The chromatograms and elution volumes of (A) n-PTEN and (B) 4p-PTEN purified by FPLC on a size exclusion column. (C) The chromatogram of protein standards on a size exclusion column (Peak A: Thyroglobulin, MW = 670,000, elution volume = 8.7 ml; Peak B: gamma globulin, MW = 158,000, elution volume = 9.7 ml; Peak C: Ovalbumin, MW = 44,000, elution volume = 14.6 ml; Peak D: Myoglobin, MW = 17,000, elution volume = 16.3 ml; Peak E: Vitamin B12, MW = 1350, elution volume = 19.5 ml). (D) Phosphatase assays using diC6 PIP3 for WT GST-PTEN and Y379C GST-PTEN purified from E. coli. Data are reported as the mean ± the SEM of three experiments performed in duplicate. (E) Generation of 4p-PTEN by expressed protein ligation in the presence or absence of 25 μM VO-OHpic, a potent PTEN inhibitor monitored by western blot. (F) 50 ng of 4p-PTEN treated with 2 μM n-PTEN (left) or 1 μM CIP (right). Dephosphorylation of the phospho-tail is monitored by western blot.