Mass spectroscopy locates the extrinsic proteins of photosystem II.

The photosynthetic oxidation of water is catalyzed by photosystem II (PSII), a multisubunit pigment-protein embedded in the thylakoid membranes of cyanobacteria, algae, and plants. This remarkable photoenzyme is capable of generating the highly oxidizing chemical species required to extract four tightly bound electrons of two substrate water-molecules, yielding biosynthetically useful reductant and by-product O2. PSII is a large homodimeric complex in vivo, with a combined mass of ∼700 kDa. Each PSII monomer comprises more than 20 different proteins collectively coordinating ∼60 cofactors, including 35 chlorophylls, 2 pheophytins, 2 plastoquinone molecules, and the Mn4Ca cluster, responsible for H2O-oxidation (1, 2). Despite much progress, including the 1.9 Å crystal structure of cyanobacterial PSII (1, 3), crucial structural information is missing. This lack includes extrinsic proteins affecting the Mn4Ca cluster, not observed in the PSII crystal structure because they are lost during the purification and crystallization process. In PNAS, Liu et al. use complementary technical approaches to construct a model for the binding of one of the lost extrinsic proteins, PsbQ, to the PSII, complex, and in doing so provide an example for the solution of the more general structural problem of defining the 3D structure of separately crystallized proteins that assemble into larger macromolecular complexes (4). Besides its prodigious catalytic capacities, the PSII complex provides a study in contrasts: its core exhibits an extremely high level of evolutionary conservation, considering the ∼2.7 billion y of evolutionary divergence, whereas the peripheral components display a dizzying degree of variation (Fig. 1). Although much of the peripheral variation is associated with the different light-harvesting complexes conferring adaptation to different light environments, there is also a great degree of diversity with respect to the extrinsic proteins associated with the water-splitting reaction. Generally, the extrinsic proteins modulate the Ca and Cl cofactor requirements of the H2O-oxidation reaction, stabilize the Mn4Ca cluster, and control accessibility to the strong oxidizing equivalents accumulated during the catalytic cycle (5–8). However, their precise role is not known, nor is the significance of the variations in the structure of this region of the PSII complex appreciated. It is not known, for example, whether the extrinsic proteins facilitate the crucial deprotonation of substrate H2O, although the proton exit pathway is modeled to include residues from PsbO (1, 9). The uncertainty is especially acute for the proteins designated PsbP and PsbQ, which have multiple homologs in and outside of PSII, including assembly factors for other membrane complexes (10, 11). Early studies revealed what appeared to be a fairly simple “one-plus-two” picture of the PSII extrinsic proteins: Three extrinsic proteins were associated with the H2O-oxidation complex, of which one protein, PsbO, is found in all species. The identity of the other two extrinsic proteins depends upon the species: PsbP and PsbQ are found in green algae and plants, while PsbV and PsbU are present in cyanobacteria, red algae, and evolutionary derivatives of red algae, including diatoms and brown algae. However, this one-plus-two picture is an oversimplification. Fig. 1. Photosystem II: A study in contrasts. The reaction center core of PSII coordinates the pigments and cofactors involved in converting photoexcitation energy to transmembrane charge separation and stabilization. This portion of the PSII complex is highly conserved despite ∼2.7 billion y of evolution. In contrast, cyanobacteria, higher plants, and the many genera of eukaryotic algae exhibit great diversity with respect to the light-harvesting antennae and the extrinsic protein domain that caps the highly reactive metal cluster, Mn4Ca, which is the metal center that accumulates the “electron holes” produced during photochemical charge separation. The electron holes are charge compensated by deprotonation, and constitute the powerful, multivalent oxidant, required to remove the hydrogens from substrate H2O molecules. Author contributions: R.L.B. wrote the paper.