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Perdeuteration and in vitro refolding of hen egg-white lysozyme impact protein thermal stability and activity. Deuteration appears to primarily affect enzymatic function through changes in protein dynamics, while refolding contributes to a small decrease in protein thermal stability., This structural and biophysical study exploited a method of perdeuterating hen egg-white lysozyme based on the expression of insoluble protein in Escherichia coli followed by in-column chemical refolding. This allowed detailed comparisons with perdeuterated lysozyme produced in the yeast Pichia pastoris, as well as with unlabelled lysozyme. Both perdeuterated variants exhibit reduced thermal stability and enzymatic activity in comparison with hydrogenated lysozyme. The thermal stability of refolded perdeuterated lysozyme is 4.9°C lower than that of the perdeuterated variant expressed and secreted in yeast and 6.8°C lower than that of the hydrogenated Gallus gallus protein. However, both perdeuterated variants exhibit a comparable activity. Atomic resolution X-ray crystallographic analyses show that the differences in thermal stability and enzymatic function are correlated with refolding and deuteration effects. The hydrogen/deuterium isotope effect causes a decrease in the stability and activity of the perdeuterated analogues; this is believed to occur through a combination of changes to hydrophobicity and protein dynamics. The lower level of thermal stability of the refolded perdeuterated lysozyme is caused by the unrestrained Asn103 peptide-plane flip during the unfolded state, leading to a significant increase in disorder of the Lys97–Gly104 region following subsequent refolding. An ancillary outcome of this study has been the development of an efficient and financially viable protocol that allows stable and active perdeuterated lysozyme to be more easily available for scientific applications.

A new algorithm automatically determines the structures of large macromolecules of unknown fold from low-resolution single-wavelength anomalous X-ray data and a partial model that failed with other methods., Determining macromolecular structures from X-ray data with resolution worse than 3 Å remains a challenge. Even if a related starting model is available, its incompleteness or its bias together with a low observation-to-parameter ratio can render the process unsuccessful or very time-consuming. Yet, many biologically important macromolecules, especially large macromolecular assemblies, membrane proteins and receptors, tend to provide crystals that diffract to low resolution. A new algorithm to tackle this problem is presented that uses a multivariate function to simultaneously exploit information from both an initial partial model and low-resolution single-wavelength anomalous diffraction data. The new approach has been used for six challenging structure determinations, including the crystal structures of membrane proteins and macromolecular complexes that have evaded experts using other methods, and large structures from a 3.0 Å resolution F1-ATPase data set and a 4.5 Å resolution SecYEG–SecA complex data set. All of the models were automatically built by the method to R free values of between 28.9 and 39.9% and were free from the initial model bias.

The metallonuclease colicin E7 is a member of the HNH family of endonucleases. It serves as a bacterial toxin in Escherichia coli, protecting the host cell from other related bacteria and bacteriophages by degradation of their chromosomal DNA under environmental stress. Its cell-killing activity is attributed to the nonspecific nuclease domain (NColE7), which possesses the catalytic ββα-type metal ion-binding HNH motif at its C-terminus. Mutations affecting the positively charged amino acids at the N-terminus of NColE7 (444–­576) surprisingly showed no or significantly reduced endonuclease activity [Czene et al. (2013 ▶), J. Biol. Inorg. Chem. 18, 309–321]. The necessity of the N-­terminal amino acids for the function of the C-terminal catalytic centre poses the possibility of allosteric activation within the enzyme. Precise knowledge of the intramolecular interactions of these residues that affect the catalytic activity could turn NColE7 into a novel platform for artificial nuclease design. In this study, the N-terminal deletion mutant ΔN4-NColE7-C* of the nuclease domain of colicin E7 selected by E. coli was overexpressed and crystallized at room temperature by the sitting-drop vapour-diffusion method. X-ray diffraction data were collected to 1.6 A resolution and could be indexed and averaged in the trigonal space group P3121 or P3221, with unit-cell parameters a = b = 55.4, c = 73.1 A. Structure determination by molecular replacement is in progress.

