Novel derivatives of ansa-titanocenes procured from 6-phenylfulvene: a combined experimental and theoretical study

The previously prepared trans-[(1,2-diphenyl-1,2-dicyclopentadienyl)ethanediyl] titanium(IV) dichloride, [1,2-(Ph)₂C₂H₂{η⁵-C₅H₄}₂]Ti(Cl)₂, was synthesised using an alternative procedure, from which its crystal structure was determined. Using this compound, a variety of other ansa-titanocene derivatives were synthesised by replacement of the chloride ligands with selected substituents. Thus RTi(X)(Y) systems where R=1,2-(Ph)₂C₂H₂η⁵-C₅H₄₂; X=Y=CH₃; X=CH₃, Y=Cl; X=Y=NCS; X=Y=NCO; X=Y=OPh and (X/Y)=O have been synthesised and characterised. DFT calculations were performed on the complexes trans-[(1,2-diphenyl-1,2-dicyclopentadienyl)-ethanediyl] titanium(IV) dichloride, bis-(6,6-diphenylfulvene)titanium and bis-(6,6-diphenylfulvene)iron. This demonstrated the role that the metal centre plays in their formation, generating either an ansa-metallocene, a ‘tucked in’ fulvene complex or a metallocene coordinating fulvene anions.     

Ebola virus glycoprotein needs an additional trigger, beyond proteolytic priming for membrane fusion

Background

Ebolavirus belongs to the family filoviridae and causes severe hemorrhagic fever in humans with 50–90% lethality. Detailed understanding of how the viruses attach to and enter new host cells is critical to development of medical interventions. The virus displays a trimeric glycoprotein (GP_1,2) on its surface that is solely responsible for membrane attachment, virus internalization and fusion. GP_1,2 is expressed as a single peptide and is cleaved by furin in the host cells to yield two disulphide-linked fragments termed GP1 and GP2 that remain associated in a GP_1,2 trimeric, viral surface spike. After entry into host endosomes, GP_1,2 is enzymatically cleaved by endosomal cathepsins B and L, a necessary step in infection. However, the functional effects of the cleavage on the glycoprotein are unknown.

Principal Findings

We demonstrate by antibody binding and Hydrogen-Deuterium Exchange Mass Spectrometry (DXMS) of glycoproteins from two different ebolaviruses that although enzymatic priming of GP_1,2 is required for fusion, the priming itself does not initiate the required conformational changes in the ectodomain of GP_1,2. Further, ELISA binding data of primed GP_1,2 to conformational antibody KZ52 suggests that the low pH inside the endosomes also does not trigger dissociation of GP1 from GP2 to effect membrane fusion.

Significance

The results reveal that the ebolavirus GP_1,2 ectodomain remains in the prefusion conformation upon enzymatic cleavage in low pH and removal of the glycan cap. The results also suggest that an additional endosomal trigger is necessary to induce the conformational changes in GP_1,2 and effect fusion. Identification of this trigger will provide further mechanistic insights into ebolavirus infection.     

Kinetics of Bacterial Growth on Chlorinated Aliphatic Compounds

With the pure bacterial cultures Ancylobacter aquaticus AD20 and AD25, Xanthobacter autotrophicus GJ10, and Pseudomonas sp. strain AD1, Monod kinetics was observed during growth in chemostat cultures on 1,2-dichloroethane (AD20, AD25, and GJ10), 2-chloroethanol (AD20 and GJ10), and 1,3-dichloro-2-propanol (AD1). Both the Michaelis-Menten constants (K_m) of the first catabolic (dehalogenating) enzyme and the Monod half-saturation constants (K_s) followed the order 2-chloroethanol, 1,3-dichloro-2-propanol, epichlorohydrin, and 1,2-dichloroethane. The K_s values of strains GJ10, AD20, and AD25 for 1,2-dichloroethane were 260, 222, and 24 μM, respectively. The low K_s value of strain AD25 was correlated with a higher haloalkane dehalogenase content of this bacterium. The growth rates of strains AD20 and GJ10 in continuous cultures on 1,2-dichloroethane were higher than the rates predicted from the kinetics of the haloalkane dehalogenase and the concentration of the enzyme in the cells. The results indicate that the efficiency of chlorinated compound removal is indeed influenced by the kinetic properties and cellular content of the first catabolic enzyme. The cell envelope did not seem to act as a barrier for permeation of 1,2-dichloroethane.     

