Photosynthesis of leaf cell protoplasts and permeability of the plasmalemma to some solutes.

Photosynthesis was measured in mesophyll protoplasts isolated from spinach leaves. Under high intensity illumination and in the presence of 21% O2, half-saturation of photosynthesis by CO2 required CO2 concentrations between 8 and 12 μm at different pH values of the suspending medium. Concentrations of HCO 3 (-) needed for half-saturation increased correspondingly with the pH of the media. The pH profile of protoplast photosynthesis was much broader than that of CO2 assimilation by isolated chloroplasts. The data indicate that leaf cells possess mechanisms to maintain considerable differences between external and internal pH over prolonged periods of time. Protoplast photosynthesis was inhibited by nitrite, acetate and bicarbonate; inhibition was more pronounced at low than at high pH and was attributed to stroma acidification. Nitrite was reduced in the light by protoplasts and chloroplasts. At pH 7.6, the apparent Km NO 2 (-) was about 0.6 mM for chloroplasts and 25 mM for protoplasts. Approximate permeability coefficients for NO 2 (-) and HNO2 were calculated from nitrite-dependent oxygen evolution at low nitrite concentrations, known nitrite or HNO2 gradients, data on the surface area of protoplasts and chloroplasts and the pH profile of nitrite inhibition of photosynthesis. The membrane potential was assumed to be-100 mV. For the chloroplast envelope, permeability coefficients were 1.5·10(-3) ms(-1) (HNO2) and 2·10(-8) ms(-1) (NO 2 (-) ) and for the plasmalemma 4·10(-5) ms(-1) (HNO2) and 5·10(-10) ms(-1) (NO 2 (-) ). The values calculated for anion penetration probably represent upper limits of permeability. The protoplasts appeared to be largely impermeable to phosphate and phosphate esters. A rapid metabolic response of cells or cellular strands to added anionic substrates such as phosphate esters as reported in the literature appears to be possible only in damaged cells. It requires the presence of open channels between the cytosol and external medium.