Extended Data Fig. 3: Purification and characterization of the supercomplex samples as well as evaluation of the cryo-EM maps of stacked-C2S2 and C2S2M2L2 supercomplexes.
From: Structural insight into light harvesting for photosystem II in green algae
a, Preparation of C2S2M2L2-type PSII-LHCII supercomplex through the sucrose density gradient (SDG) ultracentrifugation and amphipol method. Three major bands were identified as LHCII trimers, PSI-LHCI supercomplexes, and C2S2M2L2 PSII-LHCII supercomplexes. b, SDS-PAGE analysis of C2S2M2L2- type PSII-LHCII supercomplex (1 μg Chl/mL) stained with Coomassie Brilliant Blue R-250. M, molecular weight marker; Type I, LHCII type I (LhcbM3/4/6/8/9); Type III, LHCII type III (LhcbM2/7); Type IV, LHCII type IV (LhcbM1). Protein labels in parentheses represent co-migrated proteins. The experiment was repeated five times independently with similar results. c, Preparation of C2S2-type PSII-LHCII supercomplex through the SDG ultracentrifugation method. The components in four major bands of SDG tube (B1-B4) are labeled nearby the bands. d, SDS-PAGE analysis of the B4 fraction used for Cryo-EM data collection. The experiment was repeated five times independently with similar results. The identities of proteins labeled nearby the bands were revealed through mass spectroscopy analysis on the in-gel digestion products of the proteins. e, The Fourier Shell Correlation (FSC) curves for the maps and structural models of stacked-C2S2(left) and C2S2M2L2 (right). Black, the gold standard FSC curves between two independent half maps. Red, the FSC curve calculated between the maps and structural models. The dotted lines indicate the criterion at a FSC value of 0.143. f, Local resolution estimation of the maps of stacked-C2S2 and C2S2M2L2supercomplexes. The rainbow bar at the bottom indicate the color codes for regions with resolutions between 2.4 (blue) − 5.6 (red) Å.
