Detergent structure in tetragonal crystals of OmpF porin

Pebay‐Peyroula, Eva; Garavito, R. Michael; Rosenbusch, J P; Zulauf, M.; Timmins, P.

doi:10.1016/s0969-2126(01)00241-6

Cited by 109 publications

(93 citation statements)

References 21 publications

(39 reference statements)

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“…At the convex side of the b-barrel two rings of aromatic residues delimit the modeled detergent density (Fig. 4a), similar to the rings observed in the detergent-protein structure of OmpF (Pebay-Peyroula et al, 1995;Penel et al, 1998) and the general porin from Rhodobacter capsulatus (Penel, 1997). The aromatic rings fulfill a role in delimiting the detergent phase.…”

Section: Detergent Structuresupporting

confidence: 68%

“…Knowledge of the factors that govern their crystallisation may help to rationalise the quest for diffraction quality crystals of membrane proteins. In this respect, analyses of the packing of detergent and membrane proteins in the crystal may contribute to achieving this (Pebay-Peyroula et al, 1995;Penel et al, 1998;Roth et al, 1991Roth et al, , 1989. The structure of the detergent phase cannot be determined routinely using X-ray diffraction, owing to the fluidity of the detergent molecules and their low X-ray contrast.…”

Section: Introductionmentioning

confidence: 99%

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Detergent organisation in crystals of monomeric outer membrane phospholipase A

Snijder¹,

Timmins²,

Kalk³

et al. 2003

Journal of Structural Biology

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The structure of the detergent in crystals of outer membrane phospholipase A (OMPLA) has been determined using neutron diffraction contrast variation. Large crystals were soaked in stabilising solutions, each containing a different H 2 O=D 2 O contrast. From the neutron diffraction at five contrasts, the 12 A A resolution structure of the detergent micelle around the protein molecule was determined. The hydrophobic b-barrel surfaces of the protein molecules are covered by rings of detergent. These detergent belts are fused to neighbouring detergent rings forming a continuous three-dimensional network throughout the crystal. The thickness of the detergent layer around the protein varies from 7-20 A A. The enzymeÕs active site is positioned just outside the hydrophobic detergent zone and is thus in a proper location to catalyse the hydrolysis of phospholipids in a natural membrane. Although the dimerisation face of OMPLA is covered with detergent, the detergent density is weak near the exposed polar patch, suggesting that burying this patch in the enzymeÕs dimer interface may be energetically favourable. Furthermore, these results indicate a crucial role for detergent coalescence during crystal formation and contribute to the understanding of membrane protein crystallisation.

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Section: Detergent Structuresupporting

confidence: 68%

Section: Introductionmentioning

confidence: 99%

Detergent organisation in crystals of monomeric outer membrane phospholipase A

Snijder¹,

Timmins²,

Kalk³

et al. 2003

Journal of Structural Biology

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“…Localization of the e-and gsubunits reveals that they extend from the rest of the membranebound region distal to the c 8 -ring. The structure of the DDM micelle seen in the present map possesses a strong curvature, atypical of previously observed detergent micelles for membrane proteins from planar lipid bilayers (26,27). The results suggest that ATP synthase would produce a curved membrane structure.…”

supporting

confidence: 67%

“…detergent can be visualized in 3D maps from amorphous iceembedded protein particles. In these other maps, the detergent micelle has a torroidal or donut shape, which one would expect from standard models of detergent micelles in detergent-protein complexes (27). In the map of the bovine ATP synthase, the micelle has a marked bend.…”

Section: Resultsmentioning

confidence: 95%

Arrangement of subunits in intact mammalian mitochondrial ATP synthase determined by cryo-EM

