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A catalytically active, vanadate-sensitive P-type ATPase with enzymatic, inhibitor and immunological properties similar to the plasmalemma enzyme is associated with the rough, but not smooth, endoplasmic reticulum obtained from tendril tissue of Bryonia dioicaJacq. using a density shift discontinuous two-step gradient purification procedure. The enzyme is present in this fraction in similar abundance as in the plasmalemma and constitutes approximately 32% of the total vanadate-sensitive ATPase activity of the microsomal fraction. ATP-dependent proton pumping activity was shown for this membrane using the fluorescent dye 9-amino-6-chloro-2-methoxyacridine. Proton potentials may be of relevance for rough endoplasmic reticulum functions, at least in tendrils.