Quantitative Assessment of Intrinsic Carbonic Anhydrase Activity and the Capacity for Bicarbonate Oxidation in Photosystem II

Abstract

On the basis of equilibrium isotopic distribution experiments using (18)O-labeled water, it is generally accepted that water is the sole substrate for O(2) production by photosystem II (PSII). Nevertheless, recent studies indicating a direct interaction between bicarbonate and the donor side of PSII have been used to hypothesize that bicarbonate may have been a physiologically important substrate for O(2) production during the evolution of PSII [Dismukes, G. C., Klimov, V. V., Baranov, S. V., Kozlov, Y. N., DasGupta, J., and Tyryshikin, A. (2001) Proc. Natl. Acad. Sci. U.S.A. 98, 2170-2175]. To test out this hypothesis and to determine whether contemporary oxygenic organisms have the capacity to oxidize bicarbonate, we employed special rapid-mixing isotopic experiments using (18)O/(13)C-labeled bicarbonate to quantify the inherent carbonic anhydrase activity in PSII samples and the potential flux of oxygen from bicarbonate into the photosynthetically produced O(2). The measurements were made on PSII samples prepared from spinach, Thermosynechococcus elongatus, and Arthrospira maxima. For the latter organism, a strain was used that grows naturally in an alkaline, high (bi)carbonate soda lake in Africa. The results reveal that bicarbonate is not the substrate for O(2) production in these contemporary oxygenic photoautotrophs when assayed under single turnover conditions.

Mobile hydrogen carbonate acts as proton acceptor in photosynthetic water oxidation

“…Despite numerous experiments this point remained unresolved (21,22,35,38). Although PSII membranes isolated from spinach also possess some carbonic anhydrase activity (6,8), there is presently no evidence for a carbonic anhydrase directly connected to higher-plant PSII, and there is also no need for a CCM function (discussed above). Our finding that under continuous illumination the activity of PSII is nevertheless dependent on the HCO − 3 concentration of the buffer therefore demonstrates that HCO − 3 directly affects the activity of PSII.…”

“…The possible roles of inorganic carbon, C i ðC i = CO 2 ; H 2 CO 3 ; HCO − 3 ; CO 2− 3 Þ, in this process have been a controversial issue ever since Otto Warburg and Günter Krippahl (5) reported in 1958 that oxygen evolution by PSII strictly depends on CO 2 and therefore has to be based on the photolysis of H 2 CO 3 ("Kohlensäure") and not of water. These first experiments were indirect and, as became apparent later, were wrongly interpreted (6)(7)(8). Several research groups followed up on these initial results and identified two possible sites of C i interaction within PSII (reviewed in refs.…”

Photoassembly of the water-oxidizing complex in photosystem II

“…Some alkalophilic cyanobacteria (Arthrospira genus) that originate from volcanic soda lakes prefer very high extracellular carbonate concentrations reaching 0.4 to 1.2 M NaCO 3 . These organisms lack carbonic anhydrases all together [66]} and possess a reversible bicarbonate site within the WOC that is required for O 2 evolution ( [67]). This site may be the same as the bicarbonate site associated with the Arg 357 residue of subunit CP43 which when mutated, gives rises to lowered O 2 evolution activity, hypothesized as due to a more energetic two-electron oxidation pathway that forms H 2 O 2 [68].…”

Focusing the view on nature's water-splitting catalyst

“…On the basis of EXAFS measurements on Br K substituted samples, it was recently suggested that Cl K is not a direct ligand of the Mn 4 cluster in the S 1 state (Haumann et al 2006). In contrast to Cl K , no sound evidence exists for HCO 3 K binding within the WOC, and it is also highly controversial what kind of role it may play during photosynthetic water oxidation (Klimov & Baranov 2001;Stemler 2002;Clausen et al 2005;Hillier et al 2006). EXAFS on solution samples does not allow extracting the orientations of the Mn-Mn and MnCa vectors.…”