Click chemistry generated model DNA-peptide heteroconjugates as tools for mass spectrometry.

UV cross-linking of nucleic acids to proteins in combination with mass spectrometry is a powerful technique to identify proteins, peptides, and the amino acids involved in intermolecular interactions within nucleic acid-protein complexes. However, the mass spectrometric identification of cross-linked nucleic acid-protein heteroconjugates in complex mixtures and MS/MS characterization of the specific sites of cross-linking is extremely challenging. As a tool for the optimization of sample preparation, ionization, fragmentation, and detection by mass spectrometry, novel synthetic DNA-peptide heteroconjugates were generated to act as mimics of UV cross-linked heteroconjugates. Click chemistry was employed to cross-link peptides to DNA oligonucleotides. These heteroconjugates were fully characterized by high resolution FTICR mass spectrometry and by collision-induced dissociation (CID) following nuclease P1 digestion of the DNA moiety to a single nucleotide monophosphate. This allowed the exact site of the cross-linking within the peptide to be unambiguously assigned. These synthetic DNA-peptide heteroconjugates have the potential to be of use for a variety of applications that involve DNA-peptide heteroconjugates.

Photoluminescent carbon dots from 1,4-addition polymers.

Photoluminescent carbon dots were synthesised directly by thermopyrolysis of 1,4-addition polymers, allowing precise control of their properties. The effect of polymer composition on the properties of the carbon dots was investigated by TEM, IR, XPS, elemental analysis and fluorescence analysis, with carbon dots synthesised from nitrogen-containing polymers showing the highest fluorescence. The carbon dots with high nitrogen content were observed to have strong fluorescence in the visible region, and culture with cells showed that the carbon dots were non-cytotoxic and readily taken up by three different cell lines.

Anomeric bond-character in the pyranose sugars.

Evidence from determinations of crystal structure relating to the shortening of carbon-oxygen bonds in the anomeric position in pyranose sugars is presented. There is a high degree of probability that the C(1)-O(1) bond is shortened by about 0.04 A relative to the other C-O single-bond lengths, except in the case of an axially oriented glycosidic group where there is evidence of a distinction between the two ring C-O bonds.

Clathrate hydrates of some amines.

A crystallographic study of some alkylamine hydrates establishes that these are polyhedral clathrate hy drates. While the simpler alkylamines form hydrates with water structures re lated to those of the gas hydrates or the alkylonium salt hydrates, the di ethylamine and tert-butylamine hydrates have water frameworks forming cages which have not previously been ob served in the clathrate hydrates.

IDENTIFICATION OF CONOIDIN A AS A COVALENT INHIBITOR OF PEROXIREDOXIN II.

Conoidin A (1) is an inhibitor of host cell invasion by the protozoan parasite Toxoplasma gondii. In the course of studies aimed at identifying potential targets of this compound, we determined that it binds to the T. gondii enzyme peroxiredoxin II (TgPrxII). Peroxiredoxins are a widely conserved family of enzymes that function in antioxidant defense and signal transduction, and changes in PrxII expression are associated with a variety of human diseases, including cancer. Disruption of the TgPrxII gene by homologous recombination had no effect on the sensitivity of the parasites to 1, suggesting that TgPrxII is not the invasion-relevant target of 1. However, we showed that 1 binds covalently to the peroxidatic cysteine of TgPrxII, inhibiting its enzymatic activity in vitro. Studies with human epithelial cells showed that 1 also inhibits hyperoxidation of human PrxII. These data identify Conoidin A as a novel inhibitor of this important class of antioxidant and redox signaling enzymes.

Fluorescent Formazans and Tetrazolium Salts - Towards Fluorescent Cytotoxicity Assays.

