X-ray Structure of Gelonin at 1.8 Å Resolution

Gelonin is a single chain ribosome inactivating protein (RIP) with potential application in the treatment of cancer and AIDS. Diffraction quality crystals grown using PEG3350, belong to the space group P2(1), with it a = 49.4 Angstrom b = 44.9 Angstrom, c = 137.4 Angstrom and beta = 98.4 degrees, and contain two molecules in the asymmetric unit. Diffraction data collected to 1.8 Angstrom resolution has a R(m) value of 7.3%. Structure of gelonin has been solved by the molecular replacement method, using ricin A chain as the search model. Crystallographic refinement using X-PLOR resulted in a model for which the r.m.s deviations from ideal bond lengths and bond angles are 0.012 Angstrom and 2.7 degrees, respectively The final R-factor is 18.4% for 39,806 reflections for which I > 1.0 sigma(I).The C-alpha atoms of the two molecules in the asymmetric unit superpose to within 0.38 Angstrom for 247 atom pairs. The overall fold of gelonin is similar to that of other RIPs such as ricin A chain and alpha-momorcharin, the r.m.s.d. for C-alpha superpositions being 1.3 and 1.4 Angstrom, respectively The-catalytic residues (Glu166, Arg169 and Tyr113) in the active site form a hydrogen bond scheme similar to that observed in other RIPs. The conformation of Tyr74 in the active site, however, is significantly different from that in alpha-momorcharin. Three well defined water molecules are located in the active site cavity and one of them, X319, superposes to within 0.2 Angstrom of a corresponding water molecule in the structure of alpha-momorcharin. Any of the three could be the substrate water molecule in the hydrolysis reaction catalysed by gelonin.Difference electron density for a N-linked sugar moiety has been observed near only one of the two potential glycosylation sites in the sequence. The amino acid at position 239 has been established as Lys by calculation of omit electron density maps.The two cysteine residues in the sequence, Cys44 and Cys50, form a disulphide bond, and are therefore not available for disulphide conjugation with antibodies. Based on the structure, the region of the molecule that is involved in intradimer interactions is suggested to be suitable for introducing a Cys residue for purposes of conjugation with an antibody to produce useful immunotoxins.

Pt metal-CeO2 interaction: Direct observation of redox coupling between Pt-0/Pt2+/Pt4+ and Ce4+/Ce3+ states in Ce0.98Pt0.02O2-delta catalyst by a combined electrochemical and x-ray photoelectron spectroscopy study

Pt ions-CeO2 interaction in Ce1-xPtxO2-delta (x=0.02) has been studied for the first time by electrochemical method combined with x-ray diffraction and x-ray photoelectron spectroscopy. Working electrodes made of CeO2 and Ce0.98Pt0.02O2-delta mixed with 30% carbon are treated electrochemically between 0.0-1.2 V in potentiostatic (chronoamperometry) and potentiodynamic (cyclic voltametry) mode with reference to saturated calomel electrode. Reversible oxidation of Pt-0 to Pt2+ and Pt4+ state due to the applied positive potential is coupled to simultaneous reversible reduction of Ce4+ to Ce3+ state. CeO2 reduces to CeO2-y (y=0.35) after applying 1.2 V, which is not reversible; Ce0.98Pt0.02O2-delta reaches a steady state with Pt2+:Pt4+ in the ratio of 0.60:0.40 and Ce4+:Ce3+ in the ratio of 0.55:0.45 giving a composition Ce0.98Pt0.02O1.74 at 1.2 V, which is reversible. Composition of Pt ion substituted compound is reversible between Ce0.98Pt0.02O1.95 to Ce0.98Pt0.02O1.74 within the potential range of 0.0-1.2 V. Thus, Ce0.98Pt0.02O2-delta forms a stable electrode for oxidation of H2O to O-2 unlike CeO2. A linear relation between oxidation of Pt2+ to Pt4+ with simultaneous reduction in Ce4+ to Ce3+ is observed demonstrating Pt-CeO2 metal support interaction is due to reversible Pt-0/Pt2+/Pt4+ interaction with Ce4+/Ce3+ redox couple.

