The molecular mechanism of the bicarbonate effect at the plastoquinone reductase site of photosynthesis

Abstract

It has been known for some time that bicarbonate reverses the inhibition, by formate under HCO3 (-)-depletion conditions, of electron transport in thylakoid membranes. It has been shown that the major effect is on the electron acceptor side of photosystem II, at the site of plastoquinone reduction. After presenting a historical introduction, and a minireview of the bicarbonate effect, we present a hypothesis on how HCO3 (-) functions in vivo as (a) a proton donor to the plastoquinone reductase site in the D1-D2 protein; and (b) a ligand to Fe(2+) in the QA-Fe-QB complex that keeps the D1-D2 proteins in their proper functional conformation. They key points of the hypothesis are: (1) HCO3 (-) forms a salt bridge between Fe(2+) and the D2 protein. The carboxyl group of HCO3 (-) is a bidentate ligand to Fe(2+), while the hydroxyl group H-bonds to a protein residue. (2) A second HCO3 (-) is involved in protonating a histidine near the QB site to stabilize the negative charge on QB. HCO3 (-) provides a rapidly available source of H(+) for this purpose. (3) After donation of a H(+), CO3 (2-) is replaced by another HCO3 (-). The high pKa of CO3 (2-) ensures rapid reprotonation from the bulk phase. (4) An intramembrane pool of HCO3 (-) is in equilibrium with a large number of low affinity sites. This pool is a H(+) buffering domain functionally connecting the external bulk phase with the quinones. The low affinity sites buffer the intrathylakoid [HCO3 (-)] against fluctuations in the intracellular CO2. (5) Low pH and high ionic strength are suggested to disrupt the HCO3 (-) salt bridge between Fe(2+) and D2. The resulting conformational change exposes the intramembrane HCO3 (-) pool and low affinity sites to the bulk phase.Two contrasting hypotheses for the action of formate are: (a) it functions to remove bicarbonate, and the low electron transport left in such samples is due to the left-over (or endogenous) bicarbonate in the system; or (b) bicarbonate is less of an inhibitor and so appears to relieve the inhibition by formate. Hypothesis (a) implies that HCO3 (-) is an essential requirement for electron transport through the plastoquinones (bound plastoquinones QA and QB and the plastoquinone pool) of photosystem II. Hypothesis (b) implies that HCO3 (-) does not play any significant role in vivo. Our conclusion is that hypothesis (a) is correct and HCO3 (-) is an essential requirement for electron transport on the electron acceptor side of PS II. This is based on several observations: (i) since HCO3 (-), not CO2, is the active species involved (Blubaugh and Govindjee 1986), the calculated concentration of this species (220 μM at pH 8, pH of the stroma) is much higher than the calculated dissociation constant (Kd) of 35-60 μM; thus, the likelihood of bound HCO3 (-) in ambient air is high; (ii) studies on HCO3 (-) effect in thylakoid samples with different chlorophyll concentrations suggest that the "left-over" (or "endogenous") electron flow in bicarbonate-depleted chloroplasts is due to "left-over" (or endogenous) HCO3...

Overall kinetics of photosystem II: pH dependence and deuterium isotope effect

“…This was not due to an altered requirement for bicarbonate, which is known to be necessary for electron transport through PS II, at least on the acceptor side [21], since the addition of 10 mM bicarb()nate did not fasten the kinetics (data not shown).…”

Bicarbonate effects on photo‐inhibition

“…At present it is not obvious how to explain this paradoxal role of bicarbonate in photo-inhibition. It may be related to the recent proposal by Blubaugh and Govindjee of two sites of high-affinity bicarbonate binding per reaction center [5]. While the presence of a bicarbonate ion in the first binding site (A) is suggested to be a pre-requisite for keeping the reaction center in a functional configuration, the second binding site (B) is suggested to be involved in the protonation of QB-.…”

“…They may be a new approach in investigations on the mechanism of photo-inhibition and the functioning of the reaction center of PS2. At this stage it is not clear how the bicarbonate effects on photo-inhibition presented here are related to the several observations of bicarbonate effects that have been made over the years (see [5]). For a thorough characterization of the bicarbonate effects on photoinhibition, we are currently conducting a set of experiments where the content of oxygen and bicarbonate is carefully measured and varied systematically under controlled conditions.…”

“…Instead it was found that the formate-treated thylakoid membranes that were depleted in endogenous bicarbonate prior to the pre-illumination (Fig. 2a, open symbols) were less susceptible to photo-inhibition than were the non-depleted control thylakoid membranes ( normal rates of electron transport [5,7]. Depletion of endogenous bicarbonate with formate ions inhibits electron transport and this inhibition is reversed upon readdition of NaHCO3.…”

“…Bicarbonate is well-known to bind to one or several sites on the thylakoid membrane (see [5]). Even when 20 mM NaHCO3 is not supplied to the thylakoid membrane suspension as in the experiments described above, the medium in itself still contains some bicarbonate, originating from dissolved carbon dioxide from the air and corresponding to about 0.130 mM NaHCO3 at pH 7.4.…”