The mechanism of proton pumping by cytochrome c oxidase

Cytochrome c oxidase catalyzes the reduction of oxygen to water that is accompanied by pumping of four protons across the mitochondrial or bacterial membrane. Triggered by the results of recent x-ray crystallographic analyses, published data concerning the coupling of individual electron transfer steps to proton pumping are reanalyzed: Conversion of the conventional oxoferryl intermediate F to the fully oxidized form O is connected to pumping of only one proton. Most likely one proton is already pumped during the double reduction of O, and only three protons during conversion of the “peroxy” forms P to O via the oxoferryl form F. Based on the available structural, spectroscopic, and mutagenesis data, a detailed mechanistic model, carefully considering electrostatic interactions, is presented. In this model, each of the four reductions of heme a during the catalytic cycle is coupled to the uptake of one proton via the D-pathway. These protons, but never more than two, are temporarily stored in the regions of the heme a and a3 propionates and are driven to the outside (“pumped”) by electrostatic repulsion from protons entering the active site during turnover. The first proton is pumped by uptake of one proton via the K-pathway during reduction, the second and third proton during the P → F transition when the D-pathway and the active site become directly connected, and the fourth one upon conversion of F to O. Atomic structures are assigned to each intermediate including F′ with an alternative route to O.

Rapid Regulation by Acid pH of Cell Wall Adjustment and Leaf Growth in Maize Plants Responding to Reversal of Water Stress1

The role of acid secretion in regulating short-term changes in growth rate and wall extensibility was investigated in emerging first leaves of intact, water-stressed maize (Zea mays L.) seedlings. A novel approach was used to measure leaf responses to injection of water or solutions containing potential regulators of growth. Both leaf elongation and wall extensibility, as measured with a whole-plant creep extensiometer, increased dramatically within minutes of injecting water, 0.5 mm phosphate, or strong (50 mm) buffer solutions with pH ≤ 5.0 into the cell-elongation zone of water-stressed leaves. In contrast, injecting buffer solutions at pH ≥ 5.5 inhibited these fast responses. Solutions containing 0.5 mm orthovanadate or erythrosin B to inhibit wall acidification by plasma membrane H+-ATPases were also inhibitory. Thus, cell wall extensibility and leaf growth in water-stressed plants remained inhibited, despite the increased availability of (injected) water when accompanying increases in acid-induced wall loosening were prevented. However, growth was stimulated when pH 4.5 buffers were included with the vanadate injections. These findings suggest that increasing the availability of water to expanding cells in water-stressed leaves signals rapid increases in outward proton pumping by plasma membrane H+-ATPases. Resultant increases in cell wall extensibility participate in the regulation of water uptake, cell expansion, and leaf growth.