The Fibronectin Type 3-Like Repeat from the Clostridium thermocellum Cellobiohydrolase CbhA Promotes Hydrolysis of Cellulose by Modifying Its Surface

Fibronectin type 3 homology domains (Fn3) as found in the cellobiohydrolase CbhA of Clostridium thermocellum are common among bacterial extracellular glycohydrolases. The function of these domains is not clear. CbhA is modular and composed of an N-terminal family IV carbohydrate-binding domain (CBDIV), an immunoglobulin-like domain, a family 9 glycosyl hydrolase catalytic domain (Gh9), two Fn3-like domains (Fn3_1,2), a family III carbohydrate-binding domain (CBDIII), and a dockerin domain. Efficiency of cellulose hydrolysis by truncated forms of CbhA increased in the following order: Gh9 (lowest efficiency), Gh9-Fn3_1,2 (more efficient), and Gh9-Fn3_1,2-CBDIII (greatest efficiency). Thermostability of the above constructs decreased in the following order: Gh9 (most stable), Gh9-Fn3_1,2, and then Gh9-Fn3_1,2-CBDIII (least stable). Mixing of Orpinomyces endoglucanase CelE with Fn3_1,2, or Fn3_1,2-CBDIII increased efficiency of hydrolysis of acid-swollen cellulose (ASC) and filter paper. Scanning electron microscopic studies of filter paper treated with Fn3_1,2, Fn3_1,2-CBDIII, or CBDIII showed that the surface of the cellulose fibers had been loosened up and crenellated by Fn3_1,2 and Fn3_1,2-CBDIII and to a lesser extent by CBDIII. X-ray diffraction analysis did not reveal changes in the crystallinity of the filter paper. CBDIII bound to ASC and filter paper with capacities of 2.45 and 0.73 μmoles g⁻¹ and relative affinities (K_r) of 1.12 and 2.13 liters g⁻¹, respectively. Fn3_1,2 bound weakly to both celluloses. Fn3_1,2-CBD bound to ASC and filter paper with capacities of 3.22 and 0.81 μmoles g⁻¹ and K_rs of 1.14 and 1.98 liters g⁻¹, respectively. Fn3_1,2 and CBDIII contained 2 and 1 mol of calcium per mol, respectively. The results suggest that Fn3_1,2 aids the hydrolysis of cellulose by modifying its surface. This effect is enhanced by the presence of CBDIII, which increases the concentration of Fn3_1,2 on the cellulose surface.     

Synthesis of Glycols by Microbial Transformation of Some Monocyclic Terpenes

The transformation of three monocyclic terpenes by three soil microorganisms have been studied. The organisms were isolated on, and grew rapidly in, mineral salts medium containing the appropriate terpene substrates as sole carbon sources. These organisms belong to the class Fungi Imperfecti, and two of them have been tentatively identified as Cladosporium species. A Cladosporium species designated T₁ was isolated from terpene-soaked soil, using 1-menthene as the sole source of carbon. The major catabolic product isolated from the growth medium of this organism was found to be a cyclic 1,2-diol identified as trans-p-methane-1,2-diol. A similar but biochemically distinct Cladosporium sp. designated T₇ was isolated on D-limonene. After growth, the medium of this organism contained 1.5 g/liter of the analogous product, trans-limonene-1,2-diol. Minor quantities of the corresponding cis-1,2-diol were also isolated. The third organism, designated as laboratory culture T₈, was isolated on 3-menthene and yielded a diol identified as trans-p-menthane-3,4-diol. From these results it is concluded that the formation of diols is a common intermediate in the fungal metabolism of monocyclic terpenes.     

