Characterization of ATP-Hydrolyzing and ATP-Driven proton-translocating activities associated with the peribacteroid membrane from root nodules of Lupinus luteusL

Autor: Andreev, Igor, Dubrovo, Pavel, Krylova, Valeria, Andreeva, Irina N., Koren'kov, Victor, Sorokin, Evgeny M., Izmailov, Stanislav F.
Zdroj: Journal of Plant Physiology; January 1997, Vol. 151 Issue: 5 p563-569, 7p
Abstrakt: In order to detect and characterize PBM H+-ATPase from root nodules of Lupinus luteusL., ATP-hydrolyzing and ATP-dependent proton translocating activities associated with this membrane were studied using isolated symbiosomes, vesicle PBM preparations and intact plant tissues. It has been found that ATP-hydrolyzing activity of isolated PBMs is sufficiently high (about 100–150 μmoles of μmg protein-h) in the selected pH range (4.5–8.5) and is characterized by the absence of any pronounced pH optimum, little substrate specificity, the absence of selectivity in relation to Mg2+and Ca2+as stimulators of ATP-hydrolysis and even more sensitivity to Ca2+. Mg2+-dependent ATP-hydrolyzing activity was moderately decreased in the presence of some known inhibitors of H+-ATPases, such as vanadate, DCCD and nitrate. On the other hand, Ca2+-dependent ATP-hydrolysis was significandy inhibited by sodium fluoride. ATP-hydrolyzing activity on the PBM of Lupinus luteusL. with many similar properties was also detected cytochemically in electron microscope studies of intact plant tissues. These findings led us to conclude that at least one more Ca2+/Mg2+-dependent ATP-hydrolase, in addition to H+-ATPase, is associated with the PBM. This enzyme is likely responsible for the observed background that masks ATP-hydrolyzing activity of the PBM H+-ATPase. ATP-dependent electrogenic proton transport across the PBM was detected in isolated symbiosomes and vesicle PBM preparations using oxonol YI and acridine orange as indicators of ΔΨ and ΔpH, respectively. Transport activity of the PBM H+-ATPase was blocked by the above mentioned ATPase inhibitors, except nitrate, which acted only as permeant anion. These results indicate that the PBM of Lupinus luteusL. contains only one H+-translocating ATPase, which belongs to a P-type H+-ATPase.
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