A functional role for tyrosine-D in assembly of the inorganic core of the water oxidase complex of photosystem II and the kinetics of water oxidation

The role of D2-Tyr160 (Y-D), a photooxidizable residue in the D2 reaction center polypeptide of photosystem II (PSII), was investigated in both wild type and a mutant strain (D2-Tyr160Phe) in which phenylalanine replaces Y-D in the cyanobacterium Synechocystis sp. (strain PCC 6803). Y-D is the symmetry-related tyrosine that is homologous to the essential photoactive Tyr161(Y-Z) of the D1 polypeptide of PSII. We compared the flash-induced yield of O-2 in intact, functional PSII centers from both wild-type and mutant PSII core complexes. The yield of O-2 in the intact holo-enzyme was found to be identical in the mutant and wild-type PSII cores using long (saturating) pulses or continuous illumination, but was observed to be appreciably reduced in the mutant using short (nonsaturating) light pulses (<50 ms). We also compared the rates of the first two kinetically resolved steps of photoactivation. Photoactivation is the assembly process for binding of the inorganic cofactors to the apo-water oxidation/PSII complex (apo-WOC-PSII) and their light-induced photooxidation to form the functional Mn4Ca1Clx core required for 02 evolution. We show that the D2-Tyr160Phe mutant cores can assemble a functional WOC from the free inorganic cofactors, but at a much slower rate and with reduced quantum efficiency vs wild-type PSII cores. Both of these observations imply that the presence Of Y-D(.) leads to a more efficient photooxidation of the Mn cluster relative to deactivation (reductive processes). One possible explanation for this behavior is that the phenolic proton on Y-D is retained within the reaction center following Y-D oxidation. The positive charge, likely shared by D2-His189 and other residues, raises the reduction potential of P680(+)/P-680, thereby increasing the driving force for the oxidation of Mn4YZ. There is, therefore, a competitive advantage to organisms that retain the Y-D residue, possibly explaining its retention in all sequences of psbD (encoding the D2 polypeptide) known to date. We also find that the sequence of metal binding steps during assembly of apo-WOC-PSII centers in cyanobacteria cores differs from that in higher plants. This is seen by a reduced calcium affinity at its effector site and reduced competition for binding to the Mn(II) site, resulting in acceleration of the initial lagtime by Ca2+, in contrast to retardation in spinach. Ca2+ binding to its effector site promotes the stability of the photointermediates (IM1 and above) by suppressing unproductive decay.