FIGURE 1.

MagStrep-mediated pulldown of proteoliposomes allows for an identification of the formation tripartite complexes. Lane 1 contains molecular mass markers (PageRuler Prestained Protein Ladder from Thermo Scientific, #26616). Detergent-purified MexA, MexB, OprM are loaded (2–5 μg protein per lane) onto a 10% SDS-PAGE Laemmli gel as reference (lanes 2, 3, and 4, respectively) and stained with Coomassie Blue. Proteoliposomes containing MexA and MexB (lane 5), MexA and MexB_D407N (lane 6), MexB alone (lane 7), or MexB in the presence of a soluble version of MexA (lane 8) were mixed with proteoliposomes containing OprM as described in the Material and Methods section. Liposomes were sedimented and the pellets were mixed with 5 μL loading buffer before being loaded on the gel. The samples were not boiled in order to avoid aggregation of the membrane proteins.