Conformational Dynamics of Light-Harvesting Complex II in a Native Membrane Environment

Abstract

Photosynthetic light-harvesting complexes (LHCs) of higher plants, moss, and green algae can undergo dynamic conformational transitions, which have been correlated to their ability to adapt to fluctuations in the light environment. Herein, we demonstrate the application of solid-state NMR spectroscopy on native, heterogeneous thylakoid membranes of Chlamydomonas reinhardtii (Cr) and on Cr light-harvesting complex II (LHCII) in thylakoid lipid bilayers to detect LHCII conformational dynamics in its native membrane environment. We show that membrane-reconstituted LHCII contains selective sites that undergo fast, large-amplitude motions, including the phytol tails of two chlorophylls. Protein plasticity is also observed in the N-terminal stromal loop and in protein fragments facing the lumen, involving sites that stabilize the xanthophyll-cycle carotenoid violaxanthin and the two luteins. The results report on the intrinsic flexibility of LHCII pigment-protein complexes in a membrane environment, revealing putative sites for conformational switching. In thylakoid membranes, fast dynamics of protein and pigment sites is significantly reduced, which suggests that in their native organelle membranes, LHCII complexes are locked in specific conformational states.

Protein–Protein Interactions Induce pH-Dependent and Zeaxanthin-Independent Photoprotection in the Plant Light-Harvesting Complex, LHCII

“…Critical to this remarkable feat is a network of pigment-containing proteins, termed the antenna, that captures and delivers solar energy to drive the biochemical reactions of water splitting and carbon fixation. , In green plants, the antenna simultaneously regulates the flux of energy to prevent photooxidative damage. Under high-light conditions, a reduction of pH on the luminal side of the thylakoid membrane results in a proton gradient that activates photoprotective mechanisms, which are known as nonphotochemical quenching (NPQ). , The reduced pH protonates the non-pigment-binding protein photosystem II subunit S (PsbS), which triggers NPQ through either interactions with the antenna proteins or changes to the properties of the membrane, − and activates the xanthophyll cycle, which then converts the carotenoid pigment violaxanthin (Vio) into zeaxanthin (Zea) in the antenna. − Clustering of the primary antenna protein, light-harvesting complex II (LHCII), into oligomers or arrays, within the thylakoid membrane, has also been observed during NPQ. − These changes in pH, carotenoid composition, and antenna organization are thought to change the conformation of individual antenna proteins and, as a result, the photophysics of the embedded pigments, enhancing pathways that dissipate excitation energy as heat. − While pH and carotenoid composition have been straightforward to manipulate, − systematic changes to the antenna organization cannot be independently introduced in vivo and generally require nonphysiological architectures in vitro. Due to these challenges in replicating the macro-organization of the antenna, the mechanism by which pH, carotenoid composition, and LHCII arrays collectively and/or individually activate dissipation has not been disentangled.…”

Protein structural dynamics by Magic-Angle Spinning NMR

“…An exciting possibility, opened by MAS NMR, is to investigate proteins within native membranes [56][57][58], where the lipid composition and orientation but also the presence of other proteins strongly affects their behavior [57,59]. An example of the impact of these factors on protein dynamics is the investigation of the Light-harvesting complex II (LHCII) in native thylakoid membrane [60]. The comparison of dynamics-edited correlation spectra of both LHCII reconstituted in lipid bilayers and in thylakoid membranes highlights key protein regions for which conformational equilibria are shifted by the lipidic environment.…”

Real‐Time NMR Recording of Fermentation and Lipid Metabolism Processes in Live Microalgae Cells

“…The carbonyl and carboxyl signals from lipids and protein backbone appear between 170-180 ppm. [24][25][26][27][28][29][30] A comparison of the initial DP spectrum and the DP spectrum after 24 hours shows the formation of metabolic end-and by-products over time as indicated in Figure 2. We identify signals that emerge in time of ethanol (17.4 ppm and 58.1 ppm), glycerol (63.2 ppm and 72.6 ppm) and aqueous CO 2 (125.1 ppm).…”

“…cells. [24][25][26][27][28][29][30] The signals of protein aromatic side-chains and double-bonded 13 C signals of the unsaturated lipid chains are visible in the region between 125-135 ppm. The carbonyl and carboxyl signals from lipids and protein backbone appear between 170-180 ppm.…”