Preliminary X-ray study of crystals of human C-reactive protein

Two different crystal forms of human C-reactive protein have been grown from solutions of 2-methyl-2,4-pentanediol. Both crystal forms are tetragonal, the space group for form I is P4(1)22 (or P4(3)22), and that for form II is P4(2)22. The unit cell parameters for form I are a = b = 103.0(5) A, c = 308.5(7) A and for form II are a = b = 103.1(2) A, c = 312.7(6) A. The crystals of form II diffract to at least 3.0 A resolution, and are suitable for detailed structural studies.     

Crystallization and preliminary X-ray studies of psophocarpin B1, a chymotrypsin inhibitor from winged bean seeds

Psophocarpin B1 is a 20,000 Mr protein of winged bean (Psophocarpus tetragonolobus) seeds having chymotrypsin inhibitory activity. Single crystals of this protein suitable for X-ray crystallographic studies have been obtained by the vapour diffusion method using ammonium sulphate. The crystals are hexagonal, space group P6(4)22 or P6(2)22, cell dimensions a = b = 61 A, c = 210 A. They are stable to irradiation with X-rays and diffract to at least 2.6 A resolution.     

Preliminary X-ray crystallographic study of lysozyme produced by Streptomyces globisporus

Lysozyme from Streptomyces globisporus has been crystallized in a form suitable for X-ray structure analysis using ammonium sulfate as a precipitant. The crystals are hexagonal, space group P6(1)22 (P6(5)22) with unit cell dimensions: a = b = 129 A, c = 143 A. There are three or four molecules per asymmetric unit. The crystals diffract X-rays to at least 3.0 A resolution.     

Crystallization of mouse pancreatic ribonuclease

Mouse pancreatic ribonuclease has been crystallized in a form suitable for X-ray structure determination. The crystals grown from solutions of 2-methyl-2,4-pentanediol diffract to high resolution and belong to the hexagonal space group P6(1) (P6(5)) with unit cells dimensions a = b = 64.44 A, c = 53.91 A, y = 120 degrees and V = 1.94 x 10(5) A3 (1 A = 0.1 nm). There are six molecules per unit cell (1 molecule/asymmetric unit), and Vm = 2.3 A3/dalton.     

Crystallization and preliminary X-ray diffraction studies of the lectin from Erythrina corallodendron

The lectin from Erythrina corallodendron, specific for N-acetyllactosamine, crystallizes in the hexagonal space group P6(1) (P6(5)) with unit cell dimensions a = b = 136.3 A, c = 83.2 A and one dimer of Mr 60,000 in the asymmetric unit. The crystals are suitable for high-resolution work.     

钙敏感受体基因、白介素6基因与中国人群骨密度相关联

钙敏感受体是钙新陈代谢的一个重要因素,白介素6是参与破骨细胞分化及功能的一种多效细胞因子。因此,钙敏感受体基因和白介素6基因都为骨矿物质代谢的重要候选基因。本研究旨在利用数量性状传递不平衡检测法,检测白介素6基因和钙敏感受体基因与腰椎和髋部骨密度的关联和连锁,以证实它们是否为影响中国人群骨密度的重要候选基因。本研究的样本为来自中国上海的401个中国核心家庭,共1,263个个体,均为汉族。每个核心家庭由父母双亲和至少一个20～45岁的健康绝经前女儿组成。腰椎与髋部的骨密度采用Hologic QDR 2000＋双能X射线扫描仪进行了检测。用PE377测序仪及相关软件分别对白介素6和钙敏感受体基因中的一个CA重复多态微卫星位点进行了基因分型。分析结果表明钙敏感受体基因（CA）12等位基因（P=0．006）及（Ca）18等位基因（P=0．02）与股骨颈骨密度之间存在显著的整体关联。白介素6基因的（CA）18等位基因与整个髋部（P=0．021）、股骨颈（P=0．041）以及转子间区（P=0．029）骨密度之间均存在显著的家庭内关联。白介素6基因（CA）n位点与腰椎BMD之间存在显著的连锁（P=0．001）。本研究结果表明白介素6基因和钙敏感受体基因可能为与中国人群骨密度变异相关联的候选基因。     

Crystallization and preliminary X-ray studies of glutathione synthetase from Escherichia coli B

The glutathione synthetase from Escherichia coli B has been crystallized from 27% saturated ammonium sulfate solution (pH 5.5). The crystals are hexagonal, space group P6(2)22 or P6(4)22. The cell dimensions are a = b = 88.0 A, c = 164.2 A, and gamma = 120 degrees. The enzyme is a tetramer (Mr = 143,000) with 222 symmetry, and the asymmetric unit contains one subunit molecule (Mr = 35,600). The crystals diffract to at least 2.5 A resolution.     

