Novel approaches to identify small molecules modulating E.coli TolC protein function

  • The urge of new strategies to overcome the world wide health problem of antibiotic resistance induced researchers as well governmental and private institution to come together to increase the medical and scientific knowledge on this topic. This PhD project is part of the ITN Translocation, whose aim was to investigate the molecular and cellular mechanism at the basis of influx and efflux processes in gram-negative bacteria. The work behind the scientific advances reached during my 3-year-journey has been accomplished mainly between the Fraunhofer IME ScreeningPort in Hamburg and the Jacobs University Bremen, enriched by external collaborations with the University of Frankfurt and the Helmholtz Center for Infection Research. Through an interdisciplinary approach, from in silico studies to biophysical characterization in vitro, small molecules hits to be developed as efflux pump inhibitors (EPIs) were identified. In particular, these compounds were shown to bind TolC, part of the major efflux systems in E.coli, in docking studies targeting an acidic pocket present in the periplasmic tip of the channel; to selectively bind TolCWT against a recombinant version, where key residues in the target site are mutated, in a biophysical setup allowing the determination of binding constant; to modulate ion current in reconstituted TolCWT in single-channel electrophysiology measurements. Overall, the findings reported in this PhD thesis increase the knowledge of the biophysical characteristics of TolC through the use of cutting-edge methods and technologies, in particular in the field of membrane channels, and allowed the identification of promising compounds hits to support the development of EPIs to be employed as adjuvants in antimicrobial therapies.

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Publishing Institution:IRC-Library, Information Resource Center der Jacobs University Bremen
Granting Institution:Jacobs Univ.
Author:Alessia Gilardi
Referee:Mathias Winterhalter, Richard Wagner, Philip Gribbon, Björn Windshügel
Advisor:Mathias Winterhalter
Persistent Identifier (URN):urn:nbn:de:gbv:579-opus-1008103
Document Type:PhD Thesis
Date of Successful Oral Defense:2017/07/14
Date of First Publication:2018/06/29
Academic Department:Life Sciences & Chemistry
PhD Degree:Biochemistry
Focus Area:Health
Other Organisations Involved:Fraunhofer IME
Call No:2018/12

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