Abstract
In photosynthetic organisms, photosystem II (PSII) is a large membrane protein complex, consisting of a pair of core complexes surrounded by an array of variable numbers of light-harvesting complex (LHC) II proteins. Previously reported structures of the PSII-LHCII supercomplex of the green alga Chlamydomonas reinhardtii exhibit significant structural heterogeneity, but recently improved purification methods employing ionic amphipol A8-35 have enhanced supercomplex stability, providing opportunities for determining a more intact structure. Herein, we present a 5.8 Å cryo-EM map of the C. reinhardtii PSII-LHCII supercomplex containing six LHCII trimers (C2S2M2L2). Utilizing a newly developed nonionic amphipol-based purification and stabilizing method, we purified the largest photosynthetic supercomplex to the highest percentage of the intact configuration reported to date. We found that the interprotein distances within the light-harvesting complex array in the green algal photosystem are larger than those previously observed in higher plants, indicating that the potential route of energy transfer in the PSII-LHCII supercomplex in green algae may be altered. Interestingly, we also observed an asymmetric PSII-LHCII supercomplex structure comprising C2S2M1L1 in the same sample. Moreover, we found a new density adjacent to the PSII core complex, attributable to a single-transmembrane helix. It was previously unreported in the cryo-EM maps of PSII-LHCII supercomplexes from land plants.
Highlights
In photosynthetic organisms, photosystem II (PSII) is a large membrane protein complex, consisting of a pair of core complexes surrounded by an array of variable numbers of light-harvesting complex (LHC) II proteins
We found that the interprotein distances within the light-harvesting complex array in the green algal photosystem are larger than those previously observed in higher plants, indicating that the potential route of energy transfer in the PSII–LHCII supercomplex in green algae may be altered
We developed a procedure for purifying the PSII– LHCII supercomplex of C. reinhardtii employing an ionic amphipol, A8-35 (11)
Summary
We developed a procedure for purifying the PSII– LHCII supercomplex of C. reinhardtii employing an ionic amphipol, A8-35 (11). Upon examination of the 2D classes of the C. reinhardtii C2S2M2L2 PSII–LHCII supercomplex, it was observed that there were two extra points of density (Fig. 4 and Fig. S6), commonly attributed to transmembrane helices and potentially interacting with PsbJ, PsbK, and PsbZ, that were not present in projection maps of previously reported C2S2 or C2S2M2 PSII– LHCII structures in land plants (3, 17, 18) (Fig. S6). The misalignments of the LHCII S-trimer were 15 Å for the C2S2-type PSII–LHCII from spinach, 6 Å for the C2S2M2-type PSII–LHCII from A. thaliana, and 14 Å for the C2S2M2-type PSII–LHCII from pea, respectively (Fig. 3, F–H, arrows) Another point of interest is that the C2S2M2L2-type supercomplex was highly flexible, its yield was high using NAPol stabilization. The rotations of 3.9° for the S-trimer, 18.9° for the M-trimer and of 26.6° for the L-trimer were observed in the 2D classification images, respectively (Fig. S9, arrows)
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