Examination of the structural changes of small conductivity mechanosensitive ion channel (MscS) as an antibacterial drug target

Objective: The small conductance mechanosensitive (MS) ion channel (MscS) in Escherichia coli (E. coli) (EcMscS) is a heptameric integral membrane protein that responds to increased tension in the lipid bilayer by opening its pores to release solutes and reduce the increased cytoplasmic pressure. Therefore, MS ion channels are thought to have potential as antibacterial drug targets. However, the mechanism of cellular access of solutes passing through EcMscS is not clear yet. The aim of our thesis project is to determine the structural changes of EcMscS as an antibacterial drug target and to explain the gating mechanisms with a multidisciplinary approach using biochemical, structural biology and in-silico techniques. Method: The effects of viomycin on cell viability in E. coli strains with and without MscS ion channel were compared in a concentration-dependently. EcMscS:MSP1E3D1:(16:1) obtained by nanodisc reconstitution was resolved by Cryo-Electron Microscopy (CryoEM) techniques. The transition of viomycin from EcMscS was modeled by MD simulation techniques and the findings were compared with the CryoEM results. Results: It was determined that viomycin decreased cell viability in MscS-expressed strains in a concentration-dependently. The structure of EcMscS:MSP1E3D1:(16:1) (1:10:500) with a closed conformation at 3.4 Å resolution was obtained by CryoEM methods. It was observed that the channel was able to maintain its closed form throughout the 100 ns MD simulation for the same lipid type (DYPC 16:1). Also, the binding sites of viomycin to EcMscS were identified. Conclusion: It was observed that viomycin affected cell viability in MscS-containing E.coli strains and may have a potential to pass through the channel. As a result of cryoEM and MD experiments, it was thought that the PC:16 lipid types may be effective in the closed conformation of EcMscS. As our knowledge is deepened about the three-dimensional conformational structure of MscS and the functions of lipid types, the potential of MscS as an antibacterial drug target will increase. Keywords: MscS, Viomycin, Cell viability, CryoEM, MD simulations.