Solution NMR Approaches for Studying Molecular Chaperones

Abstract

Molecular chaperones protect proteomes against factors that perturb protein homeostasis and can lead to a range of debilitating diseases. Chaperones must interact with partially folded, improperly folded or even aggregated substrates specifically, though these interactions are usually transient because substrates must be released in a timely manner. By their nature, partially folded proteins that are chaperone substrates are dynamic and conformationally flexible, traits that are refractory to structural characterization by techniques such as cryogenic electron microscopy and X-ray crystallography. However, nuclear magnetic resonance (NMR) spectroscopy is well suited to the study of conformationally flexible molecules and is sensitive to transiently populated conformations, making it a valuable tool for the study of chaperone–substrate interactions. This chapter focuses on describing the NMR techniques and approaches used for structural determination and for delineating protein–protein interactions of chaperones. It emphasizes the use of exchange-based NMR techniques for investigating sparsely populated protein conformations, an approach that has provided significant insights into chaperone–substrate interactions. Several examples of the applications of these techniques for studies of various types of molecular chaperones are presented.