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>Characterization, Functional Reconstitution and Activation by Fusicoccin of a Ca2+-ATPase fromCorydalis sempervirensPers. Cell Suspension Cultures
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Characterization, Functional Reconstitution and Activation by Fusicoccin of a Ca2+-ATPase fromCorydalis sempervirensPers. Cell Suspension Cultures
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机译:Characterization, Functional Reconstitution and Activation by Fusicoccin of a Ca2+-ATPase fromCorydalis sempervirensPers. Cell Suspension Cultures
Cell suspension cultures ofCorydalis sempervirenshave proven ideal for the study of fusicoccin action Schulz et al. (1990)Planta183: 83 and express the fusicoccin-binding protein as well as a plasma membrane H+-ATPase which is activated by the fungal toxin. Microsomal vesicles prepared from these cells accumulate Ca2+in the presence of Mg-ATP. The protonophore car-bonylcyanidem-chlorophenylhydrazone did not inhibit the Mg-ATP dependent Ca2+-transport into the vesicles. This process is thus due to the activity of at least one primary active, ATP-driven, Ca2+-pump. The enzyme was characterized in detail. It has a pH optimum of 7.2, an apparentKmof 0.3 mu (ATP), 12pm (Ca2+), accepts ATP>ITP∼GTP>CTP∼UTP, and is strongly (Ki, app 0.75μmM) inhibited by erythrosine B but less so (Ki, app 95μM) by or-thovanadate. These characteristics are typical for the plasma membrane Ca2+-ATPase characterized from differentiated tissues Graf and Weiler (1990)Physiol. Plant.75: 634. Fusicoccin activates the erythrosine-sensitive Ca2+-pump by lowering itsKmfor ATP, when added to living cells prior to tissue homogenization. Thus, fusicoccin appears to activate at least two ion-translocating ATPases in one and the same tissue, suggesting that the toxin's mechanism of action is complex and not restricted to activation of the H+-ATPase. FC has no effect when administered to microsomes. The microsomal enzyme was solubilized and reconstituted into asolec-tin liposomes in functional form. The reconstituted, erythrosine sensitive Ca2+-ATPase was insensitive to fusicoccin. Thus, components essential for toxin action are either lost or inactivated during subcellular fractionation. It is likely that FC action requires soluble compo
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