SHP-2 dephosphorylates coimmunoprecipitated band 3.

Erythrocytes were treated with diamide, and extracts from cell membranes were immunoprecipitated with anti–SHP-2 antibody. Immunocomplexes were washed twice in the absence of sodium orthovanadate and then incubated for 30 minutes at 30°C in phosphatase assay buffer containing 1 mM sodium orthovanadate (lane 1) or 4 mM dithiothreitol (lanes 2-4). The indicated amount of calpeptin was present in the phosphatase assay buffer of experiments of lanes 3 and 4. After incubation, samples were submitted to SDS-PAGE, transferred to nitrocellulose, and immunostained with anti–P-Y (A) or anti–band 3 (B) antibodies. Figure represents at least 3 separate experiments.