Controlled Protein Translocation in Nanopores for Sequencing Applications

Project Summary In the field of proteomics, the ultimate goal is to determine the sequence and abundance of every protein variant in a sample. To reach this goal, we need techniques capable of single-molecule, high-accuracy, de novo protein sequencing. Unfortunately, current proteomics technologies lack the sensitivity, dynamic range, throughput, scale, and accuracy needed to meet this goal. During this project, we will develop technological components and methods required to enable state-of-the-art protein sequencing. More specifically, we will develop methods for controllably translocating a protein through a nanopore reader such that the protein can be sequenced with single amino acid resolution. Nanopore-based sequencing provides a path to characterizing both high- and low- abundance proteins by sequencing proteins one molecule at a time, but with the high accuracy, sensitivity and throughput needed to cover the wide dynamic range in protein abundance in the proteome. During this project, we will develop and build a novel prototype sequencing system, fully assess and optimize the associated workflow/methodology for highly accurate, reproducible, and versatile protein/peptide characterization, and demonstrate initial sequencing for various proteins and peptides. At the conclusion of this project, we will have successfully demonstrated feasibility for a practical nanopore-based protein sequencing concept.