Fig. 1. Phosphatase activity of PTPα from unsynchronized and mitotic cells. Endogenous mouse PTPα (Endog) and overexpressed human PTPα (WT), PTPα-HA (WT-HA), PTPα(Y789F) (Y789F) and PTPα(Y789F)-HA (Y789F-HA) were immunoprecipitated with anti-PTPα(D2) antibody from lysates from non-overexpressor or overexpressor NIH 3T3-derived cells that were either unsynchronized (U) or arrested in mitosis (M). (The lysates were adjusted to contain approximately equal amounts of PTPα.) Aliquots of immunoprecipitates were incubated with [³²P]pTyr-containing MBP in phosphatase buffer for 15 min at 30°C or subjected to anti-pTyr or-PTPα immunoblots. (A) Amount of ³²P released per molecule of PTPα after 15 min incubation, normalized to the amount released by overexpressed PTPα from unsynchronized cells. Error bars indicate the SEM. (B) Anti-pTyr immunoblot of the immunoprecipitated PTPα. (C) Anti-PTPα immunoblot of the immunoprecipitated PTPα. SDS–PAGE was in 10% gels. The positions of molecular weight standards are indicated in kDa.