Date Available

5-2-2013

Year of Publication

2013

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Arts and Sciences

Department/School/Program

Chemistry

First Advisor

Dr. Yinan Wei

Abstract

Multiple drug resistance (MDR) has been a severe issue in treatment and recovery from infection.Gram-negative bacteria intrinsically exhibit higher drug tolerance than Gram-positive microbes. In this thesis, two proteins involved in Gram-negative bacterial MDR were studied, AcrB and SurA.

Resistance-nodulation-cell division pump AcrAB-TolC is the major MDR efflux system in Gram-negative bacteria and efficiently extrudes a broad range of substances from the cells. To study subtle conformational changes of AcrB in vivo, a reporter platform was designed. Cysteine pairs were introduced into different regions in the periplasmic domain of the protein, and the extents of disulfide bond formation were examined. Using this platform, an inactive mutant, AcrB∆loop, was created that existed as a well-folded monomer in vivo. Next, random mutageneses were performed on a functionally compromised mutant, AcrBP223G, to identify residues that restored the function loss. The mechanism of function restoration was examined.

SurA is a periplasmic molecular chaperone for outer membrane biogenesis. Deletion of SurA decreased outer membrane density and bacterial drug resistance. The dependence of SurA function on structural flexibility and stability was examined. In addition, the effect of molecular crowding on SurA interaction with its outer membrane protein substrates was examined.

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