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Self-rotation Function (self-rotation + function)

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Chemistry100%

Terms modified by Self-rotation Function

  • self-rotation function analysis
    		•

    Selected Abstracts

    Crystallization and preliminary X-ray study of recombinant betaine,homocysteine S -­methyltransferase from rat liver

    ACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2002
    Beatriz González
    Betaine,homocysteine S -methyltransferase is one of the three enzymes involved in homocysteine catabolism. It uses betaine as the methyl donor to convert homocysteine into methionine, also producing dimethylglycine. Recombinant BHMT from rat liver was crystallized by the vapour-diffusion method in both native and seleniomethionyl-labelled forms. Crystals belong to space group P21, with unit-cell parameters a = 57.8, b = 149.3, c = 96.2,Å, , = 92.9°. Data from native, seleniomethionine-labelled and two heavy-atom derivatives were collected using synchrotron sources. Self-rotation function and sedimentation-velocity experiments suggest that the enzyme is tetrameric with 222 symmetry. [source]

    Crystallization and preliminary X-ray diffraction analysis of the fructofuranosidase from Schwanniomyces occidentalis

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2009
    Aitana Polo
    Schwanniomyces occidentalis invertase is an extracellular enzyme that releases ,-fructose from the nonreducing termini of various ,- d -fructofuranoside substrates. Its ability to produce 6-kestose by transglycosylation makes this enzyme an interesting research target for applications in industrial biotechnology. The enzyme has been expressed in Saccharomyces cerevisiae. Recombinant and wild-type forms, which showed different glycosylation patterns, were crystallized by vapour-diffusion methods. Although crystallization trials were conducted on both forms of the protein, crystals suitable for X-ray crystallographic analyses were only obtained from the wild-type enzyme. The crystals belonged to space group P212121, with unit-cell parameters a = 105.78, b = 119.49, c = 137.68,Å. A diffraction data set was collected using a synchrotron source. Self-rotation function and sedimentation-velocity experiments suggested that the enzyme was dimeric with twofold symmetry. [source]

    Cloning, purification and preliminary X-ray analysis of the C-terminal domain of Helicobacter pylori MotB

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2008
    Anna Roujeinikova
    The C-terminal domain of MotB (MotB-C) contains a putative peptidoglycan-binding motif and is believed to anchor the MotA/MotB stator unit of the bacterial flagellar motor to the cell wall. Crystals of Helicobacter pylori MotB-C (138 amino-acid residues) were obtained by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant. These crystals belong to space group P21, with unit-cell parameters a = 50.8, b = 89.5, c = 66.3,Å, , = 112.5°. The crystals diffract X-rays to at least 1.6,Å resolution using a synchrotron-radiation source. Self-rotation function and Matthews coefficient calculations suggest that the asymmetric unit contains one tetramer with 222 point-group symmetry. The anomalous difference Patterson maps calculated for an ytterbium-derivative crystal using diffraction data at a wavelength of 1.38,Å showed significant peaks on the v = 1/2 Harker section, suggesting that ab initio phase information could be derived from the MAD data. [source]

    Crystallization of Saccharomyces cerevisiae aminopeptidase 1, the major cargo protein of the Cvt pathway

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2007
    Wakana Adachi
    The vacuole hydrolase aminopeptidase 1 (Ape1) is a cargo protein transported to the vacuole by the cytosol-to-vacuole targeting (Cvt) pathway during conditions of growth and by autophagy during conditions of starvation. After transport to the vacuole, Ape1 is processed into mature Ape1 (mApe1). mApe1 has been expressed, purified and crystallized in two crystal forms. Form I belongs to space group P21, with unit-cell parameters a = 120.6, b = 219.5, c = 133.1,Å, , = 116.5°. Form II belongs to space group R3, with unit-cell parameters a = 141.2, c = 349.4,Å. Diffraction data were collected from these crystals to a resolution of 2.5,Å for form I and 1.83,Å for form II. Self-rotation functions and the volume-to-weight ratio values suggest that forms I and II contain 12 and four mApe1 molecules per asymmetric unit, respectively, and that mApe1 exists as a tetrahedral dodecamer in both crystal forms. [source]

    Structure determination of adeno-associated virus 2: three complete virus particles per asymmetric unit

    ACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2003
    Qing Xie
    The atomic structure of adeno-associated virus 2 (AAV-2) has been determined to 3.0,Å resolution. AAV-2 crystallized in space group P1, with unit-cell parameters a = 249.7, b = 249.7, c = 644.8,Å, , = 90.0, , = 101.2, , = 120.0°. The crystals contained three full virus particles in the asymmetric unit, allowing 180-fold non-crystallographic symmetry averaging. The particle orientations were determined using the self-rotation function and found to have similar but resolvably different orientations. Approximate alignment of icosahedral and interparticle threefold screw symmetry led to a native Patterson that was interpretable in terms of approximate particle positions. Accurate positions required a Patterson correlation search that was constrained to be consistent with non-crystallographic threefold projection symmetry evident in the diffraction intensities. Initial phases to 15.0,Å resolution were calculated by molecular replacement using the known structure of a distantly related homolog (23% sequence identity). Real-space averaging was performed and phases were extended from 15.0 to 3.0,Å. An atomic model was fitted and refined using a simulated-annealing real-space procedure. [source]

