Unraveling design principles of protein landscapes in photosynthetic membranes in plant chloroplasts.

The supramolecular organization of proteins within photosynthetic membranes is crucial for energy conversion in plants. Here, we introduce an analytical and computational pipeline that integrates high-resolution cryo-scanning electron microscopy, biochemical quantification, advanced Monte Carlo computer simulations, and statistical methods to elucidate the elusive protein landscapes of grana membranes in intact Arabidopsis leaves. Our integrated analysis challenges the prevailing view that particles on the exoplasmic fracture faces in freeze-fracture samples represent photosystem II exclusively. Instead, these particles also include cytochrome b6f complexes. Furthermore, our steric clash analysis demonstrates that stacked membranes contain a mixture of larger PSII supercomplexes (C2S2M2 and C2S2) in addition to a smaller complex (C2). This suggests that in vivo PSII supercomplexes exist in an equilibrium distribution of differing sizes. Furthermore, we discovered that, although size exclusion effects govern the global protein arrangement, local packing exhibits orientational order indicative of lateral attractive protein-protein interactions.