X-ray diffraction data of high quality measured to high resolution on crystals of the two pentitol epimers ribitol (centric) and xylitol (acentric) at 101, 141, and 181 K and data on the two compounds previously recorded at 122 K have formed the basis for multipole refinements with the VALRAY system. Our analysis showed that it is possible to obtain a reliable crystal electron density for an acentric compound (xylitol) from X-ray diffraction data and that the thermal motion can be deconvoluted from the static density in this temperature range. The Bader-type topological analysis of the static electron densities revealed virtually identical intramolecular interactions as well as very similar hydrogen bond interactions of ribitol and xylitol; the only minor differences are found in the weaker intermolecular interactions. The high-level periodic DFT calculations are in accordance with the thermodynamic measurements that show that the two pentitols have identical sublimation energies. A rigid body normal coordinate analysis was performed on the atomic displacement parameters obtained at the four different temperatures. The translational and librational mean square deviations derived through this analysis were used in a quantum statistical approach to derive frequencies of the corresponding harmonic oscillators. The analysis showed a consistent vibrational model for all temperatures. The frequencies were subsequently used to calculate crystal entropies assuming an Einstein-type behavior. These calculations show that the crystal entropy of ribitol is 8 J K(-1) mol(-1) higher than the crystal entropy of xylitol, confirming that it is a difference in the entropy of the two compounds that causes the difference in their free energy. Our results presented in this Article show the potential to use X-ray diffraction data to obtain physicochemical properties of crystals.

Laccases of different biological origins have been widely investigated and these studies have elucidated fundamentals of the generic catalytic mechanism. However, other features such as surface properties and residues located away from the catalytic centres may also have impact on enzyme function. Here we present the crystal structure of laccase from Myceliophthora thermophila (MtL) to a resolution of 1.62 Å together with a thorough structural comparison with other members of the CAZy family AA1_3 that comprises fungal laccases from ascomycetes. The recombinant protein produced in A. oryzae has a molecular mass of 75 kDa, a pI of 4.2 and carries 13.5 kDa N-linked glycans. In the crystal, MtL forms a dimer with the phenolic substrate binding pocket blocked, suggesting that the active form of the enzyme is monomeric. Overall, the MtL structure conforms with the canonical fold of fungal laccases as well as the features specific for the asco-laccases. However, the structural comparisons also reveal significant variations within this taxonomic subgroup. Notable differences in the T1-Cu active site topology and polar motifs imply molecular evolution to serve different functional roles. Very few surface residues are conserved and it is noticeable that they encompass residues that interact with the N-glycans and/or are located at domain interfaces. The N-glycosylation sites are surprisingly conserved among asco-laccases and in most cases the glycan displays extensive interactions with the protein. In particular, the glycans at Asn88 and Asn210 appear to have evolved as an integral part of the asco-laccase structure. An uneven distribution of the carbohydrates around the enzyme give unique properties to a distinct part of the surface of the asco-laccases which may have implication for laccase function–in particular towards large substrates.

The short hydrogen bonds in rhamnogalacturonan acetylesterase have been investigated by structure determination of an active-site mutant, 1H NMR spectra and computational methods. Comparisons are made to database statistics. A very short carboxylic acid carboxylate hydrogen bond, buried in the protein, could explain the low-field (18 p.p.m.) 1H NMR signal., An extremely low-field signal (at approximately 18 p.p.m.) in the 1H NMR spectrum of rhamnogalacturonan acetylesterase (RGAE) shows the presence of a short strong hydrogen bond in the structure. This signal was also present in the mutant RGAE D192N, in which Asp192, which is part of the catalytic triad, has been replaced with Asn. A careful analysis of wild-type RGAE and RGAE D192N was conducted with the purpose of identifying possible candidates for the short hydrogen bond with the 18 p.p.m. deshielded proton. Theor­etical calculations of chemical shift values were used in the interpretation of the experimental 1H NMR spectra. The crystal structure of RGAE D192N was determined to 1.33 Å resolution and refined to an R value of 11.6% for all data. The structure is virtually identical to the high-resolution (1.12 Å) structure of the wild-type enzyme except for the interactions involving the mutation and a disordered loop. Searches of the Cambridge Structural Database were conducted to obtain information on the donor–acceptor distances of different types of hydrogen bonds. The short hydrogen-bond inter­actions found in RGAE have equivalents in small-molecule structures. An examination of the short hydrogen bonds in RGAE, the calculated pK a values and solvent-accessibilities identified a buried carboxylic acid carboxylate hydrogen bond between Asp75 and Asp87 as the likely origin of the 18 p.p.m. signal. Similar hydrogen-bond interactions between two Asp or Glu carboxy groups were found in 16% of a homology-reduced set of high-quality structures extracted from the PDB. The shortest hydrogen bonds in RGAE are all located close to the active site and short interactions between Ser and Thr side-chain OH groups and backbone carbonyl O atoms seem to play an important role in the stability of the protein structure. These results illustrate the significance of short strong hydrogen bonds in proteins.

