Fig. 1. Purification of DPM synthase complex. GST- and FLAG-tagged hDPM1 was isolated by two-step affinity purification using anti-FLAG and glutathione beads from the digitonin lysates of 3 × 109 JY25 transfectant cells. A part of the purified protein was analysed by SDS–PAGE and silver staining (A) and the rest was used for DPM synthase assay as shown in (B). (A) M, molecular weight markers. The protein indicated by the asterisk is a contaminant. (B) Liposomes only (lane 1), DPM synthase complexes reconstituted in liposomes (lane 2) and membranes of JY25 cells (lane 3) were assayed for DPM synthase activity. The lipids were separated by TLC with a solvent system of chloroform/methanol/H2O (10:10:3).