Crystallographic characterization of recombinant human CuZn superoxide dismutase

Recombinant human CuZn superoxide dismutase as expressed in yeast has been crystallized in three different crystal forms. Hexagonal plates grow from 2.4 M ammonium sulfate, pH 7.5, and belong to the space group P6(3)22, with cell dimensions a = b = 113.5(3), c = 151.5(5) A, and Vm = 2.21 A3/dalton for two dimers per asymmetric unit. At 2.0 M ammonium sulfate, pH 7.5, chunky wedges grow in space group C222(1), a = 205.2(6), b = 166.5(4), c = 145.4(4) A with a Vm of 2.43 A3/dalton for eight dimers per asymmetric unit. With polyethylene glycol 8000, pH 7.5-8.0, hexagonal prisms are obtained with cell dimensions a = b = 197.4(6), c = 43.1(2) A, space group P6, and Vm = 2.53 A3/dalton for three dimers per asymmetric unit. All of these forms diffract to high resolution, are stable to x-rays, and appear suitable for determination of the atomic structure. Crystals of the doubly mutated enzyme (Cys6----Ala, Cys111----Ser) grown from both micro- and macroseeds of the wild type protein demonstrate the feasibility of isomorphous crystallization of site-directed mutants of the cloned parent enzyme for comparative structure-function studies.     

Crystallization of an anti-2,2,6,6-tetramethyl-1-piperidinyloxy-dinitrophenyl monoclonal antibody Fab fragment with and without bound hapten

The Fab fragment of a monoclonal antibody, AN02, specific for a 2,2,6,6-tetramethyl-1-piperidinyloxy-dinitrophenyl hapten was crystallized both with and without bound hapten. Both crystals formed in phosphate-buffered saline (150 mM-NaCl, 10 mM-Na2PO4, 0.02% (w/v) NaN3 (pH 7.4) at 4 degrees C and diffracted beyond 2.2 A resolution (1A = 0.1 nm). Fab with bound hapten crystallizes in space group P6(1)22 or P6(5)22, with cell dimensions a = b = 73.23 A, c = 373.8 A, alpha = beta = 90 degrees and gamma = 120 degrees. Unoccupied Fab also crystallizes in space group P6(1)22 or P6(5)22 with cell dimensions a = b = 73 A, c = 377 A, alpha = beta = 90 degrees and gamma = 120 degrees.     

Crystallization studies of cAMP-dependent protein kinase. Cocrystals of the catalytic subunit with a 20 amino acid residue peptide inhibitor and MgATP diffract to 3.0 A resolution.

Crystallographic studies of the catalytic subunit of cAMP-dependent protein kinase demonstrate that the presence of a 20 amino acid residue peptide inhibitor and MgATP during crystallization yields crystals with a different space group and, more significantly, makes an important difference in the quality of the resulting crystals. Under identical experimental conditions, the kinase crystallizes in a cubic space group P4(1)32 (a = b = c = 169.24 A), when no substrates or inhibitors are present, and in the hexagonal space group P6(1)22 (or P6(5)22) (a = b = 80.16 A, c = 288.07 A, alpha = beta = 90 degrees, gamma = 120 degrees) when a 20-amino acid residue peptide inhibitor and MgATP are present. Moreover, the hexagonal crystal diffracts to a resolution of 3.0 A, while the cubic crystals diffract to a resolution of 4.0 A.     

Personality endophenotypes for bipolar affective disorder: a family-based genetic association analysis

Genetic analyses of complex conditions such as bipolar disorder (BD) may be facilitated by the use of intermediate phenotypes. Various personality traits are overrepresented in people with BD and their unaffected relatives, and may constitute genetically transmitted risk factors or endophenotypes of the illness. In this study, we administered a battery of seven different personality questionnaires comprising 19 subscales to 31 Caucasian BD families (n = 241). Ten of these personality traits showed significant evidence of heritability and were therefore selected as candidate endophenotypes. In addition, a principal components analysis produced two heritable components (negative affect and appetitive drive), which accounted for a considerable proportion of the variance (29% + 12%) and were also used in the analysis. A family-based quantitative association study was carried out using the orthogonal model from the quantitative transmission disequilibrium tests (QTDT) program. Monte Carlo permutations (M = 500), which allow for non-normal data and provide a global P value, corrected for multiple testing, were used to calculate empirical P values for the within-family component of association. The 3' untranslated region repeat polymorphism of the dopamine transporter gene (SLC6A3) was associated with self-directedness (P < 0.0001) and negative affect (P = 0.010). The short allele of the serotonin transporter gene (SLC6A4) promoter polymorphism showed a trend toward association with higher harm avoidance (P = 0.016) and negative affect (P = 0.028). The catechol-o-methyltransferase val158met polymorphism was weakly associated with the personality traits, 'Spirituality' (P = 0.040) and irritable temperament (P = 0.022). Furthermore, the met allele of the brain-derived neurotrophic factor val66met polymorphism was associated with higher hyperthymic temperament scores. We raise the possibility that the 10R allele of the SLC6A3 repeat polymorphism and the short allele of the SLC6A4 promoter variant constitute risk factors for irritable-aggressive and anxious-dysthymic subtypes of BD, respectively.     