Anisotropic displacement parameters (ADPs) are compared for H atoms estimated using three recently described procedures, both among themselves and with neutron diffraction results. The results convincingly demonstrate that all methods are capable of giving excellent results for several benchmark systems and identify systematic discrepancies for several atom types. A revised and extended library of internal H-atom mean-square displacements is presented for use with Madsen's SHADE web server [J. Appl. Cryst. (2006), 39, 757–758; http://shade.ki.ku.dk], and the improvement over the original SHADE results is substantial, suggesting that this is now the most readily and widely applicable of the three approximate procedures. Using this new library – SHADE2 – it is shown that, in line with expectations, a segmented rigid-body description of the heavy atoms yields only a small improvement in the agreement with neutron results. The SHADE2 library, now incorporated in the SHADE web server, is recommended as a routine procedure for deriving estimates of H-atom ADPs suitable for use in charge-density studies on molecular crystals, and its widespread use should reveal remaining deficiencies and perhaps overcome the inherent bias in the majority of such studies.

The syntheses of the square-planar platinum(II) complexes [Pt(L)(tba)] (L = 5,5′-dichloro-2,2′-dihydroxyazobenzenate (dhab), (5-chloro-2-hydroxyphenylazo)-3-oxo-N-phenylbutanamidate (hpab) or (5-chloro-2-hydroxyphenylazo)-2-naphtholate (hpan) and (tba) = tributylamine) is reported, together with those of the complete set of analogous complexes [M(L)(py)] (M = nickel(II), palladium(II), platinum(II) and (py) = pyridine). The coordinating nitrogen has been assigned to the azo-nitrogen attached to the 5-chloro-2-hydroxyphenyl substituent in both [Ni(hpab)(py)] and [Pt(hpan)(tba)] by single crystal X-ray diffraction methods. It has been established that complexes with different group 10 metals are iso-structural by isomorphology of the crystals as determined by X-ray powder diffraction studies on the complexes containing pyridine in the fourth coordination site. Furthermore, we present a method to determine the coordination geometry by comparison of calculated 13C chemical shifts for possible coordination modes optimized by ab initio methods with experimentally measured 13C chemical shifts.

Tri-t-butylgallium has been reacted with the macrocycle 1,12-diaza-3,4:9,10-dibenzo-5,8-dioxocyclopentadecane which could be a potential ligand for gallium(III). A reaction product was analyzed and single crystal X-ray diffraction experiments showed that it consisted of the cyclic dimer of di-t-butylgallium-hydroxide hydrogen bonded to the macrocycle. Without a co-crystallized organic molecule di-t-butylgallium hydroxide crystallizes as a trimer. Density functional calculations have been used to predict the structures and the total energies for the monomer, dimer, trimer, and tetramer of dimethylgallium hydroxide in order to provide a basis for the understanding of oligomer population for the dimethylgallium hydroxides. Force field calculations are shown to be able to produce a similar strain energy difference for dimer, trimer, and tetramer forms of (CH3)2Ga(OH) and this method can economically be used for larger alkyl groups. The force field computations show that the trimeric di-t-butyl gallium(III) hydroxide is much more stable than the dimeric form which therefore must owe its existence to the association with the hydrogen-bonded macrocycle.

Extensive and precise X-ray diffraction data for xylitol have been used to test different approaches to estimate nuclear parameters for H atoms in charge-density studies. The parameters from a neutron diffraction study of the same compound were taken as a reference. The resulting static charge densities obtained for the different approaches based on a multipole model were subjected to a topological analysis. The comparative analysis led to the following results. The procedure of extending the X-H bond to match bond lengths from neutron diffraction studies provides the best agreement with the neutron positional parameters. An isotropic model for the atomic displacements of H atoms is highly unsatisfactory and leads to significant deviations for the properties of the bond critical points including those that only involve non-H atoms. Anisotropic displacement parameters for H atoms can be derived from the X-ray data that are in agreement with the values from the neutron study, and the resulting charge-density models are in good agreement with the reference model. The anisotropic displacement parameters for H atoms are derived from the X-ray data as a sum of the external (rigid-body) and internal vibrations. The external vibrations are obtained from a TLS analysis of the ADPs of the non-H atoms and the internal vibrations from analysis of neutron diffraction studies of related compounds. The results from the analysis of positional and thermal parameters were combined to devise a 'best anisotropic' model, which was employed for three other systems where X-ray and neutron data were available. The results from the topological analysis of these systems confirm the success of the 'best anisotropic' model in providing parameters for the H atoms that give charge densities in agreement with the reference models based on H-atom parameters derived from neutron diffraction.