Studies on glutathione-S-arene oxidase transferase. A sensitive assay and partial purification of the enzyme from sheep liver

A sensitive assay has been devised for glutathione-S-arene oxidase transferase using as substrates naphthalene-1,2-oxide or styrene oxide along with [³⁵S]glutathione. Activity of the order of 2–3 nmoles of conjugate formed during a 5-min incubation can be detected. This yields about 2000 cpm above a blank of about 1500 cpm. Transferase activity was found mainly in liver and kidney but was also present in most other tissues of rats. Glutathione-S-arene oxide transferase has been purified 70- to 80-fold from sheep liver 100,000 g supernatants using the conventional procedures. After electrofocusing, enzyme activity separated into two major peaks and two or three minor peaks, ranging in isoelectric point from pH 6.5 to 7.5. Activities assayed with naphthalene-1,2-oxide or styrene oxide as substrates were found to almost parallel each other in all the peaks.The sheep liver transferase required neither metal ions nor cofactors such as FAD, pyridoxal-phosphate and thiamine pyrophosphate. The molecular weight of the transferase has been estimated to be about 40,000.K_m values for glutathione, naphthalene-1,2-oxide, and styrene oxide are 1.6, 0.11, and 0.13 mm, respectively. K_m values for glutathione decreased with increasing pH, whereas the K_m values for naphthalene-1,2-oxide were independent of pH in the range of 6.5–8.     

Bidentate palladium(II) chelation by the common aldoses

The [Pd^II{(R,R)-chxn}(OH)₂] reagent (chxn = 1,2-diaminocyclohexane) is introduced as a metal probe for the detection of the bidentate chelating sites of a glycose. Two moles of hydroxide per mole palladium support double deprotonation of potentially chelating diol functions at a glycose’s backbone. The individual chelating sites are detected using one- and two-dimensional NMR techniques. At equimolar amounts of palladium(II) and aldose, the metal-binding sites include mostly the hydroxy function at the anomeric carbon atom. Chelators are derived from both the pyranose and the furanose isomers. Most pyranose-based chelators form five-membered chelate rings by using their 1,2-diol function. Though 1,2-diolate bonding is also common to the furanoses, the formation of six-membered chelate rings by 1,3-bonding is more significant for them. Metal-excess conditions provoke mostly bis-bidentate 1,2;3,4-chelation but unusual isomers form also: thus d-xylose is dimetallated in its all-axial β-pyranose form, and erythrose’s dimetallation results in the formation of two isomers of a metal derivative of the open-chain hydrate. The spectroscopic results are supported by crystal-structure determinations on [Pd{(R,R)-chxn}(α-d-Xylp1,2H₋₂-κO^1,2)]·H₂O (Xyl = xylose), [Pd{(R,R)-chxn}(α-d-Ribp1,2H₋₂-κO^1,2)]·2.25H₂O (Rib = ribose), [Pd{(R,R)-chxn}(α-l-Thrf1,3H₋₂-κO^1,3)]·2H₂O (Thr = threose) and [Pd{(R,R)-chxn}(α-d-Eryf1,3H₋₂-κO^1,3)]·3H₂O (Ery = erythrose).     

Synthesis and X-ray structure of a C5-C4-linked glucofuranose-oxazolidin-2-one

The formation of (4R)-4-carbamoyl-4-[(4R)-3-O-benzyl-1,2-O-isopropylidene-β-l-threofuranos-4-C-yl]-oxazolidin-2-one instead of expected imidazolidin-2,4-dione (hydantoin) derivative from 5-amino-5-cyano-5-deoxy-3-O-benzyl-1,2-O-isopropylidene-α-d-glucofuranose or 3-O-benzyl-1,2-O-isopropylidene-α-d-xylo-hexofuranos-5-ulose under Bucherer-Bergs reaction conditions is reported. Single crystal X-ray diffraction data revealed that ³T₄ is the prefered conformation for the furanose ring, while E₂ and ²T₁ conformations are adopted by the 1,3-dioxolane and 2-oxazolidinone five-membered rings, respectively.     