Crystallization of phosphatidylinositol-specific phospholipase C from Bacillus cereus.

Phosphatidylinositol-specific phospholipase C (PI-PLC) cleaves phosphoinositides into two parts, lipid-soluble diacylglycerol and the water-soluble phosphorylated inositol. Two crystal forms of Bacillus cereus PI-PLC have been obtained by the vapor diffusion technique. Hexagonal crystals were grown from solutions containing polyethylene glycol (PEG; 4,000 to 8,000 D). The space group of these hexagonal crystals is P6(1)22 (or the enantiomorphic space group P6(5)22), with cell constants a = b = 133 A, and c = 231 A. The crystals diffract to 2.8 A. The second crystalline form was grown from a two-phase PEG (600 D)-sodium citrate solution. The phase diagram and PI-PLC distribution between phases has been determined. The enzyme crystallizes from the PEG-rich phase. The crystals are orthorhombic with space group P2(1)2(1)2(1) (a = 45 A, b = 46 A, c = 160 A), and contain one PI-PLC monomer per asymmetric unit. The orthorhombic crystals diffract to 2.5 A. Both the hexagonal and orthorhombic forms are suitable for crystallographic studies.     

Crystallization and preliminary X-ray analysis of a fungal endoglucanase I.

An endoglucanase I (EG1) from a fungal source (Humicola insolens) has been crystallized in a number of forms suitable for X-ray diffraction analysis. Four crystal forms have been grown from various precipitants using vapour phase diffusion methods in hanging drops. Three of these crystal forms diffract to beyond 2.5 A resolution. Two forms, obtained from ammonium sulphate at pH 5.4, or 8.0, grow as tetragonal bipyramids in space group P4(1)22 or P4(3)22, with approximate cell dimensions a = b = 102 A, c = 282 A. The other crystal forms were grown from polyethylene glycol 8000 at pH 8.0. One grows as monoclinic plates, space group P2(1), with cell dimensions a = 66.9 A, b = 75.2 A, c = 86.9 A and beta = 102.9 degrees and the other as long hexagonal rods in space group P6(1)22 or P6(5)22, with cell dimensions a = b = 119 A, c = 83 A.     

Peptidic phosphonylating agents as irreversible inhibitors of serine proteases and models of the tetrahedral intermediates

Peptide analogues incorporating an electrophilic phosphorus moiety (2-6) have been synthesized and studied as inhibitors of a variety of serine proteases. Inhibition is irreversible and, for alpha-lytic protease (ALP), shown to result from covalent binding to the active site serine hydroxyl [Bone, R., Sampson, N. S., Bartlett, P.A., & Agard, D. A. (1991) Biochemistry (following paper in this issue)]. For reaction of human leukocyte elastase (HLE) with the thiophenyl esters 6s-V (Boc-AAPV psi [P = O(SPh)O]AA-OMe), 4s-V (BocAAPV psi [P = O(SPh)O]-Me), and 3s-V (Boc-V psi [P = O(SPh)O]AA-OMe), evidence is presented to suggest that the S4-S1 subsites, but not the S1' and S2' positions, are occupied by the inhibitors during the inactivation process. The selectivity that is observed between the proteases and the hexapeptide phosphonates 6o-V (Boc-AAPV psi [P = O(OPh)O]AA-OMe) and 6o-F (Boc-AAPF psi [P = O(OPh)O]AA-OMe) parallels that between these enzymes and their substrates: ALP and HLE are selectively inactivated by the ValP-containing analogue 6o-V, while subtilisin (SUB) shows a preference for the PheP derivative 6o-F. A detailed kinetic analysis of the enzyme-inhibitor interactions was complicated by the susceptibility of the inhibitors to enzymatic degradation. The configuration at phosphorus was found not to have a significant influence on the rate at which the inhibitors react with the peptidases. Moreover, in the case of inactivation of ALP by the hexapeptide 6o-V, the same covalent adduct is formed from both stereoisomers (Bone et al., 1991), indicating that one of these diastereomers undergoes substitution with retention of configuration.     

Preliminary crystallographic study of the Erythrina rubrinervia lectin

The galactose-specific lectin from Erythrina rubrinervia crystallizes in the hexagonal space group P6, (or P6(5)) with unit cell dimensions a = b = d = 135.1 A, c = 83.0 A. These parameters are compatible with the presence of a dimer with Mr = 60,000 in the asymmetric unit. The crystals are suitable for high-resolution X-ray analysis.     