The glycoprotein rhamnogalacturonan acetylesterase from Aspergillus aculeatus has been crystallized in two crystal forms, an orthorhombic and a trigonal crystal form. In the orthorhombic crystal form, the covalently bound carbohydrate at one of the two N-glycosylation sites is involved in crystal contacts. The orthorhombic crystal form was obtained at pH 5.0 and the trigonal crystal form at pH 4.5. In one case, the two crystal forms were found in the same drop at pH 4.7. The differences in crystal packing in the two crystal forms can be explained by the pH-dependent variation in the protonation state of the glutamic acid residues on the protein surface.

Recombinant rhamnogalacturonan lyase from Aspergillus aculeatus has been crystallized by a stepwise procedure and X-ray diffraction data have been collected. The crystals were grown using hanging-drop vapour-diffusion and microseeding techniques. Crystals were obtained showing a flat plate morphology. The crystallization conditions were 20% PEG 4000, 9% PEG 400, 0.1 M (NH(4))(2)SO(4) and 0.1 M sodium acetate pH 4.4. These crystals diffracted to a resolution of 1.5 A. The unit-cell parameters are a = b = 77.0, c = 170.8 A with the possible space group P4(3)2(1)2 or P4(1)2(1)2. There is most likely to be one molecule in the asymmetric unit, leading to a calculated solvent content of approximately 47% for the crystals.

The crystal structure of Thermoascus aurantiacus endoglucanase (Cel5A), a family 5 glycoside hydrolase, has been determined to 1.62 Å resolution by multiple isomorphous replacement with anomalous scattering. It is the first report of a structure in the subfamily to which Cel5A belongs. Cel5A consists solely of a catalytic module with compact eight-fold β/α barrel architecture. The length of the tryptophan-rich substrate binding groove suggests the presence of substrate binding subsites −4 to +3. Structural comparison shows that two glycines are completely conserved in the family, in addition to the two catalytic glutamates and six other conserved residues previously identified. Gly 44 in particular is part of a type IV C-terminal helix capping motif, whose disruption is likely to affect the position of an essential conserved arginine. One aromatic residue (Trp 170 in Cel5A), not conserved in term of sequence, is nonetheless spatially conserved in the substrate binding groove. Its role might be to force the bend that occurs in the polysaccharide chain on binding, thus favoring substrate distortion at subsite −1.

The nuclease domain of colicin E7 metallonuclease (NColE7) contains its active centre at the C-terminus. The mutant ΔN4-NColE7-C∗-where the four N-terminal residues including the positively charged K446, R447 and K449 are replaced with eight residues from the GST tag-is catalytically inactive. The crystal structure of this mutant demonstrates that its overall fold is very similar to that of the native NColE7 structure. This implicates the stabilizing effect of the remaining N-terminal sequence on the structure of the C-terminal catalytic site and the essential role of the deleted residues in the mechanism of the catalyzed reaction. Complementary QM/MM calculations on the protein-DNA complexes support the less favourable cleavage by the mutant protein than by NColE7. Furthermore, a water molecule as a possible ligand for the Zn2+-ion is proposed to play a role in the catalytic process. These results suggest that the mechanism of the Zn2+-containing HNH nucleases needs to be further studied and discussed.

The structure of the dihydroxyphenylselenonium ion (C_{6}H_{7}O_{2}Se^{+}) has been determined in its benzenesulfonate (C_{6}H_{5}O_{3}Se^{-}) and p-toluenesulfonate (C_{7}H_{7}O_{3}S ^{-}) salts. Whereas the former salt is disordered, the latter less dense salt is well defined. This difference in crystallization behaviour is attributed to a C—H...O hydrogen bond involving the methyl group of the p-toluenesulfonate ion. The two salts display very similar hydrogen-bond arrangements and differ only with respect to the stacking of the phenyl groups. The dihydroxyselenonium ion is a strong acid with a pK value of −0.9 determined from the variation of the 77Se chemical shift. A comparison with the two deprotonated species reveals a systematic increase in the Se—O bond lengths and the pyramidal configuration around Se with the number of protons attached.

