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The vault particle, with a molecular weight of about 10 MDa, is the largest ribonucleoprotein that has been described. The X-ray structure of intact rat vault has been solved at a resolution of 3.5 Å [Tanaka et al. (2009), Science, 323, 384-388], showing an overall barrel-shaped architecture organized into two identical moieties, each consisting of 39 copies of the major vault protein (MVP). The model deposited in the PDB includes 39 MVP copies (half a vault) in the crystal asymmetric unit. A 2.1 Å resolution structure of the seven N-terminal repeats (R1-7) of MVP has also been determined [Querol-Audí et al. (2009), EMBO J. 28, 3450-3457], revealing important discrepancies with respect to the MVP models for repeats R1 and R2. Here, the re-refinement of the vault structure by incorporating the high-resolution information available for the R1-7 domains, using the deformable elastic network (DEN) approach and maintaining strict 39-fold noncrystallographic symmetry is reported. The new refinement indicates that at the resolution presently available the MVP shell can be described well as only one independent subunit organized with perfect D39 molecular symmetry. This refinement reveals that significant rearrangements occur in the N-terminus of MVP during the closing of the two vault halves and that the 39-fold symmetry breaks in the cap region. These results reflect the highly dynamic nature of the vault structure and represent a necessary step towards a better understanding of the biology and regulation of this particle. [ABSTRACT FROM AUTHOR]

Intercellular signalling is an essential characteristic of multicellular organisms. Gap junctions, which consist of arrays of intercellular channels, permit the exchange of ions and small molecules between adjacent cells. Here, the structural determination of a gap junction channel composed of connexin 26 (Cx26) at 3.5 Å resolution is described. During each step of the purification process, the protein was examined using electron microscopy and/or dynamic light scattering. Dehydration of the crystals improved the resolution limits. Phase refinement using multi-crystal averaging in conjunction with noncrystallographic symmetry averaging based on strictly determined noncrystallographic symmetry operators resulted in an electron-density map for model building. The amino-acid sequence of a protomer structure consisting of the aminoterminal helix, four transmembrane helices and two extracellular loops was assigned to the electron-density map. The amino-acid assignment was confirmed using six selenomethionine (SeMet) sites in the difference Fourier map of the SeMet derivative and three intramolecular disulfide bonds in the anomalous difference Fourier map of the native crystal. [ABSTRACT FROM AUTHOR]

The absolute configuration of haem A, the prosthetic group of cytochrome c oxidase, was determined to be S by analysis of the bond angles surrounding the chiral centre of haem A after refinement with X-PLOR starting from respective initial structures with R and S configurations under absolute configuration constraints at 1.8 Å resolution. The same result was obtained by refinement at 1.8 Å resolution without the absolute configuration constraints. Both of these methods were applicable down to a resolution of about 2.8 Å. The constrained refinement converges more quickly than the unconstrained refinement. [ABSTRACT FROM AUTHOR]

Ab initio phasing by non-crystallographic symmetry averaging coupled with solvent flattening has previously been used to determine the structure of canine parvovirus [CPV; Tsao et al. (1992), Acta Cryst. A48, 293-301; Chapman et al. (1992), Acta Cryst. A48, 301-312]. As CPV particles were located at general positions, initial phases were generated with a spherical shell deviating from the centre of symmetry. In many virus crystals, the viral particles are located at positions with a centre of symmetry in the unit cell. Thus, the initial phases calculated with a spherical shell model have a centre of symmetry. The inherent difficulty in structural determination was breaking the centre of symmetry in the initial phase. The centric nature of the initial phases of the rice dwarf virus crystal, however, was successfully broken at low resolution by iteration of the density-modification method described by Tsao et al. (1992) [Naitow et al. (1999), Acta Cryst. D55, 77-84]. In this study, ab initio phasing was tested for seven viruses ranging from 82 to 344 Å in radius. Although the crystal structures of the initial spherical shell models for each virus had a centre of symmetry, the centric natures of the initial phases were successfully broken by non-crystallographic symmetry averaging coupled with solvent flattening at a low resolution; these phases were then successfully extended to high-resolution measurements. A novel procedure of ab initio phasing for spherical virus crystals is proposed. [ABSTRACT FROM AUTHOR]

