Factors associated with the instability of nitrate-insensitive proton transport by maize root microsomes

Proton transport catalyzed by the nitrate-insensitive, vanadate-sensitive H⁺-ATPase in microsomes from maize (Zea mays L.) roots washed with 0.25 molar KI decreased as a function of time at 0 to 4°C. The rate of proton transport was approximately one-half of that by freshly isolated microsomes after 6 to 18 hours of cold storage. The decrease in proton transport coincided with losses in membrane phosphatidylcholine and was not associated with a change in vanadate-sensitive ATP hydrolysis. A technique based on a protocol developed for the reconstitution of Neurospora crassa plasma membrane H⁺-ATPase (DS Perlin, K Kasamo, RJ Brooker, CW Slayman 1984 J Biol Chem 259: 7884-7892) was employed to restore proton transport activity to maize microsomes. These results indicated that the decline in proton transport by maize root membranes during cold storage was not due to degradation of the protein moiety of the H⁺-ATPase, but was due to the loss of phospholipids.