Syntheses and characterisation by H-1 NMR and IR of the platinum(II) complexes [PtI2(Ph2C=NH)(2)], [PtCl2(Ph2C=NH)(2)], [PtCl(Ph(Ph-H)C=NH)-(NH2CH2CH2NH2)] and trans-(N,N)-[PtCl(Ph(Ph-H)C=NH)(Ph2C=NH)] are described. Absorption and emission spectra at room temperature for the latter complex are reported. The crystal structure of trans-(N,N)[PtCl(Ph(Ph-H)C=NH)(Ph2C=NH)] . 1/2{(CH3)(2)CO} was determined by X-ray diffraction methods. Space group C2/c, a = 14.318(4) Angstrom, b = 22.854(4) Angstrom. c = 15.212(3) Angstrom and beta = 108.42(2)degrees using 10 368 reflections in the refinement of 291 parameters gave R = 0.037 and wR2 = 0.074 (for all data). The ligands surround platinum in a planar configuration with bond lengths of Pt-Cl 2.404(1) Angstrom, Pt-N (monodentate imine ligand) 2.004(3) Angstrom, Pt-N (bidentate imine ligand) 1.979(3) Angstrom and Pt-C 1.988(3) Angstrom. The chemical shifts in the H-1 NMR spectrum of trans-(N,N)-[PtCl(Ph(Ph-H)C=NH)(Ph2C=NH)] (in CDCl3) display variations with concentration of the complex that indicate dimerisation. The dimerisation constant was determined from the change in chemical shift for the NH proton of the ortho-metalated ligand, K = 1.25(4) M-1 at 300 K. The structure of the dimer in solution is proposed to resemble one of the types of interactions that are encountered between platinum complexes in the solid state.

Two independent ion pairs [(6-methoxy-4-quinolyl)(5-vinyl-1 -azoniabicyclo[2.2.2]octan-2-yl)methanol hydrogen 2,3-dihydroxybutanedioate] and a water molecule are found in the asymmetric unit of 2C 20 H 25 N 2 O 2 + .-2C 4 H 5 O - 6 .H 2 O. The two cations are virtually identical, but the two anions have markedly different stereochemistries. One of these anions adopts a unique conformation not observed previously for hydrogen tartrate ions. The packing resembles the arrangement in cinchonidinium (S)-mandelate, with hydrogen-bonded chains of alternating cations and anions. The herring-bone stacking of the quinoline ring systems of the cations resembles the pattern seen in other cinchona structures.

The title compound, [N(C4H9)4+]2.[Ni(C8H4N2S2)2]2−, crystallizes with the Ni atom on a crystallographic inversion center. The phenyl rings of the ligands make an angle of 50.50 (5)° with the plane spanned by the central nickel-sulfur fragment. The bond lengths do not indicate that the formal negative charge on the dithiocarbimato moiety is delocalized into the phenyl ring.

The experimental crystal charge density of deuterated methylammonium hydrogen maleate has been determined from neutron and X-ray diffraction data collected at 122.4 K. Refinements of the neutron diffraction data showed that the very short intramolecular O−H−O hydrogen bonds found in the two independent anions are symmetric with D(H) on crystallographic mirror planes. The experimental charge density was subjected to a topological analysis. It is positive throughout the unit cell and constitutes a topological space that fulfills the Poincare−Hopf relation. The topological analysis enabled us to characterize different types of interatomic interactions in the crystal and showed that the two very short intramolecular hydrogen bonds have covalent character. Newly developed algorithms made it possible to determine the atomic basins in the crystal charge density, which were used to calculate integrated properties such as volume, charge, and the integrated Laplacian. Those of the methylammonium ion have been compa...

Diffraction data recorded at low temperature with Mo K\alpha from two different crystals have been used in the refinement of the structure of the acid rhodo complex salt, \mu-hydroxo-bis[pentaamminechromium(III)] chloride monohydrate [(NH3)5CrOHCr(NH3)5]Cl5.H2O. Several structural models in two different tetragonal space groups have been investigated before a satisfactory description of the structure was obtained. The compound crystallizes in the orthorhombic space group P212121, with a= 16.155 (3), b = 16.154 (3), c = 14.750 (7) Å and Z = 8. The apparent tetragonal symmetry arises due to rotation twinning of the crystals. The diffraction pattern is further complicated by tetragonal pseudo-symmetry of the chromium sites, which means that reflections with l = 2n + 1 are systematically weak. The two independent dimeric cations are involved in slightly different hydrogen-bond patterns. This observation and the relative orientation of the dimers in the crystal is in accordance with spectral and magnetic measurements.

A series of methylcobalt(III) compounds with solely classical ligands such as NH3, H2O, NO2-, CN-, and 1,2-ethanediamine (en) has been prepared by ligand substitution of the pentaamminemethylcobalt(III) cation. The new compounds have been characterized by 1H, 13C, and 59Co nuclear magnetic resonance spectroscopy and by absorption spectroscopy. Single-crystal X-ray structure determinations at 122.0(5) K were performed on [Co(NH3)5(CH3)]S2O6 (1), trans-[Co(en)2(NH3)(CH3)]S2O6 (2), and [Co(NH3)6]-mer,trans-[Co(NO2)3(NH3)2(CH3)]2-trans-[Co(NO2)4(NH3)2] (3). Compound 1 is monoclinic, P21/n, with cell dimensions a = 6.4856(9) A, b = 18.0720(11) A, c = 9.7854(14) A, and β = 98.280(13)°. Compound 2 is orthorhombic, P21212, with cell dimensions a = 12.211(3) A, b = 10.107(3) A, and c = 11.514(3) A. Compound 3 is monoclinic, P21/n, with cell dimensions a = 8.390(2) A, b = 8.970(3) A, c = 20.650(9) A, and β = 91.29(2)°. Virtually identical Co−C distances in the range 1.980−1.984 A are found in the three compounds. T...