Polymorphism in B-DNA - X-ray-Diffraction Studies on Li-DNA Fibers

From X-ray diffraction studies it is generally believed that B-DNA has the structural parameters n = 10 and h = 3.4 Å. However, for the first time we report that polymorphism in the B-form can be observed in DNA fibres. This was achieved by the precise control of salt and humidity in fibres and by the application of the precession method of X-ray diffraction to DNA fibres. The significant result obtained is that n = 10 is not observed for crystalline fibre patterns. In fact, n = 10 and h = 3.4 Å are not found to occur simultaneously. Instead, a range of values, n = 9.6–10.0 and h = 3.35 Å–3.41 Å is observed.

x-ray studies on crystalline complexes involving amino-acids and peptides .10. head-to-tail sequences in the crystal-structure of l-lysine acetate

l-Lysine acetate crystallises in the monoclinic space group P21 with a = 5.411 (1), b = 7.562(1), c= l2.635(2) Å and β = 91.7(1). The crystal structure was solved by direct methods and refined to an R value of 0.049 using the full matrix least squares method. The conformation and the aggregation of lysine molecules in the structure are similar to those found in the crystal structure of l-lysine l-aspartate. A conspicuous similarity between the crystal structures of l-arginine acetate and l-lysine acetate is that in both cases the strongly basic side chain, although having the largest pK value, interacts with the weakly acidic acetate group leaving the α-amino and the α-carboxylate groups to take part in head-to-tail sequences. These structures thus indicate that electrostatic effects are strongly modulated by other factors so as to give rise to head-to-tail sequences which have earlier been shown to be an almost universal feature of amino acid aggregation in the solid state.

Study of strontium compounds and minerals by X-ray absorption spectroscopy. I. K-edge shifts

The chemical shifts in the X-ray K-absorption edge of strontium in various compounds and in six minerals are measured using a single crystal X-ray spectrometer. Besides valence, the shifts are found to be governed by ionic charges on the absorbing ions, which are calculated employing Pauling's method. For the minerals the plot of chemical shift against the theoretically calculated ionic charges is used to determine the charges on the strontium ions.

X-Ray Structure Of A Ternary Metal-Complex - Copper(Ii) Inosine 5'-Monophosphate Imidazole - Metal-Binding To The N(7) Atom Of The Nucleotide In A Mixed-Ligand System Containing An Aromatic Amine

A ternary metal-nucleotide complex, Na2[Cu(5’-IMP)2(im)o,8(H20)l,2(H20)2h]as~ 1be2e.n4 pHr2ep0a,r ed and its structure analyzed by X-ray diffraction (5’-IMP = inosine 5’-monophos hate; im = imidazole). The complex crystallizes in space group C222, with a = 8.733 (4) A, b = 23.213 (5) A, c = 21.489 (6) 1, and Z = 4. The structure was solved by the heavy-atom method and refined by full-matrix least-squares technique on the basis of 2008 observed reflections to a final R value of 0.087. Symmetry-related 5’-IMP anions coordinate in cis geometry through the N(7) atoms of the bases. The other cis positions of the coordination plane are statistically occupied by nitrogen atoms of disordered im groups and water oxygens with occupancies 0.4 and 0.6, respectively. Water oxygens in axial positions complete the octahedral coordination of Cu(I1). The complex is isostructural with C~S-[P~(S’-IMP),(NH~)~a] m”,o del proposed for Pt(I1) binding to DNA. The base binding observed in the present case is different from the typical ”phosphate only” binding shown from earlier studies on metal-nucleotide complexes containing various other ?r-aromatic amines.

Ternary metal complexes of anionic and neutral pyridoxine (vitamin B6) with 2,2'-bipyridine. Syntheses and x-ray structures of (pyridoxinato)bis(2,2'-bipyridyl)cobalt(III) perchlorate and chloro(2,2'-bipyridyl)(pyridoxine)copper(II) perchlorate hydrate

