Abstract

The cellular processes underpinning life are orchestrated by proteins and the interactions they make with themselves and other biomolecules. A range of techniques has been developed to characterise these associations, operating from the ensemble all the way to the single molecule level. Structural and dynamic heterogeneity, however, continues to pose a fundamental challenge to existing analytical and structural methodologies, despite being key to protein and drug function. I will present recent developments demonstrating that interferometric scattering mass spectrometry (iSCAMS) can mass-image single biomolecules in solution with simultaneous nanometre precision and mass accuracy comparable to native mass spectrometry in the gas phase. As a result, we can resolve oligomeric distributions at high dynamic range, detect small-molecule binding, and quantitatively mass-image not only biomolecules composed of amino acids, but also heterogeneous species such as glyco- and lipoproteins. These capabilities enable us to determine the equilibrium and rate constants and thereby the molecular mechanisms of processes as diverse as homo- and hetero-oligomeric protein assembly, amyloidogenic protein aggregation and actin polymerisation. Our results illustrate how single molecule mass imaging provides universally-applicable and spatially-resolved access to the mechanisms and dynamics of protein assemblies, their interactions and how they form nano- and mesoscopic structures, one molecule at a time.