A novel Emericella nidulans endo-β-1,4-galactanase (EnGAL) demonstrates a strong capacity to generate high levels of very potent prebiotic oligosaccharides from potato pulp, a by-product of the agricultural potato-starch industry. EnGAL belongs to glycoside hydrolase family 53 and shows high (72.5%) sequence identity to an endo-β-1,4-galactanase from Aspergillus aculeatus. Diffraction data extending to 2.0 A resolution were collected from a crystal of EnGAL grown from conditions containing 0.2 M zinc acetate. The crystal structure showed a high similarity between EnGAL and other endo-β-1,4-galactanases belonging to GH53. It also revealed 15 zinc ions bound to the protein, one of which is located in the active site, where it is coordinated by residues Glu136 and Glu246 which comprise the catalytic machinery. The majority of the zinc ions are located on the surface of the enzyme, in some cases with side chains from two different molecules as ligands, thus explaining why the presence of zinc ions was essential for crystallization.

Lactococcus lactis is the only organism known to contain two dihydroorotate dehydrogenases, i.e., the A- and B-forms. In this paper, we report the overproduction, purification, and crystallization of dihydroorotate dehydrogenase A. In solution, the enzyme is bright yellow. It is a dimer of subunits (34 kDa) that contain one molecule of flavin mononucleotide each. The enzyme shows optimal function in the pH range 7.5-9.0. It is specific for L-dihydroorotate as substrate and can use dichlorophenolindophenol, potassium hexacyanoferrate (III), and, to a lower extent, also molecular oxygen as acceptors of the reducing equivalents, whereas the pyridine nucleotide coenzymes (NAD+, NADP+) and the respiratory quinones (i.e., vitamins Q6, Q10 and K2) were inactive. The enzyme has been crystallized from solutions of 30% polyethylene glycol, 0.2 M sodium acetate, and 0.1 M Tris-HCl, pH 8.5. The resulting yellow crystals diffracted well and showed little sign of radiation damage during diffraction experiments. The crystals are monoclinic, space group P21 with unit cell dimensions a = 54.19 A, b = 109.23 A, c = 67.17 A, and beta = 104.5 degrees. A native data set has been collected with a completeness of 99.3% to 2.0 A and an Rsym value of 5.2%. Analysis of the solvent content and the self-rotation function indicates that the two subunits in the asymmetric unit are related by a noncrystallographic twofold axis perpendicular to the crystallographic b and c axes.

The crystal structures of racemic and optically active cis-3-hydroxy-2-(4-methoxyphenyl)-2,3-dihydro-1,5-benzothiazepin-4(5H)-one (C16H15NO3S) were determined to investigate the enantiomer associations formed in the solid state. X-ray diffraction data were collected with Cu Kα ( λ = 1.54184 A ) radiation at 122.0(5) K. Optically active: monoclinic, space group P21, with a = 7.8605(8), b = 16.148(2), c = 11.3477(11) A , β = 97.607(8)°, V = 1427.7(3) A 3 , Z = 4, Dx = 1.402 g cm−3, μ = 20.53 cm−1 refined to R = 0.0282 for 2922 observed reflections. Racemic: monoclinic, space group P 2 1 n , with a = 10.3322(14), b = 7.7110(11), c = 18.901(3) A , β = 99.781(13)°, V = 1484.0(4) A 3 , Z = 4, Dx = 1.349 g cm−3, μ = 19.75 cm−1, refined to R = 0.0277 for 2909 observed reflections. In both structures the strongest intermolecular interactions are NH⋯O and OH⋯O hydrogen bonds formed between molecules of identical chirality. The racemate and enantiomer crystal structures differ in that there are two different conformers in the crystal structure of the optically pure compound, in the symmetry relation between molecules connected by strong hydrogen bonds, and in the formation of weak CH⋯O hydrogen bonds. The structure determinations do not support the formation of the hydrogen-bonded dimers that had been proposed to cause the difference in NMR chemical shifts of the methoxy protons.

