Bicarbonate accelerates assembly of the inorganic core of the water-oxidizing complex in manganese-depleted photosystem II: A proposed biogeochemical role for atmospheric carbon dioxide in oxygenic photosynthesis

The proposed role for bicarbonate (HCO3-) as an intrinsic cofactor within the water-oxidizing complex (WOC) of photosystem II (PSII) [Klimov et al. (1997) Biochemistry 36, 16277-16281] was tested by investigation of its influence on the kinetics and yield of photoactivation, the light-induced assembly of the functional inorganic core (Mn4OyCa1Clx) starting from the cofactor-depleted apo-WOC-PSII center and free Mn2+, Ca2+, and Cl-. Two binding sites for bicarbonate were found that stimulate photoactivation by accelerating the information and suppressing the decay, respectively, of the first light-induced assembly intermediate, IM1 [apo-WOC-Mn(OH)(2)(+)]. A high-affinity bicarbonate site (K-D less than or equal to 10 mu M) stimulates both the rate of recovery of O-2 evolving centers and decreases (by a factor of 1.2-3) t(lag), the time for formation of IM1. This stimulation involves enhanced binding of the initial Mn2+ and occurs only at concentrations of Mn2+ at or below the stoichiometric requirements for water oxidation (less than or equal to 4 Mn/PSII) and disappears above 4 Mn/PSII. The absence of an effect from added bicarbonate on photoactivation kinetics and yield at saturating concentrations of Mn2+ and Ca2+ may be due to the availability of atmospheric bicarbonate dissolve in the buffers (similar to 4 mu M at pH 6.0) sufficient for photoactivation. The second bicarbonate site also stimulates the rate of formation of IM1 but has much lower affinity (K-D approximately millimolar) and becomes observable only at low concentrations of Ca2+ that are limiting for photoactivation. This stimulation effect appears to occur by complexation of free Ca2+, thereby reducing its activity in competing with Mn2+ in the formation of IM1. Bicarbonate had no effect on the calcium effector site responsible for the rate-limiting dark step of photoactivation (Ca2+ binding to IM1). Four interpretations of the high-affinity bicarbonate effect may be advanced as testable hypothesis: bicarbonate may (1) act as an integral cofactor within the WOC (possible ligand to the first Mn), (2) act as a Bronsted base to accelerate proton release during formation of either the dark precursor [apo-WOC-Mn(OH)(+)] or IM1 [apo-WOC-Mn(OH)(2)(+)], (3) directly deliver one or more hydroxide ions during formation of the latter two species (with release of CO2), or (4) act as a membrane-soluble anion that electrostatically elevates the local concentration of Mn2+ in PSII. These results support a possible biogeochemical role for bicarbonate in the evolution of the first oxygenic photosynthetic organism. An improvement in the illumination method for photoactivation is presented in which light flashes of increasing during are used to extend the pre-steady-state lag phase and to suppress photoinhibition, thereby improving the accuracy of t(lag) determination.