Jayesh Arun Bafna

• The outer membrane of Gram-negative bacteria is associated with a channel called porins, designed to facilitate the transport (uptake) of nutrients and small molecules. In biological channels, the flux is very low (typically 1-1,000 s-1), and it is challenging to detect such small quantity through a direct and straightforward approach. To characterise the uptake, a general and simple method is required to quantify such molecules. To access direct information on the flux of nutrient and small molecules, we try to exploit electrophysiology as a method. In this thesis, we use electrophysiology (single molecule electrical detection) as a tool to understand the transport and flux through outer membrane channels. Before approaching the problem of flux quantification, one needs to address a few open questions like: What is the difference between a real translocation and binding? Can we detect small molecules (size < 300 Da)? How can we improvise the detection process? etc. . In electrophysiology, we often observe fluctuation in time series of ion current signal in the presence of a molecule of interest, and thus specific conclusion on the transport properties (event rate and residence time) can be drawn, but it is often difficult to conclude on the real translocations. Next, we probe the forces involved in the translocation of such molecules. Based on the channel preference (selectivity) under the application of biased transmembrane potential, it induces a transverse fluid flow (Electroosmotic flow (EOF)) inside a channel, which drives these molecules through them by either enhancing the effect on the detection (translocation) or by hindering their pathway. Overall, the research carried out in this thesis contributes to the understanding of the electrophysiology-based study of flux quantification and the dynamics of molecular through Outer membrane channel of Gram-negative bacteria in the artificial membrane and native membrane system.