Unveiling the biological interface of protein complexes by mass spectrometry‐coupled methods

Abstract

Most biomolecules become functional and bioactive by forming protein complexes through interaction with ligands that are diverse in size, shape, and physicochemical properties. In the complex biological milieu, the interaction is ligand-specific, driven by molecular sensing, and involves the recognition of a binding interface localized within a protein structure. Mapping interfaces of protein complexes is a highly sought area of research as it delivers fundamental insights into proteomes and pathology and hence strategies for therapeutics. While X-ray crystallography and electron microscopy remain the gold standard for structural elucidation of protein complexes, their artificial and static analytic nature often produces a non-native interface that otherwise might be negligible or non-existent in a biological environment. Recently, the mass spectrometry-coupled approaches, chemical crosslinking (CLMS) and hydrogen-deuterium exchange (HDMS) have become valuable analytic complements to the traditional techniques. These methods explicitly identify hot residues and motifs embedded in binding interfaces, especially when the interaction is predominantly dynamic, transient, and/or caused by an intrinsically disordered domain. Here, we review the principal role of CLMS and HDMS in protein structural biology with a particular emphasis on the contribution of recent examples to exploring biological interfaces. Additionally, we describe recent studies that utilized these methods to expand our understanding of protein complex formation and the related biological processes, to increase the probability of structure-based drug design.