3D Proteomics: Structural, Functional, Chemical and Biomarker Discovery Proteomics With LiP-MS

Protein structural dynamics drive changes in protein function, making the capture of such dynamics essential for interrogating biological systems. Here we review limited proteolysis coupled to mass spectrometry (LiP-MS), a structural and chemical proteomics method that uses changes in susceptibility to protease cleavage to profile proteome-wide protein structural changes within complex biological samples. In the decade since its development, LiP-MS has become a broadly used structural proteomics method, with peptide-level resolution. It has identified drug targets, delineated altered cellular pathways in response to complex perturbations, revealed structural information on otherwise challenging protein targets, and demonstrated the new concept of structural biomarkers of disease. Because LiP-MS simultaneously probes numerous types of molecular events, such as molecular binding, changes in enzyme activity, chemical modifications, allosteric conformational changes, aggregation, and unfolding, it supports a new proteomics workflow which we term 3D proteomics. This workflow enables the detection of specific functional sites within proteins that are altered upon perturbation, thereby guiding the generation of molecular hypotheses. Further, by globally profiling structural in addition to protein abundance changes, LiP-MS has proven able to greatly increase the information content of functional proteomics screens. In sum, LiP-MS has supported the development of a novel conceptual framework for generating, visualizing, and interpreting structural proteomics data with peptide level resolution, thereby comprehensively probing biological systems. Here we survey the applications of LiP-MS, discuss methodological variants developed by us and others, and describe the use of this new type of omics readout for structural, functional, chemical, and biomarker discovery proteomics.