Formazan-based colorimetric cytotoxicity assays, such as the MTT assay, are typically used to assess cell viability with only metabolically active cells reducing tetrazolium salts into the formazans, which is then quantified by absorbance. Fluorescence offers several advantages compared to colorimetric assays and would enable techniques such as flow cytometry and confocal microscopy to be used for analysis. Here, fluorescent formazans 10, 11 and 12, and their corresponding tetrazolium salts 13, 16 and 24, respectively, were synthesised by incorporation of a known fluorophore backbone (coumarin, fluorescein and rhodol) with disruption of the conjugated system preventing or reducing fluorescence of the tetrazolium salts. These tetrazolium salts were successfully reduced to the fluorescent formazans with cells and offer a step forward in the development of fluorescent cytotoxicity assays.

The lyotropic liquid crystal properties of n-octyl 1-O-beta-D-glucopyranoside and related n-alkyl pyranosides.

X-ray diffraction patterns have been obtained for the lyotropic phases of n-octyl 1-O-beta-D-glucopyranoside and the related n-heptyl, n-nonyl and n-decyl compounds with water. The octyl compound exhibits all three liquid crystal phases and forms a micellar solution with increasing solvation, when the crystal come into contact with water at room temperature. The X-ray diffraction patterns show a one-dimensional lamellar phase with [dx] = 28.4 A, a three-dimensional face-centered cubic phase with [a] = 51.2 A, and a two-dimensional hexagonal phase with [a] = [b] = 36.7 A. The micellar solution has a distribution pattern with a maximum at [dx] = 33.8 A. Crystals of the heptyl, nonyl and decyl derivatives form only the lamellar phases and the micellar solution on contact with water at room temperature.

Synthesis and anti-herpes-virus activity of acyclic 2'-deoxyguanosine analogues related to 9-[(1,3-dihydroxy-2-propoxy)methyl]guanine.

Several "sugar" modified acyclic nucleoside analogues related to 9-[(1,3-dihydroxy-2-propoxy)methyl]guanine (DHPG, 2) were synthesized and evaluated for antiviral activity. The preparation generally involved the condensation of the acetoxymethyl ether of alcohols 6c-g and 10-12a with diacetylguanine to give adducts 7c-g and 14-16, which were then deprotected to afford analogues 9c-g and 17-19. Alternatively, alcohols 12a and 13a were converted to iodides via their tosylates 12b and 13b and then reacted with the sodium salt of guanine to afford, after deprotection, analogues 22 and 23. A crossed aldol-Cannizzaro reaction on aldehyde 27 readily afforded 28, which was deprotected to give analogue 29. An in vitro assay against HSV-1 showed that all compounds tested were less active than DHPG, though several were good substrates for the viral thymidine kinase. The more promising acyclic nucleosides 9c, 19, and 29 were evaluated in a mouse encephalitis model and proved ineffective at preventing death at a dose of 20 mg/kg.

Acyclic analogues of 2'-deoxynucleosides related to 9-[(1,3-dihydroxy-2-propoxy)methyl]guanine as potential antiviral agents.

A series of acyclic analogues of 2'-deoxynucleosides related in structure to 9-[(1,3-dihydroxy-2-propoxy)methyl]guanine (DHPG, 1) have been synthesized and evaluated for antiviral activity against herpes simplex virus type 1 (F strain). Additionally, the ability of these analogues to function as substrates for the virus-specified thymidine kinase was examined. Phosphorylation by this kinase is essential for antiviral activity. Although the acyclic 4-oxopyrimidine nucleosides were substrates for the kinase, they were devoid of antiviral activity. In the purine series, most analogues similar in structure to DHPG did exhibit significantly lower antiviral activity, indicating that even small modifications in the purine substituents substantially reduce the antiviral potency. The most active agent, 2,6-diaminopurine 27, was only poorly phosphorylated by the viral kinase; therefore, its activity was most likely due to a prior enzymatic deamination to give DHPG. Evaluation of 27 in a mouse encephalitis model has shown it to be nearly as potent as DHPG (1).

The crystal structure of the C-nucleoside, 1,3-dimethyl-8-beta-D-ribofuranosylxanthine monohydrate.

