BPS2025 – Label-free identification of full-length proteins using a nanopore

Proteins have critical importance in all cellular processes and are implicated in a plethora of diseases. Methods for proteoform identification are therefore of high interest but are so far struggling e.g., with low efficiency and yield due to complicated analysis protocols. Here, we demonstrate the ability of an aerolysin nanopore to directly identify full length proteins without the need for chemical linkage, or enzymatic sample-processing. By generating a strong electroosmotic flux, we can translocate full length proteins and demonstrate identification at the single-molecule level using machine learning classifiers. In a comparison of signals from seven related proteins of the Turandot family we explore the influence of the analyte's physicochemical properties on the nanopore readout. We anticipate that this in-depth investigation of protein translocations will help to accelerate the path towards fast and simple nanopore based protein identification.