Nanopore Sensing: Towards A Dual-Modality Platform for Simultaneous Electrical and Optical Tracking

Biological nanopores have become central tools in single-molecule sensing, yet the fundamental mechanisms governing their stability and gating are not yet fully understood. Pore-forming toxins (PFTs) represent an essential and conserved class of proteins. Based on their transmembrane secondary structures, they are broadly divided into alpha-PFTs and beta-PFTs, with beta-barrel nanopores playing key roles in biological processes such as selective ion transport, intercellular communication, and pathogenic mechanisms. Beyond their physiological roles, these structures serve as biosensors for DNA and protein sequencing, as well as single-molecule detection. However, translating biological pores into robust analytical platforms presents significant challenges, mainly due to an incomplete understanding of their structure-function relationships. Based on an internship carried out at the LBEN (Laboratory of Nanoscale Biology) of EPFL, this thesis focuses on the physical and chemical principles underlying nanopore behavior. Specifically, the main goal is to develop a dual platform that enables the synchronization of electrical and optical readouts for real-time channel characterization. To do so, planar lipid bilayers were reconstituted using DPhPC lipids to monitor wild-type and engineered aerolysin mutants. By conjugating specific fluorophores to the protein structure, single-channel ionic current measurements were simultaneously coupled with optical tracking. This multimodal approach allows transient gating events and membrane-insertion dynamics to be directly investigated, providing a clearer link between structural transitions and functional electrical signatures.