Functional asymmetry of photosystem II D1 and D2 peripheral chlorophyll mutants of Chlamydomonas reinhardtii

Abstract

The peripheral accessory chlorophylls (Chls) of the photosystem II (PSII) reaction center (RC) are coordinated by a pair of symmetry-related histidine residues (D1-H118 and D2-H117). These Chls participate in energy transfer from the proximal antennae complexes (CP43 and CP47) to the RC core chromophores. In addition, one or both of the peripheral Chls are redox-active and participate in a low-quantum-yield electron transfer cycle around PSII. We demonstrate that conservative mutations of the D2-H117 residue result in decreased Chl fluorescence quenching efficiency attributed to reduced accumulation of the peripheral accessory Chl cation, Chl(Z)(+). In contrast, identical symmetry-related mutations at residue D1-H118 had no effect on Chl fluorescence yield or quenching kinetics. Mutagenesis of the D2-H117 residue also altered the line width of the Chl(Z)(+) EPR signal, but the line shape of the D1-H118Q mutant remained unchanged. The D1-H118 and D2-H117 mutations also altered energy transfer properties in PSII RCs. Unlike wild type or the D1-H118Q mutant, D2-H117N RCs exhibited a reduced CD doublet in the red region of Chl absorbance band, indicative of reduced energetic coupling between P680 and the peripheral accessory Chl. In addition, transient absorption measurements of D2-H117N RCs, excited on the blue side of the Chl absorbance band, exhibited a ( approximately 400 fs) pheophytin Q(X) band bleach lifetime component not seen in wild-type or D1-H118Q RCs. The origin of this component may be related to delayed fast-energy equilibration of the excited state between the core pigments of this mutant.

Figure 1

Chl fluorescence emission spectra (77 K) of dark-adapted (solid line) and illuminated (15 min at 200 K, dotted line) from WT (A), D1-H118Q (B), D2-H117N (C), and D2-H117Q (D) PSII membranes.

Figure 2

Time course of Chl fluorescence (695 nm) quenching of WT (solid line), D1-H118Q (dashed line), D2-H117N (dotted line), and D2-H117Q (dashed, dotted line) PSII membranes during illumination at 200 K.

Figure 3

EPR spectra of Chl generated in PSII RC preparations from WT (solid line), D1-H118Q (dashed line), and D2-H117N (dotted line). Instrument conditions: measuring temperature, 4.7 K; microwave frequency, 9.48 GHz; modulation amplitude, 0.2 mT; power, 10 μW; and modulation frequency, 100 kHz.

Figure 4

Chl CD spectra of PSII RC preparations from WT (solid line), D1-H118Q (dashed line), and D2-H117N (dotted line) mutants at 4°C.

Figure 5

Transient Pheo absorption kinetics at 542 nm in WT and peripheral Chl mutant RCs pumped at either 687 nm (A) or 655 nm (B). (Insets) Early time behavior.