Biochemical and structural studies of Escherichia coli leucine-responsive regulatory protein

Transcriptional regulation of gene expression is an essential component for the life of any organism. For uropathogenic Escherichia coli, expression of the pap genes, generating the structural and regulatory components of pap pili, has been seen to be correlated with the ability of these bacteria to colonize and infect the urinary tract. Transcription at the pap operon is regulated by a large number of proteins including the global regulator Lrp (leucine-responsive regulatory protein) and the pap-specific regulator PapI. Transcription activation is sensitive to the methylation states of the two GATC sequences contained within the pap promoter. We have been interested in studying specifically how these two regulators, Lrp and PapI, interact with the pap promoter DNA using x-ray crystallography.; Biochemical studies of Lrp and PapI were initiated to learn more about these proteins and enhance our ability to crystallize them. A crystal structure of Lrp in complex with one of its binding sites from the pap promoter has been solved to 3.2A. While there is no clear electron density in the structure for the DNA, comparison of the Lrp structure and the homologous LrpA structure, crystallized in the absence of DNA, provide insight into conformational changes that may be related to DNA binding. In addition, several Lrp mutants were made, with the goal of aiding crystal growth, that have shed light on the ability of Up to bind DNA and the role of cooperativity in this process.