Plasma membrane purification from roots of sunflower by phase partitioning.
Plasma membrane vesicles were purified from the roots of sunflower (Helianthus annuus L. cv. Topflor) by aqueous polymer two-phase partitioning. The optimal conditions for separation were determined by systematic variation of the polymer concentration and salt composition. The phase system containing 6% (w/w) dextran T-500, 6% (w/w) polyethylene glycol 3350, 250 mM sucrose, 5 mM potassium phosphate, pH 7.8, without added salts proved to be the best. The ATPase activity had a pH optimum at 6.5 and it was stimulated by Mg2+, but not by Ca2+. The plasma membrane MgATPase activity was inhibited by vanadate but not by nitrate, an inhibitor of tonoplast ATPase. Only 10% of the microsomal protein was responsible for 36% of the total MgATPase activity. Moreover IDPase activity, a Golgi marker, appeared to be very low indicating the high purity of the preparation.
Szabó-Nagy
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Institute of Biophysics, Hungarian Academy of Sciences, Szeged.
Bérczi
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Erdei
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D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism; D002458:Cell Fractionation / Q000379:methods*; D002462:Cell Membrane / Q000201:enzymology; D003377:Countercurrent Distribution; D006368:Helianthus; D006863:Hydrogen-Ion Concentration; D007290:Inosine Diphosphate / Q000378:metabolism; D059828:Plant Cells; D014638:Vanadates / Q000494:pharmacology
D014638:Vanadates; D007290:Inosine Diphosphate; D017301:Ca(2+) Mg(2+)-ATPase