Insights into the photoprotective switch of the major light-harvesting complex II (LHCII): a preserved core of arginine-glutamate interlocked helices complemented by adjustable loops

Abstract

Light-harvesting antennae of the LHC family form transmembrane three-helix bundles of which two helices are interlocked by conserved arginine-glutamate (Arg-Glu) ion pairs that form ligation sites for chlorophylls. The antenna proteins of photosystem II have an intriguing dual function. In excess light, they can switch their conformation from a light-harvesting into a photoprotective state, in which the excess and harmful excitation energies are safely dissipated as heat. Here we applied magic angle spinning NMR and selective Arg isotope enrichment as a noninvasive method to analyze the Arg structures of the major light-harvesting complex II (LHCII). The conformations of the Arg residues that interlock helix A and B appear to be preserved in the light-harvesting and photoprotective state. Several Arg residues have very downfield-shifted proton NMR responses, indicating that they stabilize the complex by strong hydrogen bonds. For the Arg Cα chemical shifts, differences are observed between LHCII in the active, light-harvesting and in the photoprotective, quenched state. These differences are attributed to a conformational change of the Arg residue in the stromal loop region. We conclude that the interlocked helices of LHCII form a rigid core. Consequently, the LHCII conformational switch does not involve changes in A/B helix tilting but likely involves rearrangements of the loops and helical segments close to the stromal and lumenal ends.

Keywords: Bioenergetics; LHC Family; Non-photochemical Quenching; Photosynthesis; Photosynthetic Light Harvesting; Photosystem II; Protein Conformation; Solid-state NMR.

FIGURE 1.

Decay-associated fluorescence spectra and associated lifetimes of LHCII concentrated in β-DM buffer solution.

FIGURE 2.

77 K fluorescence spectra of LHCII. Dashed spectrum, LHCII in β-DM buffer. Solid spectrum, LHCII in detergent-free buffer after extensive dialysis for removal of the β-DM.

FIGURE 3.

¹³C CP-MAS spectra of unquenched (dashed line) and quenched (solid line) LHCII. A, detergent peaks in unquenched LHCII are denoted with asterisks. The chemical structure of Arg is also shown. B, ¹⁵N CP-MAS spectra of unquenched (dashed line) and quenched (solid line) LHCII. C, second derivatives of the ¹⁵N spectra in B.

FIGURE 4.

Heteronuclear ¹H-¹³C correlation spectra of unquenched (blue) and quenched (red) LHCII in the Arg ¹³C_ζ region (left) and in the ¹³C_δ region (right).

FIGURE 5.

Heteronuclear ¹H-¹³C correlation spectra of unquenched (A) and quenched (B) LHCII in the Arg ¹³C_α region.

FIGURE 6.

A, homology structure of C. reinhardtii LHCII highlighting the Arg residues. B, the geometric arrangements of Arg-70, Glu-180, and Chl610 taken from the 2BHW (3) LHCII x-ray structure.

FIGURE 7.

Graphic illustrating the possible effects of membrane thinning on LHCII. Compression of the LHC structure in A is achieved by (i) increased tilting of the transmembrane helices, causing structural changes in the Arg-Glu interlocked core (B; the affected core region is encircled) or (ii) reorientation of protein segments close to the water interface, causing structural changes in the regions near the lumenal and stromal sites (C; the affected end regions are encircled). A structural change as depicted in B will affect the orientations of Chl602 and Chl610 (green diamonds) that are ligated to the Arg-Glu interlocked core. However, such a structural rearrangement is unlikely according to the preserved NMR chemical shifts of the Chl-ligating Arg. The structural change depicted in C could reorient the luteins (orange rods) that have their head groups bound to the affected protein end regions.