The copper(II), nickel(II) and cobalt(II) complexes of new S 2 N 2 ligands on Schiff bases of 4-benzoyl-3-methyl-1-phenyl-2-pyrazoline-5-thione and various diamines were prepared and characterized. Using different diamines it is possible to control the geometry around the central metal atom. An X-ray structure determination of a Cu(II) S 2 N 2 complex showed the (NCuS, NCuS) dihedral angle to be 52.11° and the following distances: CuS1 2.2274, CuS2 2.2089, CuN13 1.973, CuN23 1.998 A.

Crystals of rac-bis(1,2-ethanediamine)(2-thiooxamato-N,O)cobalt(III) trifluoromethanesulfonate monohydrate, rac-[Co(en)(2)(NHC(S)COO)]CF3SO3 . H2O, showed a reversible phase transition at 155(2) K. Diffraction data were collected al two temperatures. At 110(5) K (LT) the space group is monoclinic P2(1)/n a=6.1509(13), b=14.872(2), c=17.421(3) Angstrom, beta=93.71(2)degrees, V=1590.3(8) Angstrom(3), Z=4. Also at 295.5(5) K (RT) the space group is monoclinic P2(1)/n, a=12.558(5), b=14.978(3), c=18.139(3) Angstrom, beta=104.44(2)degrees, V=3304(3) Angstrom(3), Z=8. The major structural changes associated with the phase transition relate to the hydrogen bonding patterns. The geometries of the two independent complex cations in the RT structure and the cation of the LT structure are very similar. Two fluorine atoms of a trifluoromethanesulfonate anion in the RT structure display static disorder. The maximum variations between corresponding bond lengths and angles in the two structures amount to 0.06 Angstrom and 3 degrees, respectively.

The crystal structures have been determined for racemic and two optically active samples of 3-(2'-chloro-2'-phenylethyl)-2-thiazolidiniminium p-toluenesulfonate (C 11 H 14 ClN 2 S + .C 7 H 7 O 3 S - ) from low-temperature (122 K) X-ray diffraction data. The three crystal structures are virtually identical. The racemate crystallizes in the space group P2 1 /n, Z=4, with unit-cell dimensions a=8.5016 (10), b=8.2803 (11), c=27.447 (3) A and β=96.478 (9) o . The cation displays disorder of the Cl atom and the C atom to which it is bonded. The disorder can be rationalized in terms of the presence of two partially populated enantiomers with different conformations

The structures of the three monofluoro-substituted mandelic acids (C 8 H 7 FO 3 , M r =170.14) have been determined from low-temperature [122 (1) K] X-ray diffraction data [λ(Cu Kα)=1.54184 A]. o-Fluoromandelic acid, monoclinic, P2 1 /c, a=8.4238 (12), b=5.4766 (7), c=15.959 (2) A, β=95.962 (11) o , V=732.3 (3) A 3 , Z=4, D x =1.543 g cm -3 , μ=11.25 cm -1 , F(000)=352, R=0.040 for 1357 contributing reflections, m.p. 388.3 (5) K. m-Fluoromandelic acid (metastable modification), monoclinic, P2 1 /a, a=10.8657 (14), b=9.2663 (10), c=15.722 (2)A, β=107.474 (10)°, V=1509.9 (6)A 3 , Z=8, D x =1.497 g cm -3 , μ=10.92 cm -1 , F(000)=704, R=0.048 for 2977 contributing reflections, m.p. 368.4 (5)K

The title compound crystallizes in the orthorhombic space group C2 with a=39.191, b=5.624, c=11.247A, and β=95.750°, Z=8,. Using 3182 reflections the structure was refined to R=0.027 and R w =0.032. A short asymmetric intramolecular hydrogen bond is observed in both crystallographically independent anions; the two O-O distances are 2.42 and 2.44A. One of the hydrogen maleate ions displays a significant twist of the carboxylic groups relative to the double bond. The two (R)-1-phenylethylammonium ions are virtually identical. The water molecule links one of the cations to one of the anions

Solid-state 13 C and 2 H NMR spectra are found to very suitable for characterizing the short O H...O hydrogen bonds observed in acid salts of dicarboxylic acids. The majority of the investigated compounds are acid salts of malonic, succinic and tartaric acid with monovalent cations derived from alkali metals and small aliphatic amines. They include systems with symmetric and asymmetric hydrogen bonds. Accurate structural information about their geometry is available from low-temperature X-ray diffraction data. The 13 C chemical shifts of the C atoms in the different carboxy groups display a linear variation with the absolute difference between the two C O bond lengths. Theoretical ab initio calculations for model systems showed that the nuclear quadrupole coupling constant NQCC for 2 H increases with increasing asymmetry of the hydrogen-bonded system. NQCC values for 2 H in the short symmetric hydrogen-bonded systems are in the range 53–59 kHz compared with the larger values of up to 166kHz found in systems with longer asymmetric hydrogen bonds. The 2 H NQCC values display a perfect linear dependence on the asymmetry of the hydrogen bond. 2 H NQCC decreases with decreasing temperature in the symmetric hydrogen bonds showing that the corresponding potential has a single well.

