FIGURE 1:

Doc2b binds to the GLUT4 exocytic SNAREs. (A) Diagram of the liposome coflotation assay used to probe Doc2b-SNARE interactions. (B) Coomassie blue–stained SDS–PAGE gels showing the binding of Doc2b to protein-free or SNARE liposomes in the presence of 1 mM EGTA or CaCl₂. The liposomes were prepared with the PC lipid. Because Doc2b comigrates with syntaxin-4 on SDS–PAGE, a SUMO-tagged Doc2b protein, which exhibits the same biological activity as untagged Doc2b (Supplemental Figure S2), was used in the liposome flotation assays. Each binding reaction contained 5 μM v- or t-SNAREs. Soluble factors were added to a final concentration of 5 μM.