flhE belongs to the flhBAE flagellar operon in Enterobacteria, whose first two members function in Type III secretion (T3S). In Salmonella enterica, absence of FlhE affects swarming but not swimming motility. Based on a chance observation of a ‘green’ colony phenotype of flhE mutants on pH indicator plates containing glucose, I have established that this phenotype is associated with lysis of flagellated cells in an acidic environment created by glucose metabolism. The flhE mutant phenotype of Escherichia coli is similar overall to that of S. enterica, but is seen in the absence of glucose and unlike in S. enterica, causes a substantial growth defect. flhE mutants have a lowered cytoplasmic pH in both bacteria, indicative of a proton leak. GFP reporter assays indicate that the leak is dependent on the flagellar system, is present before the T3S system switches to secretion of late substrates, but gets worse after the switch and upon filament assembly, leading to cell lysis. I show that FlhE is a periplasmic protein, which co-purifies with flagellar basal bodies. Also, co-localization of fluorescent fusion proteins suggests a plausible interaction between FlhE and FlhA, implicated in channeling protons for PMF-driven secretion. These results imply that FlhE may act as a plug or a chaperone to regulate proton flow through the flagellar T3S system. I have obtained crystals of the FlhE protein. X-ray data show that the FlhE crystal belongs to space group P212121 and is diffracted to 2.02 Å. Completion of this study will contribute to a better understanding of the proposed role of FlhE. / text
| Identifer | oai:union.ndltd.org:UTEXAS/oai:repositories.lib.utexas.edu:2152/27205 |
| Date | 07 November 2014 |
| Creators | Lee, Jaemin |
| Source Sets | University of Texas |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf |
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