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Biophysical and biochemical characterization of TolC as a potential target for efflux pump inhibition
Ikujuni, Ayotunde Paul
Ikujuni, Ayotunde Paul
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Abstract
Antibiotic resistance is a major global threat to public health faced by healthcare services. Currently, about 1.27 million deaths are attributable to antibiotic-resistant infections globally. Global deaths attributable to antibiotic resistance have been predicted to increase to about 10 million by 2050 with an estimated global economic burden of about 100 million dollars. Overexpression of efflux pumps has been reported to be directly correlated with antibiotic resistance of different bacterial pathogens. The AcrAB-TolC system which belongs to the resistance-nodulation-division (RND) superfamily of efflux pumps, exclusively found in Gram-negative bacteria, is the best-characterized efflux pump in E. coli. Deletion of the gene that encodes TolC, the outer membrane component of the AcrAB-TolC efflux pump has been reported to significantly decreased bacterial resistance to many antibiotics. However, there is a limited understanding of the mechanism of TolC biogenesis due to its unique structure. This dissertation reports a systemic biophysical and biochemical characterization of TolC. We have developed a very easy-to-use, highly efficient method for the high-yield production of TolC from inclusion bodies. Also, we have used various biophysical techniques to systematically characterize the effect of various detergent properties on the folding of TolC. We have also used some in vitro and in vivo assays to show that the biogenesis of TolC involves the formation of folded periplasmic intermediates. We believe that the works reported in this dissertation will open the door to the design and development of TolC-assembly-pathway-targeted efflux pump inhibitors.
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Date
2023-05-31
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University of Kansas
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961637_1.pdf
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Keywords
Biochemistry, Biophysics, Antibiotic resistance, Efflux Pumps, Outer membrane proteins, Protein characterization, Protein folding, TolC