FPLC traces, western blots of caveolin purifications, and negative stain EM averages of caveolin complexes. (A–D) Indicated caveolin proteins were purified from E. coli membranes and applied to a Superose 6 10/300 Gl column. Elution profiles and western blotting results are shown for (A) human Cav1 (Type II-CAV, Q03135), (B) S. purpuratus caveolin (Type I-CAV, A0A7M7T4C2), (C) A. queenslandica caveolin (Atypical-CAV, A0A1X7UHP5), and (D) S. rosetta caveolin (Choa-CAV, F2U793). The position of the void and shoulders corresponding to various peaks (P1–P4) is indicated on each FPLC trace. Arrows on the western blots point to the expected position for monomers for each of the caveolins based on their predicted molecular weight. (E–H) Negative stain 2D class averages of human Cav1 (E), S. purpuratus caveolin (F), A. queenslandica caveolin (G), and S. rosetta caveolin (H). Classes denoted with # are shown in Fig. 6. The number of particles found in each class average is shown in the bottom left. Scale bar, 30 nm. The classes of smaller particles represent a membrane chaperone complex that is a structured protein contaminant in the purifications. For the case of A. queenslandica caveolin, the majority of 2D classes consist of these contaminant proteins. Consequently, only one class is marked as the caveolin complex. FPLC, fast protein liquid chromatography. Source data are available for this figure: SourceData FS4.