Threading of Single Protein Molecules Through a Solid-State Nanopore.

The building blocks and machinery of cells that maintain homeostasis and functional biological architecture are proteins. Protein analysis is a crucial aspect of modern biology, biophysics, biochemistry, and medicine, with numerous applications ranging from disease diagnostics to drug development. In recent years, nanopore technology has emerged as a promising approach for probing and characterizing proteins at the single-molecule level. A tiny hole in a biological or solid-state membrane is known as a nanopore, which can be used to detect the physical and chemical behavior of single protein molecules as they pass through the analyzer nanopore. This instructional book chapter helps users with a manual and a point of reference for employing nanopores for single-molecule measurements and analysis of protein translocation through a nanopore. Nanopore technology requires simple and easy protein sample preparation and can detect ultra-low-abundance of water-soluble proteins. One of its most important advantages is the capacity to deliver real-time analysis of label-free protein's structure and dynamics with sub-microseconds time resolution. As proteins pass through the nanopore, they can disrupt the ionic current flowing through the pore, which can give information about the protein's size, shape, sequences, conformation changes (folding and unfolding), short-lived intermediate states, transition states, and metastable states.