The X-ray structure of bovine heart cytochrome c oxidase solved for orthorhombic crystals showed a dimeric structure stabilized by four subunit-subunit contacts, namely, subunit Vb-subunit Vb on the matrix side, subunit I-subunit Vla, subunit Vla-subunit I in the transmembrane region and subunit VIb-subunit VIb on the intermembrane side. The same intermonomer contacts as in the orthorhombic crystals were observed in both hexagonal and tetragonal crystals, the X-ray structures of which were determined by the molecular-replacement method. These results suggest that the dimeric structure also exists under physiological conditions. These contacts, especially the subunit IVa-subunit I contact, in which the N-terminal portion of subunit IVa is placed on the surface of subunit I near the dioxygen-reduction site, indicate that the function of the bovine heart enzyme is likely to be controlled by perturbation of the monomer-monomer association. [ABSTRACT FROM AUTHOR]

Two azide ions were identified, one between the Fe and Cu atoms in the O2-reduction site and the other at the transmembrane surface of the enzyme, in the crystal structure of the azide-bound form of bovine heart cytochrome c oxidase at 2.9 Å resolution. Two geometries, a μ-1,3 type geometry between the Fe and Cu atoms and a terminal geometry on the Fe atom, are equally possible for an azide ion in the O2-reduction site. The other azide molecule was hydrogen bonded to an amide group of an asparagine and a hydroxyl group of tyrosine in a μ-1,1 type geometry. The antisymmetric infrared bands arising from these azide ions, which show essentially identical intensity [Yoshikawa & Caughey (1992), J. Biol. Chem. 267, 9757-97661, strongly suggest terminal binding of the azide to Fe. The electron density of all three imidazole ligands to CUB was clearly seen in the electron-density map of the azide-bound form of bovine heart enzyme, in contrast to the crystal structure of the azide-bound form of the bacterial enzyme [Iwata et al. (1995), Nature (London), 376, 660-669], which lacks one of the three imidazole ligands to CuB. [ABSTRACT FROM AUTHOR]

The article discusses an anomalous dispersion analysis of the mechanism of coupling the proton pumping process to the electron transfer reaction in resting-oxidized cytochrome c oxydase. The study suggests that peroxide moiety receives electron equivalents from cytochrome c without affecting the oxidation states of the metal sites. It further suggests that the metal site reduction is coupled to the proton pump.

Studies the crystallization and preliminary crystallographic analysis of rat monoamine oxidase A (MAO) complexed with clorgyline. Primary sequences but different substrate and inhibitor specificities in the two subtypes of MAO; Mechanisms underlying the unique substrate and inhibitor recognition.

The nuclear-transport protein importin-β mediates the nuclear import of the transcription factor SREBP-2 without requiring adaptor proteins such as importin-α, An importin-Β-SREBP-2 HLHZ domain complex was purified and crystallized. The crystals belong to space group P2[SUB1]2[SUB1]2[SUB1] and show diffraction to at least 3.0 Å resolution. The unit-cell parameters are a = 101.0, b = 113.2, c = 240.0 Å. Structure determination using the MAD or SAD method is under way. [ABSTRACT FROM AUTHOR]

S-RNase is a 25 kDa pistil-specific protein associated with gameto- phytic self-incompatibility. The S-RNase is secreted into the transmitting tissue of the pistil and is responsible for the discrimination of pollen S-alleles. Crystals of Japanese pear S3-RNase were obtained by the hanging-drop vapour-diffusion method. Preliminary X-ray data showed that the crystals diffract to a 1.5 Å resolution and belong to the space group P21, with unit-cell parameters a = 45.4, b = 52.4, c = 47.4 Å, a = γ = 90, β = 106.5°. [ABSTRACT FROM AUTHOR]

The β-carbonic anhydrase from the red alga Porphyridium purpureum was heterologously expressed, purified and crystallized. The crystals belong to space group P21 (unit-cell parameters a = 63.8, b = 113.9, c = 73.8 Å, β = 104.1°) with two subunits per asymmetric unit and diffract to 2.5 Å resolution. [ABSTRACT FROM AUTHOR]