Extended Data Fig. 1: Flow chart of preparation of native-MS ready proteo-liposomes.

Detail flow chart of two alternate approaches that can yield native-MS ready proteo-liposomes with desired lipid composition and membrane biophysical properties. The formed proteo-liposomes can be isolated from the free protein and the empty liposomes through a floatation assay. For better visualization, we typically add 1% RhodPE in our lipid mix. Post-floatation, the quality of the liposomes is assessed via negative stain imaging and DLS. An optional step can be introduced to control the diameter, and in turn the curvature, of the liposomes by passing them through an extruder fitted with filters of desired dimension. The size is further assessed by negative stain or DLS. Between the two protocols, while float-up method is typically longer, for certain protein-detergent scenario, dialysis can be more effective way to reconstitute. Figure graphics were created using Biorender.com.