Baker

Watt

Runswick

et al. 2012

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

112

109

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Mitochondrial ATP synthase is responsible for the synthesis of ATP, a universal energy currency in cells. Whereas X-ray crystallography has revealed the structure of the soluble region of the complex and the membrane-intrinsic c-subunits, little is known about the structure of the six other proteins (a, b, f, A6L, e, and g) that comprise the membrane-bound region of the complex in animal mitochondria. Here, we present the structure of intact bovine mitochondrial ATP synthase at ∼18 Å resolution by electron cryomicroscopy of single particles in amorphous ice. The map reveals that the a-subunit and c 8 -ring of the complex interact with a small contact area and that the b-subunit spans the membrane without contacting the c 8 -ring. The e-and g-subunits extend from the a-subunit density distal to the c 8 -ring. The map was calculated from images of a preparation of the enzyme solubilized with the detergent dodecyl maltoside, which is visible in electron cryomicroscopy maps. The structure shows that the micelle surrounding the complex is curved. The observed bend in the micelle of the detergent-solubilized complex is consistent with previous electron tomography experiments and suggests that monomers of ATP synthase are sufficient to produce curvature in lipid bilayers.A TP synthases are responsible for the synthesis of ATP from ADP and inorganic phosphate. In mammalian mitochondria, the enzyme is an ∼600-kDa membrane protein complex comprised of a catalytic F 1 region and a membrane-bound F O region. The F 1 region consists of subunits α 3 β 3 γδε (1, 2) and the F O region consists of subunits abc 8 defg(A6L)F 6 (3). The F 1 and F O regions are connected by a central stalk, comprised of the γ-, δ-, and ε-subunits from the F 1 region, and a peripheral stalk, comprised of the oligomycin sensitivity conferral protein (OSCP) and subunits b, d, and F 6 from the F O region (4, 5). Previously, electron cryomicroscopy (cryo-EM) of intact detergent-solubilized bovine ATP synthase particles embedded in a thin layer of amorphous ice revealed the overall shape of the complex at 32 Å resolution (6), and subsequent analysis of the Saccharomyces cerevisiae enzyme produced a similar map at 24 Å resolution (7). X-ray crystallography of subcomplexes of the bovine enzyme has defined the arrangement of subunits in the F 1 region (8) and the peripheral stalk (9, 10) and showed the presence of a ring of eight c-subunits in the F O region (11). The structure and arrangement of the remaining subunits in the membrane-bound region of the enzyme are not known.The ATP synthase functions by a rotary catalytic mechanism. Proton translocation through the F O region requires the a-, b-, and c-subunits (12-14) and induces rotation of the membranebound c 8 -ring (15). The structure of the c 8 -ring is thought to be stabilized by binding of cardiolipin to a lysine residue conserved throughout animalia that has been shown to be trimethylated at the ε-amino group in all animal ATP synthases tested (11,16,17). The c 8 -ring is attached to the central st...

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“…Contrast-variation methods in neutron or X-ray crystallography represent an exception to this situation. Experiments with chemical contrast variation, using X-rays (Dumas, 1988;Carter et al, 1990) or isotopical contrast variation, using neutrons (Moras et al, 1983;Roth et al, 1984;Bentley et al, 1984;Podjarny et al, 1987;Timmins et al, 1992;Pebay-Peroula et al, 1995) have been conducted. With X-rays, the main limitation in chemical contrast variation comes from the fact that several samples are used with different solvent contents that may induce structural changes in the macromolecule leading to non-isomorphism.…”

Section: Introductionmentioning

confidence: 99%

Multiwavelength anomalous solvent contrast (MASC): derivation of envelope structure-factor amplitudes and comparison with model values

Ramin¹,

Shepard²,

Fourme³

et al. 1999

Acta Crystallogr D Biol Cryst

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A previous article [Fourme et al. (1995). J. Synchrotron Rad. 2, 36±48] presented the theoretical foundations of MASC, a new contrast-variation method using multiwavelength anomalous scattering, and reported the ®rst experimental results. New experiments have been conducted both at the ESRF (Grenoble, France) and at LURE-DCI (Orsay, France), using cryocooled crystals of three proteins of known structures and very different molecular weights. Amplitudes of fÀ T h g, thè normal' structure factors of the anomalously scattering part of the crystal including the solvent zone and the ordered anomalous scattering sites (if any), have been extracted from multiwavelength data. In the very low resolution range (d ! 20 A Ê ), the agreement between experimental fjÀ T h jg and model values calculated from the bulk solvent is all the more satisfactory since the molecular weight of the protein is high. For spacings between 10 and 20 A Ê , the agreement between experimental fjÀ T h jg and model values is also satisfactory if one takes into account ordered anomalous scatterer sites. Such sites have been found in the three cases.

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Detergent structure in tetragonal crystals of OmpF porin

Cited by 109 publications

References 21 publications

Detergent organisation in crystals of monomeric outer membrane phospholipase A

Detergent organisation in crystals of monomeric outer membrane phospholipase A

Arrangement of subunits in intact mammalian mitochondrial ATP synthase determined by cryo-EM

Multiwavelength anomalous solvent contrast (MASC): derivation of envelope structure-factor amplitudes and comparison with model values

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