Studies on 17,24 kD Depleted Photosystem II Membranes

Abstract

Analyses were made of the effects of extraction of the 17,24 kilodalton extrinsic proteins from spinach versus wheat photosystem II (PSII) membranes on Ca abundance and O₂ evolution capacity determined in the absence and presence of either Cl⁻ or Ca²⁺. Extraction of these proteins from spinach PSII routinely diminished steady state O₂ evolution by about 70% when assayed in the presence of sufficient Cl⁻. Additionally, O₂ evolution of 17,24 kilodalton-less spinach PSII membranes showed about 2-fold more enhancement by Ca²⁺ than by Cl⁻ during assay. When the same extraction and assay procedures were applied to wheat PSII membranes, we observed, in contrast to 17,24 kilodalton-less spinach PSII, only about 50% inhibition of O₂ evolution and about 2-fold greater enhancement by Cl⁻ than by Ca²⁺. Irrespective of differences in the magnitude of enhancement of O₂ evolution by Ca²⁺versus Cl⁻ in spinach versus wheat, the K_m values for Cl⁻ (about 1.7 millimolar) and Ca²⁺ (about 1.5 millimolar) were similar for both type preparations. The abundance of Ca specifically associated with fully functional PSII (about 2 and about 3 Ca/200 chlorophyll for spinach and wheat, respectively) was diminished to about 1 per 200 chlorophyll upon 17.24 kilodalton protein depletion. Further treatment of wheat 17,24 kilodalton-less PSII in darkness with 2 molar NaCl/1 millimolar ethyleneglycol-bis(β-aminoethyl ether)-N,N′-tetraacetic acid/20 micromolar A23187² made O₂ evolution highly dependent on Ca²⁺ addition, much like the 17,24 kilodalton-less spinach PSII. Analyses of this Ca²⁺ effect on O₂ evolution revealed both high (K_m about 65 micromolar) and low (K_m about 1.5 millimolar) affinity Ca²⁺ sites in wheat 17,24 kilodalton-less PSII. The results suggest that during 17,24 kilodalton extraction by NaCl, spinach PSII is more susceptible than wheat PSII to loss of high affinity Ca and irreversible inhibition of O₂ evolution.