Cryo-EM structure of the dimeric Rhodobacter sphaeroides RC-LH1 core complex at 2.9 Å: the structural basis for dimerisation

Abstract

The dimeric reaction centre light-harvesting 1 (RC-LH1) core complex of Rhodobacter sphaeroides converts absorbed light energy to a charge separation, and then it reduces a quinone electron and proton acceptor to a quinol. The angle between the two monomers imposes a bent configuration on the dimer complex, which exerts a major influence on the curvature of the membrane vesicles, known as chromatophores, where the light-driven photosynthetic reactions take place. To investigate the dimerisation interface between two RC-LH1 monomers, we determined the cryogenic electron microscopy structure of the dimeric complex at 2.9 Å resolution. The structure shows that each monomer consists of a central RC partly enclosed by a 14-subunit LH1 ring held in an open state by PufX and protein-Y polypeptides, thus enabling quinones to enter and leave the complex. Two monomers are brought together through N-terminal interactions between PufX polypeptides on the cytoplasmic side of the complex, augmented by two novel transmembrane polypeptides, designated protein-Z, that bind to the outer faces of the two central LH1 β polypeptides. The precise fit at the dimer interface, enabled by PufX and protein-Z, by C-terminal interactions between opposing LH1 αβ subunits, and by a series of interactions with a bound sulfoquinovosyl diacylglycerol lipid, bring together each monomer creating an S-shaped array of 28 bacteriochlorophylls. The seamless join between the two sets of LH1 bacteriochlorophylls provides a path for excitation energy absorbed by one half of the complex to migrate across the dimer interface to the other half.

Keywords: bacteriochlorophylls; carotenoids; light harvesting; photosynthesis; reaction centre; transmembrane proteins.

Figure 1.. Cryo-EM structure of the dimeric RC-LH1 complex from Rba. sphaeroides.

(A,C,E) Views of the density map, coloured as in the key at the bottom of the figure. Detergent and other disordered molecules are in grey. (B,E,F) Ribbon models of the complex, made using ChimeraX [34]. (A) View of the cytoplasmic face of the density map of the complex, showing the diameters of the short axes of the detergent belt and the complex. (B) View as in (A), as a ribbon model; the LH1 subunits are numbered. (C) View of the density map in the plane of the membrane showing the height of the complex. (D) View as in (C), as a ribbon model. (E) Perpendicular view of the density map from the periplasmic side with measurements of the long-axis of the complex and detergent micelle. (F) Ribbon model corresponding to (E).

Figure 2.. The positions and transmembrane interactions of the two protein-Z polypeptides.

(A) View of part of the LH1 ring near the monomer–monomer interface, perpendicular to the membrane from the cytoplasmic side. The transmembrane regions of protein-Z1 and protein-Z2 are closely appressed against the LH1 β1 and β2 polypeptides. (B) View in the plane of the membrane, showing hydrogen bonds (dashed lines) between Z1/Z2 and the respective β1 and β2 polypeptides. Other bonds with LH1 α1 and PufX near the cytoplasmic face of the membrane are labelled. Supplementary Table S2 lists the hydrogen bonds relating to Figure 2B. (C) ‘Open book’ format to show the opposing, interacting hydrophobic faces of Z1 and Z2 and the respective β1 and β2 polypeptides. The labelled residues are predicted to be in van der Waals contact.

Figure 3.. Protein–protein interactions at the dimer interface.

(A) View of the cytoplasmic side of the dimer. For clarity, all components are faded except for those at the interface. (B) Detailed view of the region indicated by the box in (A) showing PufX and PufX’ interacting near their N-termini on the cytoplasmic side of the membrane, with important sidechains labelled. (C) View of the periplasmic side of the dimer, with other features as in (A). (D) Detailed view of the region indicated by the left-hand box in (C), and rotated by 90°, showing C-terminal interactions between opposing LH1 α, α′ and β, β′ subunits at positions 1 and 1′ (see Figure 1B for numbering). Dashed lines indicate hydrogen bonds, which are 3.3 Å (see also Supplementary Table S2). (E) The components shown in the right-hand box in (C), but viewed from a different angle, showing C-terminal interactions between PufX on one side of the dimer complex, and protein-Z′ and the LH1 β′ subunit at position 1′ on the other side. The hydrogen bond between X-Arg53 and β1′-Ile44 is 2.8 Å (see also Supplementary Table S2).

Figure 4.. Lipid–protein interactions at the dimer interface.

(A) View of the interface from the periplasmic side; the diagonal dashed line indicates the approximate position of the interface. For clarity, BChl and carotenoid pigments have been omitted. SQDG lipids are shown within a mesh representing the density; the solid grey densities for another, unassigned, lipid (Lipid-2) are also shown. (B) Details of the hydrogen bonds (dashed lines) between the SQDG headgroup and backbone (bb) oxygen or nitrogen, and also with Arg49 and Arg53 near the C-terminus of PufX. Supplementary Table S2 lists the hydrogen bonds relating to Figure 4B. (C) View of the dimer interface, showing the disposition of the SQDG lipid tails as they reach up towards the cytoplasmic side of the complex. Sidechains of RC-L, PufX and the opposing LH1β′ subunit that interact with the lipid tails are labelled. For convenience, the numbering of RC-L residues follows the other entries for these RCs in the PDB in omitting the first Met and starting at the second residue (see also Supplementary Figure S5).

Figure 5.. The role of PufX in imposing a bent conformation on the RC-LH1 dimer complex.

Pigments and lipids, as well as most of the LH1 subunits and the RC, have been omitted for clarity. (A) Top view, from the cytoplasmic side of the membrane. The arrows within the ellipse illustrate the attractive interactions between N-terminal regions of opposing PufX polypeptides, which help to bind the two halves of the dimer together. The diverging arrows represent the effects of each PufX as it lies diagonally across its adjacent LH1αβ subunit, pushing them apart. (B) As in (A) but viewed in the plane of the membrane, with the N-terminal PufX interactions within the circle, and multiple attractive interactions near the periplasmic face, within the ellipse, that hold the bottom halves of the complex tightly together. As in (A) the diverging arrows represent the central LH1αβ subunits being pushed apart by the PufX transmembrane regions.

Figure 6.. The bacteriochlorophyll and carotenoid pigments in the RC-LH1 dimer.

Proteins are faded for clarity. The dashed line divides the complex into the two halves. The central pigments belong to the RCs. (A) View from the periplasmic side of the membrane showing the two arcs of 28 BChls. Within one monomer half the BChls are coloured in two shades of green to distinguish between pairs belonging to individual LH1 αβ subunits, and in the other half two shades of blue are used. Similarly, two shades of orange or magenta indicate pairs of carotenoids belonging to individual LH1 αβ subunits on each half of the dimer. (B) View in the plane of the membrane. The box indicates the interface zone magnified in panel (C). (C) BChl and carotenoid pigments at the interface, coloured as in (A), and with the α and β polypeptides numbered as in Figure 1B. Rings A–E of the BChl macrocycles are labelled, and Mg–Mg distances are shown in Ångstroms for intra- and inter-subunit BChls.