Catabolism of Naphthalenesulfonic Acids by Pseudomonas sp. A3 and Pseudomonas sp. C22

Naphthalene and two naphthalenesulfonic acids were degraded by Pseudomonas sp. A3 and Pseudomonas sp. C22 by the same enzymes. Gentisate is a major metabolite. Catabolic activities for naphthalene, 1-naphthalenesulfonic acid, and 2-naphthalenesulfonic acid are induced by growth with naphthalene, 1-naphthalenesulfonic acid, 2-naphthalenesulfonic acid, methylnaphthalene, or salicylate. Gentisate is also an inducer in strain A3. Inhibition kinetics show that naphthalene and substituted naphthalenes are hydroxylated by the same naphthalene dioxygenase. Substrates with nondissociable substituents such as CH₃, OCH₃, Cl, or NO₂ are hydroxylated in the 7,8-position, and 4-substituted salicylates are accumulated. If CO₂H, CH₂CO₂H, or SO₃H are substituents, hydroxylation occurs with high regioselectivity in the 1,2-position. Thus, 1,2-dihydroxy-1,2-dihydronaphthalene-2-carboxylic acids are formed quantitatively from the corresponding naphthalenecarboxylic acids. Utilization of naphthalenesulfonic acids proceeds by the same regioselective 1,2-dioxygenation which labilizes the C—SO₃⁻ bond and eliminates sulfite.     

Enzymatic deacylation of l,2-diacyl-sn-glycero-3-phosphocholines to sn-glycerol-3-phosphocholine

This research was undertaken to study the enzymatic deacylation of l,2-diacyl-sn-glycero-3-phosphocholines (sn-1,2-PC) to sn-glycerol-3-phosphocholine (GPC); this compound could be an useful intermediate in the synthesis of “structured” sn-1,2-PC, after re-acylations of the two sn-positions of the glycerol backbone. The enzymatic reactions represent a valid alternative to the chemical deacylation that can be simply obtained in alkaline conditions. High conversion were achieved using a lipase selective for the sn-1-position of sn-1,2-PC (Lipozyme IM, from Mucor miehei) together with a Phospholipase A₂ from hog pancreas, enzyme selective for the sn-2-position; the best results were obtained carrying out the enzymatic reaction in a microemulsion system.     

Chair-form [Ag2(1,2-bimb)2] in silver(I) complexes containing the ditopic ligand 1,2-bis(1-imidazolylmethyl)benzene (1,2-bimb)

The ditopic ligand 1,2-bis(1-imidazolylmethyl)benzene (1,2-bimb) and its silver(I) complexes [Ag₂(1,2-bimb)₂](PF₆)₂ (1) and {[Ag₂(1,2-bimb)₂]₂(SbF₆)₄}_n (2) were prepared and their structures characterized by X-ray crystallography. Both complexes contain the chair-form unit [Ag₂(1,2-bimb)₂]²⁺ with Ag(I) linearly coordinated by N_Im (Im=1-imidazolyl) from the Im groups of two 1,2-bimb. However, the [Ag₂(1,2-bimb)₂]²⁺ units are positioned differently in forming 1D infinite chains through weak argentophilic interactions: linear chains in 1 with the units oriented in same direction are formed via Ag?π interactions, while polymeric chains constructed via Ag?Ag interactions in 2 are observed with the units arranged in alternate directions. Differences in supramolecular structures may be a result of different size of the anions.     

