Membrane insertion of Rhodopseudomonas acidophila light harvesting complex 2 investigated by high resolution AFM

Gonçalves, Rui Pedro; Busselez, Johan; Lévy, Daniel; Seguin, Jérôme; Scheuring, Simon

doi:10.1016/j.jsb.2004.09.001

Cited by 35 publications

(39 citation statements)

References 31 publications

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“…A similar observation was made for membrane-reconstituted LH2 complexes from Rps. acidophila, where clustering of the LH2 complexes coexisting with protein-free lipid areas was found (Gonçalves et al 2005). A recent AFM study on LH complexes directly reconstituted into supported lipid bilayers also showed a network of clustered LH complexes (Milhiet et al 2006).…”

Section: Discussionmentioning

confidence: 92%

Comparison of the fluorescence kinetics of detergent-solubilized and membrane-reconstituted LH2 complexes from Rps. acidophila and Rb. sphaeroides

et al. 2007

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Picosecond time-resolved fluorescence spectroscopy has been used in order to compare the fluorescence kinetics of detergent-solubilized and membrane-reconstituted light-harvesting 2 (LH2) complexes from the purple bacteria Rhodopseudomonas (Rps.) acidophila and Rhodobacter (Rb.) sphaeroides. LH2 complexes were reconstituted in phospholipid model membranes at different lipid:protein-ratios and all samples were studied exciting with a wide range of excitation densities. While the detergent-solubilized LH2 complexes from Rps. acidophila showed monoexponential decay kinetics (tau(f )= 980 ps) for excitation densities of up to 3.10(13) photons/(pulse.cm(2)), the membrane-reconstituted LH2 complexes showed multiexponential kinetics even at low excitation densities and high lipid:protein-ratios. The latter finding indicates an efficient clustering of LH2 complexes in the phospholipid membranes. Similar results were obtained for the LH2 complexes from Rb. sphaeroides.

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Section: Discussionmentioning

confidence: 92%

Comparison of the fluorescence kinetics of detergent-solubilized and membrane-reconstituted LH2 complexes from Rps. acidophila and Rb. sphaeroides

et al. 2007

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“…An AFM image of the 2D-crystal of the LH2 complexes from Rps. acidophila in DOPC/cardiolipin lipid bilayer has recently reported (Gonçalves et al 2005b): They analyzed two types of 2D-crystals, both have two ''up'' and two ''down'' orientations of molecules, showing an (ortho)rhombic lattice. The type-1 crystal shown in Fig.…”

Section: Discussionmentioning

confidence: 99%

Construction of hybrid photosynthetic units using peripheral and core antennae from two different species of photosynthetic bacteria: detection of the energy transfer from bacteriochlorophyll a in LH2 to bacteriochlorophyll b in LH1

et al. 2007

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Typical purple bacterial photosynthetic units consist of supra-molecular arrays of peripheral (LH2) and core (LH1-RC) antenna complexes. Recent atomic force microscopy pictures of photosynthetic units in intact membranes have revealed that the architecture of these units is variable (Scheuring et al. (2005) Biochim Bhiophys Acta 1712:109-127). In this study, we describe methods for the construction of heterologous photosynthetic units in lipid-bilayers from mixtures of purified LH2 (from Rhodopseudomonas acidophila) and LH1-RC (from Rhodopseudomonas viridis) core complexes. The architecture of these reconstituted photosynthetic units can be varied by controlling ratio of added LH2 to core complexes. The arrangement of the complexes was visualized by electron-microscopy in combination with Fourier analysis. The regular trigonal array of the core complexes seen in the native photosynthetic membrane could be regenerated in the reconstituted membranes by temperature cycling. In the presence of added LH2 complexes, this trigonal symmetry was replaced with orthorhombic symmetry. The small lattice lengths for the latter suggest that the constituent unit of the orthorhombic lattice is the LH2. Fluorescence and fluorescence-excitation spectroscopy was applied to the set of the reconstituted membranes prepared with various proportions of LH2 to core complexes. Remarkably, even though the LH2 complexes contain bacteriochlorophyll a, and the core complexes contain bacteriochlorophyll b, it was possible to demonstrate energy transfer from LH2 to the core complexes. These experiments provide a first step along the path toward investigating how changing the architecture of purple bacterial photosynthetic units affects the overall efficiency of light-harvesting.