六种云杉的核型分析

对６种（含１变种）云杉属植物的染色体数目及形态特征进行了分析研究。研究树种均为我国特有种,其中４种在本文中首次报道。分析材料全部取自各树种中心产区的原始林内。其核型公式：鳞皮云杉（Ｐ．ｒｅｔｒｏｆｌｅｘａ）Ｋ（２ｎ）＝２４＝２２ｍ＋２ｓｍ;云极（Ｐ．ａｓｐｅｒａｔａ）Ｋ（２ｎ）＝２４＝２０ｍ＋４ｓｍ;青插（Ｐ．ｗｉｌｓｏｎｉｉ）Ｋ（２ｎ）＝２４＝２０ｍ＋４ｓｍ＋１Ｂ;紫果云极（Ｐ．ｐｕｒｐｕｒｅａ     

Preliminary crystallographic analysis of the breakage-reunion domain of the Escherichia coli DNA gyrase A protein

The 64 x 10(3) Mr N-terminal breakage-reunion domain of the Escherichia coli DNA gyrase A protein was purified from an over-expressing strain. When complexed with the gyrase B protein, this truncated A protein has all of the enzymic properties of the full-length counterpart, although with reduced efficiency in some cases. The 64 x 10(3) Mr protein has been crystallized in several forms, a number of which were too small for crystallographic analysis. However, two forms grew to sufficient size for preliminary X-ray analysis. Both forms were tetragonal with a primitive lattice. One form (type I) had cell dimensions of a = b = 170 A, c = 145 A a space group of either P41212 (P43212) or P42212, and diffracted to 6 A resolution. The type II crystals had cell dimensions of a = b = 177 A, c = 175 A, a space group of P41212 (P43212) or P42212, and diffracted to at least 4.5 A resolution. Both crystal forms apparently contained four subunits (possibly a tetramer) in the asymmetric unit. We are attempting to increase the size and quality of these crystals.     

Crystallization and preliminary X-ray analysis of the cAMP-dependent protein kinase catalytic subunit from Saccharomyces cerevisiae

A truncated variant of TPK1, the yeast cAMP-dependent protein kinase catalytic subunit, was overexpressed in an engineered strain of Saccharomyces cerevisiae, purified by liquid chromatography, and crystallized from solutions of 2-propanol and magnesium at alkaline pH. The crystals are hexagonal dipyramids, space group P6(1)22 (P6(5)22), with unit-cell parameters a = b = 61 A, c = 320 A. Large single crystals suitable for diffraction analysis are obtainable by microseeding, and diffract beyond 2.8-A resolution. Crystal density measurements reveal 12 kinase monomers per unit cell with a single kinase monomer per asymmetric unit.     

HIV-1 Nef protein: purification, crystallizations, and preliminary X-ray diffraction studies.

Human immunodeficiency virus Nef protein accelerates virulent progression of AIDS by its interaction with specific cellular proteins involved in cellular activation and signal transduction. Here we report the purification and crystallization of the conserved core of HIV-1LAI Nef protein in the unliganded form and in complex with the wild-type SH3 domain of the P59fyn protein-tyrosine kinase. One-dimensional NMR experiments show that full-length protein and truncated fragment corresponding to the product of HIV-1 protease cleavage have a well-folded compact tertiary structure. The ligand-free HIV-1 Nefcore protein forms cubic crystals belonging to space group P23 with unit cell dimensions of a = b = c = 86.4 A. The Nef-Fyn SH3 cocrystals belong to the space group P6(1)22 or its enantiomorph, P6(5)22, with unit cell dimensions of a = b = 108.2 A and c = 223.7 A. Both crystal forms diffract to a resolution limit of 3.0 A resolution using synchrotron radiation, and are thus suitable for X-ray structure determination.     

New packing of B-DNA in crystals]

Two crystal forms of the self-complementary tetramer GpGpCpC have been obtained by phase diagram technique: P6(2)22/P6(4)22. a = b = 67.7 A, c = 105.6 A and P3(2)12/P3(1)12, a = b = 116.9 A. c = 116.4 A. Both crystals form diffract at least up to 3.2 A. Diffraction patterns of both crystal forms have strongest base-stacking reflections corresponding to the Bragg spacing 3.38 A which is typical for B-DNA. Moreover the self-rotation function of the first crystal form shows regular located two-fold pseudo-axes periodicity of which also indicates that this is B-conformation. The same conclusion can be reached on the basis of the crystal packing of the duplexes in the unit cell. It should be emphasized that this is a new example of B-DNA crystal packing.