1,3-Dimethyl-8-beta-D-ribofuranosylxanthine monohydrate, C12H16N4O6.H2O, is monoclinic, P21, with two molecules in a unit cell having a = 8.186(1), b = 19.222(3), c = 4.7655(8) A, beta = 103.79(1) degrees, V = 728.2 A, and Dx = 1.506 g.cm-3. The structure was solved by the direct methods, using MoK alpha radiation and refined with anisotropic temperature-factors. The final agreement factors were R = 0.061, Rw = 0.042 for 2254 structure amplitudes having [Fo[ greater than 2 sigma (Fo). The ribose ring has the C-2'-exo-C-3'-endo (2T3) conformation with pseudorotation parameters P = -3.2, tau m = 39.9 degrees, and a gauche-gauche conformation around the ribose C-4'-C-5' bond. The beta-link conformation is anti, with O-4'-C-1'-C-8-N-9 = -162.6 degrees. The pyrimidine and imidazole rings are planar, making an interplanar angle of 1.0(1) degree. The molecular conformation is stabilized by an unsymmetrical, three-center hydrogen bond from N-7-H to O-5'-H as the major component and to O-4'-H as the minor component. The hydrogen bonding includes a cooperative cycle involving the HN-C-C=O moiety, two hydroxyl groups, and a water molecule.

The crystal structure of L-chiro-inositol.

The crystal structure of L-chiro-inositol is monoclinic, P21, with a = 6.867(3), b = 9.133(4), c = 6.217(3) A, beta = 106.59(4) degrees, Z = 2. The structure was solved by using MULTAN, and refined to R = 0.028 for 1065 intensities observed with Ni-filtered MoK alpha radiation. The molecule has the expected chair conformation, with puckering parameters Q = 0.561 A, theta = 4.4 degrees, phi = 51.2 degrees. The non-hydrogen molecular symmetry is close to C2, with deviations of less than 0.07 A from a weighted fit. The intramolecular hydrogen-bonding forms infinite chains which are cross-linked through the weaker component of a three-center bond. The C-C bond lengths range from 1.515 to 1.528 A, and the C-O bond lengths from 1.418 to 1.436 A. The C-C-C angles range from 109.7 to 113.1 degrees, and the C-C-O angles from 106.5 to 112.0 degrees.

The crystal structure of an unusual chromophoric dimeric furan.

The red-orange crystals of (E)-2-[1-(3-hydroxy-2-furanyl)ethylidene]-(2H)-furan-3-one, C10H8O4, crystallize in space group P2(1)/c with Z = 8, cell dimensions at 123 K [293 K], a = 16.222(4) [16.360(8)] A, b = 7.089(2) [7.219(4)] A, c = 16.631(5) [16.722(8)] A, beta = 115.20(3) [115.50(7)]degrees. There are two symmetry nonequivalent molecules in the unit cell, each of which has an unsymmetrical configuration with an unsymmetrical O-H...O = C intramolecular hydrogen bond. This is contrary to a previous report based on the solid-state i.r. spectrum. The two furan rings in each molecule are planar, but not coplanar. They are inclined at angles of 33.4 degrees and 26.1 degrees to each other, in opposite senses in the two molecules.

The crystal structure of D-threitol at 119 K and 198 K.

A sample of DL-threitol, C4H10O4 (Sigma Chemical Co.), on recrystallization provided crystals of D- and L-threitol. The crystal structure was determined at 119 K and 298 K. The space group of D-threitol is P3(1)21, with three molecules in a unit cell at 119 K [298 K] of a = 10.0995(5) [10.1405(8)], c = 4.8407(4) [4.8767(4)] A. The final agreement R-factor was 0.050 for 302 intensities [0.069 for 244 intensities]. The molecules have the straight carbon-chain conformation with twofold axial symmetry. The hydroxyl groups are hydrogen bonded in infinite chains extending in the direction of the threefold screw axis. One of the hydroxyl hydrogens is twofold disordered, so that alternate chains have reversed donor-acceptor directions.

Hydrogen bonding in the crystal structure of alpha,beta-panose.

