Inside‐out plant plasma membrane vesicles of high purity obtained by aqueous two‐phase partitioning

Highly purified plasma membranes obtained from leaves of sugar beet ( Beta vulgaris L.) by aqueous two‐phase partitioning were separated into two fractions by further phase partition steps. The vesicles partitioning to the interface showed an ATP‐dependent H + ‐uptake (measured using the pH probe acridine orange) and a negligible K + ,Mg 2+ ‐ATPase latency, while the vesicles partitioning in the upper phase showed only slow H + ‐uptake and a high ATPase latency on addition of Triton X‐100. Based on these results the material at the interface is estimated to contain ∼90% sealed, inside‐out vesicles, and the material in the upper phase ∼90% sealed, right‐side‐out vesicles