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“…The LH2 structure is well described by X-ray crystallography (16,17) and AFM in native membranes (12,(18)(19)(20). Typically, the LH2 complexes appear as circular oligomers of nine αβ subunits, including 18 long-wavelength-absorbing bacteriochlorophyll (BChl) molecules sandwiched between the inner ring of α-polypeptides and the outer ring of β-polypeptides.…”

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confidence: 99%

Forces guiding assembly of light-harvesting complex 2 in native membranes

Liu

Duquesne

Oesterhelt

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Interaction forces of membrane protein subunits are of importance in their structure, assembly, membrane insertion, and function. In biological membranes, and in the photosynthetic apparatus as a paradigm, membrane proteins fulfill their function by ensemble actions integrating a tight assembly of several proteins. In the bacterial photosynthetic apparatus light-harvesting complexes 2 (LH2) transfer light energy to neighboring tightly associated core complexes, constituted of light-harvesting complexes 1 (LH1) and reaction centers (RC). While the architecture of the photosynthetic unit has been described, the forces and energies assuring the structural and functional integrity of LH2, the assembly of LH2 complexes, and how LH2 interact with the other proteins in the supramolecular architecture are still unknown. Here we investigate the molecular forces of the bacterial LH2 within the native photosynthetic membrane using atomic force microscopy single-molecule imaging and force measurement in combination. The binding between LH2 subunits is fairly weak, of the order of k B T , indicating the importance of LH2 ring architecture. In contrast LH2 subunits are solid with a free energy difference of 90 k B T between folded and unfolded states. Subunit α-helices unfold either in one-step, α-and β-polypeptides unfold together, or sequentially. The unfolding force of transmembrane helices is approximately 150 pN. In the two-step unfolding process, the β-polypeptide is stabilized by the molecular environment in the membrane. Hence, intermolecular forces influence the structural and functional integrity of LH2.atomic force microscopy | photosynthesis | protein unfolding | membrane protein assembly M embrane protein function is intimately linked and influenced by its structural integrity and molecular environment (1-3). Photosynthesis is a paradigm of such orchestrated molecular action, for which the implicated proteins have evolved to form supramolecular assemblies (4). Knowledge of supramolecular architecture and interaction forces and energies are indispensable for a complete understanding of a membrane protein structure and function.The atomic force microscope (AFM) (5) can be used as a high-resolution imaging (6) and as a force-measurement tool (7). Most elegantly the two applications were combined allowing the attribution of structural changes to force events (8). More recently, the AFM has proven the unique tool for studying the supramolecular assembly of the bacterial photosynthetic unit (9-12) and has provided a solid basis for the understanding of the ensemble function of the photosynthetic proteins (4, 13-15).The light-harvesting process in photosynthetic bacteria starts with photon capture at the peripheral light-harvesting complex 2 (LH2) that transfers the absorbed excitation energy to the light-harvesting complex 1 (LH1) associated with the reaction center (RC). The LH2 structure is well described by X-ray crystallography (16, 17) and AFM in native membranes (12,(18)(19)(20). Typically, the LH2 complexes appe...

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Membrane insertion of Rhodopseudomonas acidophila light harvesting complex 2 investigated by high resolution AFM

Cited by 35 publications

References 31 publications

Comparison of the fluorescence kinetics of detergent-solubilized and membrane-reconstituted LH2 complexes from Rps. acidophila and Rb. sphaeroides

Comparison of the fluorescence kinetics of detergent-solubilized and membrane-reconstituted LH2 complexes from Rps. acidophila and Rb. sphaeroides

Construction of hybrid photosynthetic units using peripheral and core antennae from two different species of photosynthetic bacteria: detection of the energy transfer from bacteriochlorophyll a in LH2 to bacteriochlorophyll b in LH1

Forces guiding assembly of light-harvesting complex 2 in native membranes

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