Functional dissection of the interactions of stonin 2 with the adaptor complex AP-2 and synaptotagmin -- Data Supplement - HTML Page - 07862Fig9Legend.htslp -- Proceedings of the National Academy of Sciences

Fig. 9.
(A) The ability of GST-WVxF single-point mutants to bind AP-2. Samples were separated by SDS/PAGE and immunoblotting for clathrin and AP-2. RBE, 10 or 100 m g of total rat brain extract taken as a standard. (B) Ponceau S staining of material affinity-purified from rat brain by using the N terminus of stonin 2 fused to GST in the presence of increasing concentrations of synthetic WVxF peptide or the inactive AVxA double-point mutant. (C) Affinity purification from rat brain homogenate by using the GST-fused NT domain of stonin 2 or a triple W to A mutant (D WWW). Samples were analyzed by SDS/PAGE and immunoblotting for Eps15, AP-2a , hsc70, and clathrin RBE, with 10 or 100 m g of total rat brain extract taken as a standard. (D) Binding of ³⁵S-labeled in vitro-translated AP-2a or purified m 2 (amino acids 157-435) to GST-WVxF peptide or an inactive mutant. Samples were separated by SDS/PAGE and analyzed by PhosphorImager analysis (Upper) or Coomassie blue staining (Lower).