The crystal structure of panose, O-alpha-D-glucopyranosyl-(1----6)-O-alpha-D-glucopyranosyl-(1----4)-D-gl ucose, C18H32O16, has been refined using low-temperature, 123 K, CuK alpha X-ray data. Difference syntheses and least-squares refinement showed a 16% substitution of alpha-panose by the beta anomer. All the hydrogen atoms were located on difference synthesis with the exception of one attached to the low occupancy (beta) O-1". The final R-factor was 0.036 for 2135 observed structure amplitudes. The molecular conformation is stabilized by two interresidue intramolecular hydrogen bonds. All the hydroxyls and glycosidic oxygen atoms are involved in the hydrogen bonding, which is a two-dimensional network formed from finite and infinite chains.

Hydrogen bonding in the crystal structure of the tetrasaccharide stachyose hydrate: a 1:1 complex of two conformers.

The crystal structure of stachyose hydrate, O-alpha-D-galactopyranosyl-(1----6)-O-alpha-D-galactopyrano- syl-(1----6)-alpha-D-glucopyranosyl beta-D-fructofuranoside tetrahydrate, has been refined using X-ray data at 119 K. The crystal structure is a 1:1 complex of conformers which differ in the fructofuranosyl and glucopyranosyl residues. Each conformer has an associated hydrogen-bond structure which includes different sites for the water molecules. When superimposed in the crystal, this gives rise to two sites of 0.5 occupancy for one carbon and one oxygen atom of the fructofuranose component, and for three oxygen atoms of the glucopyranose component. The corresponding three carbon atoms of the glucopyranose component have anomalous thermal motion parameters. The four water molecules are distributed over nine sites, six with occupancy 0.5, two with occupancy 0.4, and one with occupancy 0.2. Four of the water sites are associated with one conformer, and four with the other. The hydrogen bonding includes infinite chains which are linked to form irregular ribbons, extending in the direction of the alpha axis. All hydroxyl, ring, and linkage oxygen atoms are involved in the hydrogen bonding, which includes two-centered, three-centered, and four-centered hydrogen bonds.

The crystal structure of anhydrous alpha,alpha-trehalose at -150 degrees.

The crystal structure of alpha,alpha-trehalose (alpha-D-glucopyranosyl alpha-D-glucopyranoside), C12H22O11, is monoclinic, P2(1), Z = 2, with unit cell dimensions at -150 degrees [20 degrees] of a = 12.971(5) [13.003(5)], b = 8.229(4) [8.252(4)], c = 6.789(3) [6.799(3)] A, beta = 98.12(4) [98.33(4)]. The crystal structure was solved by using MULTAN, and refined to R = 0.059, Rw = 0.048 for 1564 intensities, measured with MoK alpha radiation. The molecular structure is very similar to that observed in the dihydrate crystals. It has approximate C2 symmetry. Both glucopyranosyl groups are in the 4C1 conformation. The linkage torsion angles, O-5-C-1-O-1-C-1, are +60.8 degrees and +60.1 degrees. The primary alcohol groups are oriented gauche/gauche and gauche/trans, as in the dihydrate structure. A comparison of the cross-polarization, magic-angle-spinning (c.p.-m.a.s.), 13C-n.m.r. spectra for powders of the crystalline anhydrous and dihydrate forms shows differences in resonances assigned to C-1 and C-4 that would not be anticipated from the results of the crystal-structure analyses.

The crystal structure of isopropyl 1-thio-beta-D-galactopyranoside monohydrate at 123 K.