1,10-Phenanthroline-dithiolate mixed ligand transition metal complexes. Synthesis, characterization and EPR spectroscopy

A series of new N₂S₂ mixed ligand transition metal complexes, where N₂ is phenanthroline and S₂ is 1,2-dithiooxalate (dto) or 1,2-dithiosquarate (dtsq), has been synthesized and characterized. IR spectra reveal that the 1,2-dithiolate ligands are coordinated via the sulfur atoms forming a N₂S₂ coordination sphere. The copper(II) complex [Cu(phen)(dto)] was studied by EPR spectroscopy as a diamagnetically diluted powder. The diamagnetic dilution resulted from doping of the copper complex into the isostructural host lattice of the nickel complex [Ni(phen)(dto)]. The electronic situation in the frontier orbitals of the copper complex calculated from the experimental data is compared to the results of EHT and DFT calculations. Furthermore, one side product, chlorobis(1,10-phenanthroline)copper(I) ethanol solvate hydrate [(phen)₂CuCl]·C₂H₅OH·H₂O, was formed by a reduction process and characterized by X-ray diffraction. In the crystal packing one-dimensional columns of dimers are formed, stabilized by significant π-π interactions.     

Purification and Characterization of Two Epoxide Hydrolases from Corynebacterium sp. Strain N-1074

Enzymes II_a and II_b, which catalyze the conversion of epichlorohydrin (ECH) to 3-chloro-1,2-propanediol (MCP), were purified from Corynebacterium sp. strain N-1074, which catalyzes the formation of (R)-MCP from prochiral 1,3-dichloro-2-propanol via ECH. The specific activity of enzyme II_a for the formation of MCP from ECH was about 6.4-fold higher than that of enzyme II_b. Both enzymes catalyzed the conversion of 1,2-epoxides to the corresponding diol, although they differed in several enzymatic properties.     

Highly stereoselective synthesis of C-vinyl furanosides through acid-catalyzed S(N)2 inversion at the C-3 position of 1,2-dideoxy-hept-1-enitols

A highly stereoselective synthesis of C-vinyl furanosides through the S_N2 inversion at the C-3 position of the 1,2-dideoxy-hept-1-enitols is disclosed. Treatment of the 1,2-dideoxy-hept-1-enitols with diphenylammonium trifluoromethanesulfonate as the acid catalyst produced the C-vinyl furanosides (3,6-anhydro-1,2-dideoxy-hept-1-enitol derivatives) via a subsequent S_N2 intramolecular debenzyloxyation-cycloetherification reaction at the C-3 position.     

An improved method for the preparation of 1,2-isopropylidene-SN-glycerol

A simple and fast route for the preparation of 1,2-isopropylidene-sn-glycerol from D-mannitol in 45% yield is described. The value of optical rotation, [α]_D²⁰ + 15.2°, is higher than usual indicating considerable racemization for other procedures. Since 1,2-isopropylidene-sn-glycerol serves as general intermediate for the synthesis of glycerides and of phosphoglycerides these lipids contain substantial amounts of the isomer, for instance 1,2-dipalmitoyl-sn-glycerol-3-phosphocholine may consist of up to 15% of 2,3-dipalmitoyl-sn-glycerol-1-phosphocholine in earlier preparations.     

Catechol 1,2-dioxygenase from the new aromatic compounds - Degrading Pseudomonas putida strain N6

This study aimed to characterization of catechol 1,2-dioxygenase from a Gram-negative bacterium, being able to utilize a wide spectrum of aromatic substrates as a sole carbon and energy source. Strain designated as N6, was isolated from the activated sludge samples of a sewage treatment plant at Bentwood Furniture Factory Jasienica, Poland. Morphology, physio-biochemical characteristics and phylogenetic analysis based on 16S rDNA sequence indicate that strain belongs to Pseudomonas putida. When cells of strain N6 grown on protocatechuate or 4-hydroxybenzoic acid mainly protocatechuate 3,4-dioxygenase was induced. The activity of catechol 1,2-dioxygenase was rather small. The cells grown on benzoic acid, catechol or phenol showed high activity of only catechol 1,2-dioxygenase. This enzyme was optimally active at 35 °C and pH 7.4. Kinetic studies showed that the value of K_m and V_max was 85.19 ??M and 14.54 ??M min⁻¹ respectively. Nucleotide sequence of gene encoding catechol 1,2-dioxygenase in strain N6 has 100% identity with catA genes from two P. putida strains. The deduced 301-residue sequence of enzyme corresponds to a protein of molecular mass 33.1 kDa. The deduced molecular structure of the catechol 1,2-dioxygenase from P. putida N6 was very similar and characteristic for the other intradiol dioxygenases.     

