Properties of a plasmalemma ATPase of the maize scutellum

Abstract

Properties of a plasmalemma phosphatase of the maize scutellum, tentatively identified as an ATPase in a previous paper, were investigated. Fresh and frozen-thawed scutellum slices, that had been treated with 10 mM HCl to destroy acid phosphatases, were used as a source of enzyme. With the exceptions of the Na +, K + and dinitrophenol experiments, the two kinds of slices gave similar results. ATP and CTP were the best substrates for the enzyme followed by TTP, UTP, CDP, ADP and GTP. UDP, nucleoside monophosphates, sugar phosphates, inorganic pyrophosphate and p-nitrophenyl phosphate were relatively ineffective as substrates. The K m's for ATP and ADP were 0.65 and 5 mM, respectively, but the two substrates gave the same V max (49.8 μmol P i/hr/g slices). Previously, it was shown that the products of ATP hydrolysis are ADP, AMP and P i. Using these previous results and from the time courses of ATP disappearance from the bathing solution and the appearance of P i and ADP, it was concluded that ATP and ADP were hydrolysed by the same enzyme. The ATPase was not inhibited by oligomycin. N- N′-Dicyclohexylcarbodiimide (DCCD) was a poor inhibitor, and a water soluble analog of DCCD, 1-ethyl-3 (3 dimethyl-aminopropyl)-carbodiimide, gave only 33% inhibition. The relative effectiveness of divalent cations for stimulating ATPase activity was Mn 2+ > Mg 2+ ⩾ Ca 2+ > Co 2+ · Na + and K + gave a small additional stimulation in the presence of Mg 2+. However, Na + and K + gave a much greater stimulation when no divalent cation was added, and this occurred only when fresh slices were used. Dinitrophenol also increased ATPase activity only when fresh slices were used. Since it is likely that both the uptake of Na + and K + and the action of dinitrophenol would lower the electrochemical gradient of protons across the plasmalemma, the different results obtained with fresh slices indicate that the ATPase in these slices was under the constraint of a proton gradient.