The effect of <5 KDa (low molecular weight, LMW) and >5 KDa (high molecular weight, HMW) humic fractions on transport activities of isolated plasma membrane vesicles was studied. The K+-stimulated component of the ATP-hydrolyzing activity was considerably increased by LMW humic substances at concentrations ranging from 0.075 mg org CL-1 to 1 mg org CL-1. The stimulation was still evident when the detergent Brij-35 was added in the assay mixture, indicating a direct effect of LMW humic substances on plasma membrane ATPase activity. The LMW humic fraction stimulated ATP-dependent intravesicular H+-accumulation with a pattern similar to that recorded for ATP hydrolysis. LMW humic substances induced also an increase in passive membrane permeability to protons, as revealed by following the dissipation of an artificially imposed pH gradient. Membrane permeability to anions, as measured by the anion-dependent active proton accumulation was affected by LMW humic substances. In the presence of NO3- these molecules clearly enhanced proton transport, while Cl--dependent activity was almost unaffected, thus suggesting a specific action of LMW humic fraction on transmembrane NO3- fluxes. On the other hand, HMW humic substances decreased the passive permeability to protons and reduced the anion-dependent intravesicular H+-accumulation. The results suggest that the stimulatory effect of soil humic substances on plant nutrition and growth might be, at least in part, explained on the basis of both direct action of LMW humic molecules on plasma membrane H+-ATPase and specific modification of cell membrane permeability.
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