Structure of the Dimeric PufX-containing Core Complex of Rhodobacter blasticus by in Situ Atomic Force Microscopy

Abstract

We have studied photosynthetic membranes of wild type Rhodobacter blasticus, a closely related strain to the well studied Rhodobacter sphaeroides, using atomic force microscopy. High-resolution atomic force microscopy topographs of both cytoplasmic and periplasmic surfaces of LH2 and RC-LH1-PufX (RC, reaction center) complexes were acquired in situ. The LH2 is a nonameric ring inserted into the membrane with the 9-fold axis perpendicular to the plane. The core complex is an S-shaped dimer composed of two RCs, each encircled by 13 LH1 ␣/␤-heterodimers, and two PufXs. The LH1 assembly is an open ellipse with a topography-free gap of ϳ25 Å. The two PufXs, one of each core, are located at the dimer center. Based on our data, we propose a model of the core complex, which provides explanation for the PufX-induced dimerization of the Rhodobacter core complex. The Q B site is located facing a ϳ25-Å wide gap within LH1, explaining the PufXfavored quinone passage in and out of the core complex.In purple photosynthetic bacteria, highly organized transmembrane pigment-protein complexes perform absorption of light and its conversion into chemical energy. Two light harvesting (LH) 1 complexes, LH2 and LH1, ensure the collection of light. The excitation energy is funneled toward the special pair (P) of bacteriochlorophylls in the reaction center (RC) followed by an electron transfer from P to the ubiquinone (Q) acceptors, Q A and Q B . After two photoreactions and proton captures, ubiquinol (Q-H 2 ) is formed at the Q B site that dissociates from the RC into the membrane. The cytochrome bc 1 complex utilizes Q-H 2 and oxidized cytochrome c 2 as reductant and oxidant, respectively. The net result is a cyclic electron transfer that promotes the formation of a proton gradient across the membrane, which is utilized for ATP synthesis by the F 1 F 0 -ATP synthase (for review see Ref. 1).The description of the bacterial photosynthetic apparatus at atomic level is nearly complete. Two RC structures (2-4), two LH2 structures (5, 6), and the structure of the homologue bc 1 complex from the respiratory chain (7) are known. However, the structure of the core complex composed of the LH1 and the RC remains undetermined.Over the last two years, the data of core complexes of Rhodospirillum (Rsp.) rubrum (8), Blastochloris viridis (9), and Rhodopseudomonas (Rps.) palustris (10) have been acquired at a resolution sufficient to delineate the LH1 subunit arrangement around the RC. Whereas Rsp. rubrum and B. viridis have monomeric core complexes with 16 LH1 subunits arranged around the RC, Rps. palustris has 15 LH1 subunits plus an unknown polypeptide forming a single trans-membrane helix. The closed architecture of the LH1 assembly raised the question of the quinone exchange through the LH1 assembly between the RC and the cytochrome bc 1 .In this context, the structural analysis of Rhodobacter core complexes is of particular interest, because it is the only species in which a small trans-membrane protein named PufX has been identified as crucial ...

Structure of the dimeric RC–LH1–PufX complex from Rhodobaca bogoriensis investigated by electron microscopy

“…The N-terminal part of PufX was found to be critical for the formation of dimers of the RC-LH1 and, in the refined dimer model, the N-termini of the two PufX proteins point to the interface between the monomers. In a second type of model, PufX is placed at the dimerization interface in the centre of the dimer complex and is itself also thought to be dimerized [5,10,12]. In another model, the two PufX molecules are also placed in the centre of the dimer, but not in close contact [8].…”

The Organization of LH2 Complexes in Membranes from Rhodobacter sphaeroides

“…We have demonstrated that the comparatively disordered intracytoplasmic membrane from the model photosynthetic bacterium Rhodobacter sphaeroides can be successfully imaged by AFM (9) to reveal the hidden architecture that nature has developed to harvest, transfer, and finally utilize light energy with great efficiency. Scheuring and co-workers have also produced high resolution images of the photosynthetic membranes of Blastochloris viridis (10), Rhodospirillum photometricum (11), Rhodobacter blasticus (12), and Phaeospirillum molischianum (13), allowing a comparison of the differing strategies that purple bacteria have evolved for the harvesting and utilization of light energy (14).…”

Refinement of the x-ray structure of the RC–LH1 core complex from Rhodopseudomonas palustris by single-molecule spectroscopy

“…Because of the lack of a high-resolution x-ray structure, most proposed geometries are based on low-resolution electron microscopy/atomic force microscopy data (24)(25)(26)(27)(28). A detailed discussion on this issue can be found in ref.…”