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Plasma membrane was isolated from mature green bell pepper fruit by two-phase partitioning or by sucrose density gradient. The yield of plasma membrane was higher from the sucrose density gradient, but the two-phase system was less contaminated by other membranes, particularly those from chloroplasts and mitochondria. In the two-phase partitioned membranes, ATPase activity was stimulated by Triton X-100 by 100% and in sucrose density gradient membranes by 40%. Plasma membranes from two-phase partitioning exhibited simultaneous proton pumping and ATP hydrolysis, while the sucrose density purified membranes did not. Immunoblotting with ATPase antibody showed enrichment of plasma membrane ATPase in both the U3 phase of the two-phase system, and the 34% sucrose fraction of the sucrose gradient. However, the two-phase partitioned membranes were superior to those prepared by sucrose density for investigating functions of the ATPase.

Affiliations: 1: Department of Postharvest Science, Volcani Center, Agricultural Research Organization


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