophore. This self-transcriptional regulation is conserved amongst all Gram-negative bacteria. Our central hypothesis is that siderophore binding on the cell surface activates TBDT transcriptional regulation by a series of protein-protein interaction events that cross both the outer and inner membranes to release an intracellular transcription factor that turns on operon transcription. The goal of this proposal is to investigate the structural basis of these protein-protein interaction events at the membrane interfaces as well as in the periplasmic space. We have selected the Gram-negative bacteria Pseudomonas putida to study these protein interaction events. To accomplish our goal we will use structural (X-ray crystallography and NMR spectroscopy) and biophysical methods to determine three-dimensional structures of the proteins involved and to investigate the mechanism of signal transduction that results in up-regulation of transcription of TBDTs.