Recombinant GST-SH2 domain counteracts the interaction between Lyn and phosphorylated band 3.

(A) Immunostaining of band 3 coimmunoprecipitated in anti-Lyn IP. Erythrocytes were treated with pervanadate, and the proteins extracted from the cell membranes were immunoprecipitated with anti-Lyn antibody as described in “Materials and methods.” The immunoprecipitations shown in lanes 2 to 5 were performed in the presence of increasing concentrations of recombinant GST/SH2 domain. (B) Immunostaining of band 3 coimmunoprecipitated in anti-GST IP. Human erythrocytes were treated as in Figure 2B, and the proteins extracted from the cell membranes were immunoprecipitated with anti-GST antibody in the presence of 4-μmol/L recombinant GST-SH2 domain. Immune complexes were submitted to SDS-PAGE, transferred to nitrocellulose membranes, and immunostained with anti-band 3 antibody. Experimental conditions are detailed in “Materials and methods.” The figure is representative of 5 separate experiments.