How Proteins Fold Deciphered by Hydrogen Exchange

Abstract

Biochemical pathways have almost universally been solved by isolating the pathway intermediates and determining their structures. This approach fails for protein folding pathways. Folding intermediates that live for milliseconds cannot be isolated and solved. It has been possible to determine the structure and properties of intermediates during kinetic folding by hydrogen exchange (HX) methods. Having reached the native state proteins repeatedly unfold and refold. The normally invisible intermediates can be characterized by HX. Three concepts emerge. Proteins act like accretions of cooperative unfolding/refolding units called foldons. They account for the unit steps in folding pathways (and other functions). During folding the stepwise formation of foldons is guided by pre‐existing structure in a sequential stabilization process. These two factors generate predetermined stepwise pathways that progressively assemble the foldon units of the target protein. Experimental results that have been interpreted differently, in terms of multiple independent pathways, are due to chance misfolding errors that cause different population fractions to fold differently and so appear to represent different pathways. The integration of these three concepts ‐ foldon units, sequential stabilization, and optional errors ‐ explains much of the vast literature on the folding behavior of protein molecules.