Ternary metal complexes involving vitamin B6 with formulas [CO",(PN-H)](anCdI [OC)'(bpy)(PN)Cl]C10(.bpHy 0 = 2,2'-bipyridine, PN = neutral pyridoxine, PN-H = anionic pyridoxine) have been prepared for the first time and characterized by means of magnetic and spectroscopic measurements. The crystal structures of the compounds have also been determined. [CO(PN-H)](CcryIsOta,l)lize s in the space group P2,/c with a = 18.900 (3) A, b = 8.764 (1) A, c = 20.041 (2) A,p = 116.05 (l)', and Z = 4 and [Cu(bpy)(PN)C1]C104-H20in the space group Pi with a = 12.136 (5) A, b = 13.283 (4) A,c = 7.195 (2) A, a = 96.91 (Z)', 0 = 91.25 (3)', y = 71.63 (3)', and Z = 2. The structures were solved by the heavy-atom method and refined by least-squares techniques to R values of 0.080 and 0.042 for 3401 and 2094 independent reflections, respectively. Both structures consist of monomeric units. The geometry around Co(II1) is octahedral and around Cu(I1) is distorted square pyramidal. In [CO(PN-H)]t(wCo IoxOy~ge)n~s ,fro m phenolic and 4-(hydroxymethyl) groups of PN-H and two nitrogens from each of two bpy's form the coordination sphere. In [Cu(bpy)(PN)C1]C104.H20o ne PN and one bpy, with the same donor sites, act as bidentate chelates in the basal plane, with a chloride ion occupying the apical position. In both structures PN and PN-H exist in the tautomeric form wherein pyridine N is protonated and phenolic 0 is deprotonated. However, a novel feature of the cobalt compound is that PN-H is anionic due to the deprotonation of the 4-(hydroxymethyl) group. The packing in both structures is governed by hydrogen bonds, and in the copper compound partial stacking of bpy's at a distance of -3.55 also adds to the stability of the system. Infrared, NMR, and ligand field spectroscopic results and magnetic measurements are interpreted in light of the structures.

Patient exposure monitoring and radiation qualities in two-dimensional digital x-ray imaging

The methods for estimating patient exposure in x-ray imaging are based on the measurement of radiation incident on the patient. In digital imaging, the useful dose range of the detector is large and excessive doses may remain undetected. Therefore, real-time monitoring of radiation exposure is important. According to international recommendations, the measurement uncertainty should be lower than 7% (confidence level 95%). The kerma-area product (KAP) is a measurement quantity used for monitoring patient exposure to radiation. A field KAP meter is typically attached to an x-ray device, and it is important to recognize the effect of this measurement geometry on the response of the meter. In a tandem calibration method, introduced in this study, a field KAP meter is used in its clinical position and calibration is performed with a reference KAP meter. This method provides a practical way to calibrate field KAP meters. However, the reference KAP meters require comprehensive calibration. In the calibration laboratory it is recommended to use standard radiation qualities. These qualities do not entirely correspond to the large range of clinical radiation qualities. In this work, the energy dependence of the response of different KAP meter types was examined. According to our findings, the recommended accuracy in KAP measurements is difficult to achieve with conventional KAP meters because of their strong energy dependence. The energy dependence of the response of a novel large KAP meter was found out to be much lower than with a conventional KAP meter. The accuracy of the tandem method can be improved by using this meter type as a reference meter. A KAP meter cannot be used to determine the radiation exposure of patients in mammography, in which part of the radiation beam is always aimed directly at the detector without attenuation produced by the tissue. This work assessed whether pixel values from this detector area could be used to monitor the radiation beam incident on the patient. The results were congruent with the tube output calculation, which is the method generally used for this purpose. The recommended accuracy can be achieved with the studied method. New optimization of radiation qualities and dose level is needed when other detector types are introduced. In this work, the optimal selections were examined with one direct digital detector type. For this device, the use of radiation qualities with higher energies was recommended and appropriate image quality was achieved by increasing the low dose level of the system.

Electronic excitations in solids studied using inelastic x-ray scattering

Inelastic x-ray scattering can be used to study the electronic structure of matter. The x rays scattered from the target both induce and carry information on the electronic excitations taking place in the system. These excitations are the manifestations of the electronic structure and the physics governing the many-body system. This work presents results of non-resonant inelastic x-ray scattering experiments on a range of materials including metallic, insulating and semiconducting compounds as well as an organic polymer. The experiments were carried out at the National Synchrotron Light Source, USA and at the European Synchrotron Radiation Facility, France. The momentum transfer dependence of the experimental valence- and core-electron excitation spectra is compared with the results of theoretical first principles computations that incorporate the electron-hole interaction. A recently developed method for analyzing the momentum transfer dependence of core-electron excitation spectra is studied in detail. This method is based on real space multiple scattering calculations and is used to extract the angular symmetry components of the local unoccupied density of final states.

