Abstract
Photosystem II (PSII) is a large membrane-protein complex composed of about 20 subunits and various cofactors, which mediates the light-driven oxidation of water and reduction of plastoquinone, and is part of the photosynthetic electron transfer chain that is localized in the thylakoid membrane of cyanobacteria, algae, and plants. The stepwise assembly of PSII is guided and facilitated by numerous auxiliary proteins that play specific roles in this spatiotemporal process. Psb27, a small protein localized in the thylakoid lumen, appears to associate with an intermediate PSII complex that is involved in assembly of the Mn4CaO5 cluster. Its precise binding position on the PSII intermediate remains elusive, as previous approaches to the localization of Psb27 on PSII have yielded contradictory results. This was our motivation for a critical assessment of previously used methods and the development of an improved analysis pipeline. The combination of chemical cross-linking and mass spectrometry (CX-MS) with isotope-coded cross-linkers was refined and validated with reference to the PSII crystal structure. Psb27 was localized on the PSII surface adjacent to the large lumenal domain of CP43 on the basis of a cross-link connecting Psb27-K91 to CP43-K381. Additional contacts associating Psb27 with CP47 and the C-termini of D1 and D2 were detected by surface plasmon resonance (SPR) spectroscopy. This information was used to model the binding of Psb27 to the PSII surface in a region that is occupied by PsbV in the mature complex.
Highlights
Photosystem II (PSII) catalyzes the light-driven oxidation of water (McEvoy and Brudvig, 2006; Cox et al, 2013)
To check the ability of ab initio docking algorithms to predict the correct position of soluble subunits at the PSII surface, we made use of the CAPRI protocol for ‘Critical Assessment of PRedicted Interactions’ (Janin et al, 2003)
We have demonstrated a direct interaction of Psb27 with mature D1 C-terminus (mD1), which argues against the more distal binding position previously proposed for Psb27 on the basis of crosslinking data (Liu et al, 2011a), even if the whole CP43 domain were more flexible in the intermediate PSII complex lacking the adjacent extrinsic proteins
Summary
Photosystem II (PSII) catalyzes the light-driven oxidation of water (McEvoy and Brudvig, 2006; Cox et al, 2013). Cyanobacterial PSII has been successfully characterized by X-ray crystallography, and detailed structural models with a resolution down to 1.9 Å are Localization of Psb on PSII available (Ferreira et al, 2004; Guskov et al, 2009; Umena et al, 2011) These PSII structures reveal the spatial disposition of up to 20 protein subunits and multiple cofactors, such as chlorophylls, carotenoids, and lipids. Assembly of the different components during PSII biogenesis is an intricate process that is coordinated by a network of auxiliary proteins (Mulo et al, 2008; Nixon et al, 2010; Becker et al, 2011; Nickelsen and Rengstl, 2013) These accessory factors direct the spatiotemporal formation and distribution of precursor complexes in a coordinated assembly process, including a series of distinct, metastable assembly intermediates. Structural analysis of these intermediate PSII complexes is severely hampered by their low abundance and transient nature, with the consequence that none of these complexes has been successfully analyzed by X-ray crystallography so far
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