    Crystallization and preliminary X-ray analysis of NADP(H)-dependent alcohol dehydrogenases from Saccharomyces cerevisiae and Rana perezi

    ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2003
    Eva Valencia
    Different crystal forms diffracting to high resolution have been obtained for two NADP(H)-dependent alcohol dehydrogenases, members of the medium-chain dehydrogenase/reductase superfamily: ScADHVI from Saccharomyces cerevisiae and ADH8 from Rana perezi. ScADHVI is a broad-specificity enzyme, with a sequence identity lower than 25% with respect to all other ADHs of known structure. The best crystals of ScADHVI diffracted beyond 2.8,Å resolution and belonged to the trigonal space group P3121 (or to its enantiomorph P3221), with unit-cell parameters a = b = 102.2, c = 149.7,Å, , = 120°. These crystals were produced by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. Packing considerations together with the self-rotation function and the native Patterson map seem to indicate the presence of only one subunit per asymmetric unit, with a volume solvent content of about 80%. ADH8 from R. perezi is the only NADP(H)-dependent ADH from vertebrates characterized to date. Crystals of ADH8 obtained both in the absence and in the presence of NADP+ using polyethylene glycol and lithium sulfate as precipitants diffracted to 2.2 and 1.8,Å, respectively, using synchrotron radiation. These crystals were isomorphous, space group C2, with approximate unit-cell parameters a = 122, b = 79, c = 91,Å, , = 113° and contain one dimer per asymmetric unit, with a volume solvent content of about 50%. [source]

    Alliin lyase (alliinase) from garlic (Allium sativum): crystallization and preliminary X-ray characterization

    ACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2002
    Linda J. W. Shimon
    The enzyme alliinase has been isolated from garlic bulbs and crystallized. The crystals belong to space group P21, with unit-cell parameters a = 70.191, b = 127.006, c = 108.085,Å, , = 93.384°. They diffract to 2.2,Å at liquid-nitrogen temperature. Analysis of the Patterson self-rotation function suggests that the crystals contain two dimeric molecules per asymmetric unit. [source]

    The structure of l -rhamnulose-1-phosphate aldolase (class II) solved by low-resolution SIR phasing and 20-fold NCS averaging

    ACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2002
    Markus Kroemer
    The enzyme l -rhamnulose-1-phosphate aldolase catalyzes the reversible cleavage of l -rhamnulose-1-phosphate to dihydroxyacetone phosphate and l -­lactaldehyde. It is a homotetramer with an Mr of 30,000 per subunit and crystallized in space group P3221. The enzyme shows a low sequence identity of 18% with the structurally known l -fuculose-1-­phosphate aldolase that splits a stereoisomer in a similar reaction. Structure analysis was initiated with a single heavy-atom derivative measured to 6,Å resolution. The resulting poor electron density, a self-rotation function and the working hypothesis that both enzymes are C4 symmetric with envelopes that resemble one another allowed the location of the 20 protomers of the asymmetric unit. The crystal-packing unit was a D4 -symmetric propeller consisting of five D4 -­symmetric octamers around an internal crystallographic twofold axis. Presumably, the propellers associate laterally in layers, which in turn pile up along the 32 axis to form the crystal. The non-crystallographic symmetry was used to extend the phases to the 2.7,Å resolution limit and to establish a refined atomic model of the enzyme. The structure showed that the two enzymes are indeed homologous and that they possess chemically similar active centres. [source]

    Crystallization and preliminary X-ray diffraction analysis of Pseudomonas aeruginosa phosphorylcholine phosphatase

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010
    Lisandro H. Otero
    Pseudomonas aeruginosa phosphorylcholine phosphatase (PchP) catalyzes the hydrolysis of phosphorylcholine to produce choline and inorganic phosphate. Phosphorylcholine is released by the action of haemolytic phospholipase C (PlcH) on phosphatidylcholine or sphingomyelin. PchP belongs to the HAD superfamily and its activity is dependent on Mg2+, Zn2+ or Cu2+. The possible importance of PchP in the pathogenesis of P. aeruginosa, the lack of information about its structure and its low identity to other members of this family led us to attempt its crystallization in order to solve its three-dimensional structure. Crystals of the protein have been grown and diffraction data have been obtained to 2.7,Å resolution. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 137.16, b = 159.15, c = 73.31,Å, , = 117.89°. Statistical analysis of the unit-cell contents and the self-rotation function suggest a tetrameric state of the molecule with 222 point-group symmetry. [source]