Studies of electronic structure by x-ray Raman and emission spectroscopy: applications to MgB2 superconductors and high pressure physics

X-ray Raman scattering and x-ray emission spectroscopies were used to study the electronic properties and phase transitions in several condensed matter systems. The experimental work, carried out at the European Synchrotron Radiation Facility, was complemented by theoretical calculations of the x-ray spectra and of the electronic structure. The electronic structure of MgB2 at the Fermi level is dominated by the boron σ and π bands. The high density of states provided by these bands is the key feature of the electronic structure contributing to the high critical temperature of superconductivity in MgB2. The electronic structure of MgB2 can be modified by atomic substitutions, which introduce extra electrons or holes into the bands. X ray Raman scattering was used to probe the interesting σ and π band hole states in pure and aluminum substituted MgB2. A method for determining the final state density of electron states from experimental x-ray Raman scattering spectra was examined and applied to the experimental data on both pure MgB2 and on Mg(0.83)Al(0.17)B2. The extracted final state density of electron states for the pure and aluminum substituted samples revealed clear substitution induced changes in the σ and π bands. The experimental work was supported by theoretical calculations of the electronic structure and x-ray Raman spectra. X-ray emission at the metal Kβ line was applied to the studies of pressure and temperature induced spin state transitions in transition metal oxides. The experimental studies were complemented by cluster multiplet calculations of the electronic structure and emission spectra. In LaCoO3 evidence for the appearance of an intermediate spin state was found and the presence of a pressure induced spin transition was confirmed. Pressure induced changes in the electronic structure of transition metal monoxides were studied experimentally and were analyzed using the cluster multiplet approach. The effects of hybridization, bandwidth and crystal field splitting in stabilizing the high pressure spin state were discussed. Emission spectroscopy at the Kβ line was also applied to FeCO3 and a pressure induced iron spin state transition was discovered.

Preliminary X-ray crystallographic analysis of 2-methylcitrate synthase from Salmonella typhimurium

Analysis of the genomic sequences of Escherichia coli and Salmonella typhimurium has revealed the presence of several homologues of the well studied citrate synthase (CS). One of these homologues has been shown to code for 2-methylcitrate synthase (2-MCS) activity. 2-MCS catalyzes one of the steps in the 2-methylcitric acid cycle found in these organisms for the degradation of propionate to pyruvate and succinate. In the present work, the gene coding for 2-MCS from S. typhimurium (StPrpC) was cloned in pRSET-C vector and overexpressed in E. coli. The protein was purified to homogeneity using Ni-NTA affinity chromatography. The purified protein was crystallized using the microbatch-under-oil method. The StPrpC crystals diffracted X-rays to 2.4 A resolution and belonged to the triclinic space group P1, with unit-cell parameters a = 92.068, b = 118.159, c = 120.659 A, alpha = 60.84, beta = 67.77, gamma = 81.92 degrees. Computation of rotation functions using the X-ray diffraction data shows that the protein is likely to be a decamer of identical subunits, unlike CSs, which are dimers or hexamers.

Cloning, expression, purification, crystallization and preliminary X-ray crystallographic study of the putative SAICAR synthetase (PH0239) from Pyrococcus horikoshii OT3

The study of proteins involved in de novo biosynthesis of purine nucleotides is central in the development of antibiotics and anticancer drugs. In view of this, a protein from the hyperthermophile Pyrococcus horikoshii OT3 was isolated, purified and crystallized using the microbatch method. Its primary structure was found to be similar to that of SAICAR synthetase, which catalyses the seventh step of de novo purine biosynthesis. A diffraction-quality crystal was obtained using Hampton Research Crystal Screen II condition No. 34, consisting of 0.05 M cadmium sulfate hydrate, 0.1 M HEPES buffer pH 7.5 and 1.0 M sodium acetate trihydrate, with 40%(v/v) 1,4-butanediol as an additive. The crystal belonged to space group P3(1), with unit-cell parameters a = b = 95.62, c = 149.13 angstrom. Assuming the presence of a hexamer in the asymmetric unit resulted in a Matthews coefficient (V-M) of 2.3 angstrom(3) Da(-1), corresponding to a solvent content of about 46%. A detailed study of this protein will yield insights into structural stability at high temperatures and should be highly relevant to the development of antibiotics and anticancer drugs targeting the biosynthesis of purine nucleotides.

