Mass Photometry of Membrane Proteins

Here, we explore the application of mass photometry (MP) to integral membrane proteins and membrane mimetic systems at the single particle level. We apply MP to amphipathic vehicles, such as detergents and amphipols, as well as to lipid and native nanodiscs, characterizing the particle size, sample purity and heterogeneity. Using methods established for cryogenic electron microscopy, we eliminate detergent background, enabling high-resolution studies of membrane protein structure and interactions. We find evidence that, when extracted from native membranes using native styrene-maleic acid nanodiscs, the potassium channel KcsA is present as a dimer of tetramers – in contrast to results obtained using detergent purification. Finally, using lipid nanodiscs, we show that MP can help distinguish between functional and non-functional nanodisc assemblies, as well as determine the critical factors for lipid nanodisc formation. The dataset was acquired using custom-made mass photometer and commercial mass photometer OneMP, and the data was recorded either using custom-written LabView code or using commercial version, AcquireMP, respectively. The data was then processed using DiscoverMP and plotted using Matlab. To read the files, please analyse the data using DiscoverMP to generate .h5 results files as well as a .csv data set with particle contrast values. Further information on data processing and acquiring can be found here: https://www.biorxiv.org/content/10.1101/2020.02.28.969287v