Keeping It Native: Non-Denaturing Approaches to Top-Down Protein Mass Spectrometry

Top-down proteomics and top-down mass spectrometry approaches provide direct characterization of proteoforms; the specific molecular forms of proteins, including post-translational modifications, truncations, alternative splicing, and sequence variants. To date, most top-down studies have been performed under denaturing conditions, losing critical information about protein folding and noncovalent protein-protein and protein-ligand interactions. However, recent efforts to apply non-denaturing, or native, ionization to top-down mass spectrometry and top-down proteomics have shown substantial promise for characterizing these higher-order and more transient interactions. This review covers progress over the last 35 years in non-denaturing top-down approaches and addresses the inherent strengths and challenges of the technique. We cover three major areas of applications: (1) Ligand binding and drug discovery, (2) Structural biology, and (3) Proteomics aspects and bioinformatics.