X-Ray structure of a ternary complex, copper(II)–lnosine 5-monophosphate–benzimidazole. The first example for the absence of direct metal–nucleotide interaction

The unprecedented absence of direct metal–nucleotide interaction has been observed in the X-ray structure of the ternary metal nucleotide system [Cu(bzim)(H2O)5]2+[IMP]2–·3H2O [IMP = inosine 5-monophosphate(2–), bzim = benzimidazole). The complex crystallizes in the space group P21 with a= 7.013(2), b= 13.179(9), c= 14.565(9)Å, = 94.82(4)°, and Z= 2. The structure was solved by the heavy-atom method and refined by full-matrix least squares on the basis of 1 761 observed (I? 3i) reflections to final R and R values of 0.034 and 0.036 respectively. The CuII has a distorted octahedral co-ordination with a nitrogen of the bzim ligand [Cu–N 1.947(5)Å] and three oxygens of water molecules in the basal plane [mean Cu–O 2.017(3)Å] and two more water oxygens at axial positions [Cu–O 2.194(6) and 2.732(5)Å]. The nucleotide base stacks with the bzim ligand at an average distance of 3.5 Å and an angle of 22°. In the lattice, N(7) of the base is linked to a lattice water through a hydrogen bond, while all the phosphate oxygens are involved in hydrogen bonds with co-ordinated as well as lattice water molecules. The co-ordination behaviour of IMP to CuII is compared in structures containing different -aromatic amines in order to assess the influence of the ternary ligand in complex formation. The present results indicate that, apart from the commonly observed phosphate binding, other modes of co-ordination are possible, these being influenced mainly by the -accepting properties of the ternary ligand.

Interaction of metal ions with 6-oxopurine nucleotides. Part 1. X-Ray structures of ternary cobalt(III) complexes with inosine 5'-monophosphate or guanosine 5'-monophosphate

The crystal structures of two ternary metal nucleotide complexes of cobalt, [Co(en)2(H2O)2]-[Co(5?-IMP)2(H2O)4]Cl2·4H2O (1) and [Co(en)2(H2O)2][Co(5?-GMP)2(H2O)4]Cl2·4H2O (2), have been analysed by X-ray diffraction (en = ethylenediamine, 5?-IMP = inosine 5?-monophosphate, and 5?-GMP = guanosine 5?-monophosphate). Both complexes crystallize in the orthorhombic space group C2221 with a= 8.725(1), b= 25.891(5), c= 21.212(5)Å, Z= 4 for (1) and a= 8.733(2), b= 26.169(4), c= 21.288(4)Å, Z= 4 for (2). The structure of (1) was solved by the heavy-atom method, while that of (2) was deduced from (1). The structures were refined to R values of 0.09 and 0.10 for 1 546 and 1 572 reflections for (1) and (2) respectively. The two structures are isomorphous. A novel feature is that the chelate ligand en and the nucleotide are not co-ordinated to the same metal ion. One of the metal ions lying on the two-fold a axis is octahedrally co-ordinated by two chelating en molecules and two water oxygens, while the other on the two-fold b axis is octahedrally co-ordinated by two N(7) atoms of symmetry-related nucleotides in a cis position and four water oxygens. The conformations of the nucleotides are C(2?)-endo, anti, and gauche�gauche. In both (1) and (2) the charge-neutralising chloride ions are disordered in the vacant space between the molecules. These structures bear similarities to the mode of nucleotide co-ordination to PtII complexes of 6-oxopurine nucleotides, which are the proposed models for intrastrand cross-linking in DNA by a metal complex.

Physico-chemical properties of pure and x-ray irradiated single crystals of KAP

Single crystals of potassium hydrogen phthalate (KAP) have been grown by slow evaporation method from aqueous solutions. Thermal analyses indicate that KAP crystals decompose into phthalic anhydride and KOH around 520 K. Electrical properties of single crystals of KAP have been studied along with the effect of X-ray irradiation of the crystals. The electrical transport appears to be associated with tunneling of protons. The irradiated crystal exhibits lower dielectric constant and higher ac conductivity.