    Purification, crystallization and preliminary crystallographic analysis of very-long-chain acyl-CoA dehydrogenase from Caenorhabditis elegans

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2010
    Zhijie Li
    Acyl-CoA dehydrogenase [acyl-CoA:(acceptor) 2,3-oxidoreductase; EC 1.3.99.3] catalyzes the first reaction step in mitochondrial fatty-acid ,-oxidation. Here, the very-long-chain acyl-CoA dehydrogenase from Caenorhabditis elegans (cVLCAD) has been cloned and overexpressed in Escherichia coli strain BL21 (DE3). Interestingly, unlike other very-long-chain acyl-CoA dehydrogenases, cVLCAD was found to form a tetramer by size-exclusion chromatography coupled with in-line static light-scattering, refractive-index and ultraviolet measurements. Purified cVLCAD (12,mg,ml,1) was successfully crystallized by the hanging-drop vapour-diffusion method under conditions containing 100,mM Tris,HCl pH 8.0, 150,mM sodium chloride, 200,mM magnesium formate and 13% PEG 3350. The crystal has a tetragonal form and a complete diffraction data set was collected and processed to 1.8,Å resolution. The crystal belonged to space group C2, with unit-cell parameters a = 138.6, b = 116.7, c = 115.3,Å, , = , = 90.0, , = 124.0°. A self-rotation function indicated the existence of one noncrystallographic twofold axis. A preliminary molecular-replacement solution further confirmed the presence of two molecules in one asymmetric unit, which yields a Matthews coefficient VM of 2.76,Å3,Da,1 and a solvent content of 55%. [source]

    Crystallization and preliminary X-ray analysis of AAMS amidohydrolase, the final enzyme in degradation pathway I of pyridoxine

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2009
    Jun Kobayashi
    ,-(N -Acetylaminomethylene)succinic acid (AAMS) amidohydrolase from Mesorhizobium loti MAFF303099, which is involved in a degradation pathway of vitamin B6 and catalyzes the degradation of AAMS to acetic acid, ammonia, carbon dioxide and succinic semialdehyde, has been overexpressed in Escherichia coli. To elucidate the reaction mechanism based on the tertiary structure, the recombinant enzyme was purified and crystallized by the sitting-drop vapour-diffusion method using PEG 8000 as precipitant. A crystal of the enzyme belonged to the monoclinic space group C2, with unit-cell parameters a = 393.2, b = 58.3, c = 98.9,Å, , = 103.4°, and diffraction data were collected to 2.7,Å resolution. The VM value and calculation of the self-rotation function suggested that three dimers with a threefold symmetry were possibly present in the asymmetric unit. [source]

    Crystallization and preliminary X-ray analysis of the human respiratory syncytial virus nucleocapsid protein

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2008
    K. El Omari
    Human respiratory syncytial virus (HRSV) has a nonsegmented negative-stranded RNA genome which is encapsidated by the HRSV nucleocapsid protein (HRSVN) that is essential for viral replication. HRSV is a common cause of respiratory infection in infants, yet no effective antiviral drugs to combat it are available. Recent data from an experimental anti-HRSV compound, RSV-604, indicate that HRSVN could be the target site for drug action. Here, the expression, purification and preliminary data collection of decameric HRSVN as well as monomeric N-terminally truncated HRSVN mutants are reported. Two different crystal forms of full-length selenomethionine-labelled HRSVN were obtained that diffracted to 3.6 and ,5,Å resolution and belonged to space group P212121, with unit-cell parameters a = 133.6, b = 149.9, c = 255.1,Å, and space group P21, with unit-cell parameters a = 175.1, b = 162.6, c = 242.8,Å, , = 90.1°, respectively. For unlabelled HRSVN, only crystals belonging to space group P21 were obtained that diffracted to 3.6,Å. A self-rotation function using data from the orthorhombic crystal form confirmed the presence of tenfold noncrystallographic symmetry, which is in agreement with a reported electron-microscopic reconstruction of HRSVN. Monomeric HRSVN generated by N-terminal truncation was designed to assist in structure determination by reducing the size of the asymmetric unit. Whilst such HRSVN mutants were monomeric in solution and crystallized in a different space group, the size of the asymmetric unit was not reduced. [source]

    Expression, purification and preliminary X-ray characterization of dl -2-haloacid dehalogenase from Methylobacterium sp.

    ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2007

    dl -2-Haloacid dehalogenase from Methylobacterium sp. CPA1 (dl -DEX Mb) is a unique enzyme that catalyzes the dehalogenation reaction without the formation of an ester intermediate. A recombinant form of dl -DEX Mb has been expressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. The crystal belongs to the hexagonal space group P63, with unit-cell parameters a = b = 186.2, c = 114.4,Å. The crystals are likely to contain between four and eight monomers in the asymmetric unit, with a VM value of 4.20,2.10,Å3,Da,1. A self-rotation function revealed peaks on the , = 180° section. X-ray data have been collected to 1.75,Å resolution. [source]