The crystal structure of isopropyl 1-thio-beta-D-galactopyranoside monohydrate is orthorhombic, P2(1)2(1)2(1), Z = 4, with cell dimensions at 123 K [293 K] of a = 7.983(1) [8.037(1)], b = 24.574(5) [24.709(4)], c = 6.329(1) [6.3736(8)] A, V = 1241.84 [1265.71] A3. The calculated and measured density is Dx = 1.371 [1.345] g cm-3, Dm = [1.340] g cm-3. Diffraction data were obtained with CuK alpha radiation and a Nonius CAD-4 diffractometer. The structure was solved by using MULTAN, and refined to R(F2) = 0.051, RW(F2) = 0.078, R(F) = 0.029, S = 1.16 for 1502 reflections. The molecule has the 4C1(D) conformation. The orientation of the primary alcohol group is gauche/trans, and that about the glycosidic C-S bond is (-)synclinal relative to the ring C-O bond. Although this compound does not form thermotropic liquid crystals, it has two crystal-to-crystal phase-transitions, at 70 and 104 degrees, prior to melting at 126 degrees. The crystal structure has a characteristic, amphiphilic, head-to-head bilayer molecular packing, with intercalated alkyl groups. The water molecule is included in the hydrogen-bond structure that links the galactoside moieties.

The tetrasaccharide nystose trihydrate: crystal structure analysis and hydrogen bonding.

The crystal structure of nystose trihydrate, O-beta-D-fructofuranosyl-(2-->1)-O-beta-D-fructofuranosyl-(2-->1)-beta-D -fructofuranosyl alpha-D-glucopyranoside trihydrate, C24H42O21.3H2O, has been determined using Cu K alpha X-ray data at 121 K. The space group is P2(1)2(1)2(1), with 4 molecules in a unit cell of dimensions a = 10.155(1), b = 13.506(1), and c = 23.278(2) A. The structure was refined to R = 0.052 and RW = 0.048 for 2734 observed structure amplitudes. The alpha-D-glucopyranose unit of the molecule has the normal 4C1 chair conformation and the three fructofuranose units have twist conformations lying between E3 and 4T5. One of the three water molecules is distributed over two sites: W-3 with occupancy 0.80 and W-3' with occupancy 0.20. All the hydrogen atoms were located on the difference synthesis with the exception of those attached to the low-occupancy water site. All hydroxyls, two of the three linkage oxygen atoms, and the water molecules are involved in a complex three-dimensional network which can be decomposed into a series of infinite chains intersecting at the water molecules to form homo- and hetero-dromic cycles.

The hydrogen bonding in the crystal structure of raffinose pentahydrate.

The crystal structure of raffinose pentahydrate, O-alpha-D-galactopyranosyl-(1----6)-O-alpha-D-glucopyranosyl-(1----2)- beta-D- fructofuranose pentahydrate, C18H32O16.5H2O, has been redetermined using low-temperature, 119 K, CuK alpha X-ray data. All hydrogen atoms were unambiguously located on difference syntheses. The final R-factor is 0.036 for 2423 observed structure amplitudes. The hydrogen bonding is composed of infinite chains, which are linked through the water molecules to form a three-dimensional network containing a chain of five linked water molecules. Three of the infinite chains extend in the directions of the crystallographic axis of the space group P2(1)2(1)2(1). Four of the water molecules accept two hydrogen bonds and one accepts one. All the hydroxyls and the ring and glycosidic oxygen atoms are involved in the hydrogen bonding. With one exception, the ring and glycosidic oxygens are hydrogen-bonded by means of the minor components of unsymmetrical three-center bonds.

The crystal structure and thermotropic liquid-crystal properties of N-n-undecyl-D-gluconamide.

N-n-Undecyl-D-gluconamide, C17H35O6, crystallizes in space group P1, with one molecule in a unit cell a = 5.2267(6), b = 19.628(9), c = 4.7810(4) A, alpha = 93.23(2), beta = 95.60(1), gamma = 89.58(2) degrees, V = 487.35 A3, Dx = 1.19 g.cm-3. The crystal lattice is isostructural with N-n-heptyl-D-gluconamide having monolayer head-to-tail molecular packing. The molecules have a V-shaped conformation. The hydrogen bonding of the gluconamide moieties includes a four-link homodromic cycle. The transition to a smectic A liquid-crystal phase at 156.7 degrees is preceded by two crystal-to-crystal phase transitions at 77.2 degrees and 99.4 degrees. The long d-spacing of the intermediate crystal phase of 39 A, and the d-spacing of the liquid-crystal phase of 32 A, are consistent with a transition to a bilayer head-to-head molecular packing.