The plasma membrane ATPase from oat roots has been purified near homogeneity by a simple procedure. Plasma membranes isolated from sucrose gradients are first extracted with Triton X-100 and KC1 and the residue solubilized with lysolecithin. Rate-zonal centrifugation in a vertical rotor with a glycerol gradient results in a preparation of very high specific activity (6 mumoles min-1 mg protein-1 at 30 degrees C) and where over 70% of the protein corresponds to a polypeptide of about 100 kilodaltons previously identified as the ATPase. The purified enzyme could be reconstituted in proteoliposomes catalyzing ATP-driven proton transport sensitive to vanadate.