Solid-state 13 C and 2 H NMR spectra are found to very suitable for characterizing the short O H...O hydrogen bonds observed in acid salts of dicarboxylic acids. The majority of the investigated compounds are acid salts of malonic, succinic and tartaric acid with monovalent cations derived from alkali metals and small aliphatic amines. They include systems with symmetric and asymmetric hydrogen bonds. Accurate structural information about their geometry is available from low-temperature X-ray diffraction data. The 13 C chemical shifts of the C atoms in the different carboxy groups display a linear variation with the absolute difference between the two C O bond lengths. Theoretical ab initio calculations for model systems showed that the nuclear quadrupole coupling constant NQCC for 2 H increases with increasing asymmetry of the hydrogen-bonded system. NQCC values for 2 H in the short symmetric hydrogen-bonded systems are in the range 53–59 kHz compared with the larger values of up to 166kHz found in systems with longer asymmetric hydrogen bonds. The 2 H NQCC values display a perfect linear dependence on the asymmetry of the hydrogen bond. 2 H NQCC decreases with decreasing temperature in the symmetric hydrogen bonds showing that the corresponding potential has a single well.

Four circle neutron diffraction data were recorded for a single crystal of KMnCl3 cooled to 110 K and 10 K, respectively. Analysis of the data measured at 110 K showed that they correspond to the diffraction from a twinned crystal of α-KMnCl3. The twinning explains the luminescence behaviour of the crystal. At 110 K the cell dimensions corresponding to the orthorhombic space group Pnma are a = 7.05(3) A , b = 9.90(2) A and c = 6.96(1) A . The two grains are oriented with the a-axis from one grain parallel to the c-axis from the other. The relative proportions of the two grains were included in the refinement based on 170 observed reflections to R -values: R = 0.116, Rw = 0.085. The second data set was measured well below the transition temperature (TN = 98 K) where α-KMnCl3 transforms into a simple antiferromagnetic structure. An analysis of the magnetic intensities showed that the magnetic moment is parallel to the crystallographic c-axis and has the value (4.0 ± 0.1)μB in agreement with previous powder work.

Two mutant dCTP deaminase-dUTPases from Methanocaldococcus jannaschii were crystallised and the crystal structures were solved: E145A in complex with the substrate analogue alpha,beta-imido-dUTP and E145Q in complex with diphosphate. Both mutant enzymes were defect in the deaminase reaction and had reduced dUTPase activity. In the structure of E145Q in complex with diphosphate, the diphosphate occupied the same position as the beta- and gamma-phosphoryls of the nucleotide analogue in the E145A complex. The C-terminal region that is unresolved in the apo-form of the enzyme was ordered in both complexes and closed over the active site by interacting with the phosphate backbone of the nucleotide or with the diphosphate. A magnesium ion was readily observed to complex with all three phosphoryls in the nucleotide complex or with the diphosphate. A water molecule that is likely to be involved in the nucleotidyl diphosphorylase reaction was observed in the E145A:alpha,beta-imido-dUTP complex and positioned similarly as in the monofunctional trimeric dUTPase. A comparison of the active sites of the bifunctional enzyme and the monofunctional family members, dCTP deaminase and dUTPase, suggests similar reaction mechanisms. The similar side chain conformations in the deaminase site between the nucleotide and diphosphate complexes indicated a concerted re-arrangement, or induced fit, of the whole active site promoted by enzyme and nucleotide phosphoryl interactions. A pre-steady state kinetic analysis of the bifunctional reaction and the dUTPase half-reaction supported a conformational change upon substrate binding in both reactions and a concerted catalytic step for the bifunctional reaction.

CH 6 N + •C 4 H 5 O 4 − •H 2 O (I) cristallise dans P2 1 /m avec a=4,9788, b=13,077, c=6,133 A, β=92,34°, Z=2, T=110 K; affinement jusqu'a R=0,035. C 2 H 8 N + •C 4 H 5 O 4 − (II) cristallise dans Cmem avec a=6,7863, b=8,069, c=14,284 A, Z=4, T=110 K; affinement jusqu'a R=0,028. Les ions hydrogenosuccinate sont lies par des liaisons hydrogene tres courtes formant des chaines infinies