Coordination chemistry of the heterotopic 1,2-phenylenebis(thio)diacetic acid ligand: Rhodium(I), palladium(II) and nickel(II) complexes

Starting from the heterotopic multidentate ligand 1,2-phenylenebis(thio)diacetic acid (1), cis-rac-[PdCl₂{1,2-(HOOCCH₂S)₂C₆H₄-κ²S,S′}] (2), cis-rac-[Rh{1,2-(HOOCCH₂S)₂C₆H₄-κ²S,S′}(cod)]BF₄ (3) and cis-rac-[Ni{1,2-(OOCCH₂S)₂C₆H₄-κ⁴O,O′S,S′}{cis-(C₃H₄N₂)}₂] (4) were prepared and characterised by X-ray diffraction and conventional spectroscopic techniques. Compounds 1-4 show extensive hydrogen-bonded networks (XH?O, X = O, N) in the solid state.     

1,2-Hydrogen shifts during the biosynthesis of patchoulenes in Pogostemon cablin

The isotope ratio in α- and γ-patchoulenes in Pogostemon cablin, that has been fed with [2-¹⁴C, 4R-³H₁]MVA, suggests that a proton loss is followed by a 1,2-alkyl shift and two 1,2-hydrogen shifts during the biosynthesis of these two sesquiterpene hydrocarbons. Whereas isotope ratios in β- and δ-patchoulene suggests that a proton loss is followed by one 1,2-hydrogen shift in β-patchoulene and two 1,2-hydrogen shifts in δ-patchoulene.     

NifI inhibits nitrogenase by competing with Fe protein for binding to the MoFe protein

Reduction of substrate by nitrogenase requires direct electron transfer from the Fe protein to the MoFe protein. Inhibition of nitrogenase activity in Methanococcus maripaludis occurs when the regulatory protein NifI_1,2 binds the MoFe protein. This inhibition is relieved by 2-oxoglutarate. Here we present evidence that NifI_1,2 binding prevents association of the two nitrogenase components. Increasing amounts of Fe protein competed with NifI_1,2, decreasing its inhibitory effect. NifI_1,2 prevented the co-purification of MoFe protein with a mutant form of the Fe protein that forms a stable complex with the MoFe protein, and NifI_1,2 was unable to bind to an -stabilized Fe protein:MoFe protein complex. NifI_1,2 inhibited ATP- and MoFe protein-dependent oxidation of the Fe protein, and 2OG relieved this inhibition. These results support a model where NifI_1,2 competes with the Fe protein for binding to MoFe protein and prevents electron transfer.     

Assessing the impact of inductive electronic effects on the metrical parameters and reactivity of a series of ferrous complexes

Reported are four iron(II) complexes with N-benzyl-N,N′-bis(2-pyridylmethyl)-1,2-ethanediamine (L^H) and three electronically modified derivatives: N-(4-methoxy)benzyl-N,N′-bis(2-pyridylmethyl)-1,2-ethanediamine (L^OMe), N-(4-chloro)benzyl-N,N′-bis(2-pyridylmethyl)-1,2-ethanediamine (L^Cl), and N-(4-nitro)benzyl-N,N′-bis(2-pyridylmethyl)-1,2-ethanediamine (L^NO₂). The four ligands react with FeCl₂ to form a series of mononuclear species with the general formula [Fe(L^R)Cl₂]. The cis-α conformation of the ligand places the amine N-donors trans to the Fe-Cl bonds. The identity of the 4-benzyl substituent has profound influences on the lengths of the iron-ligand bonds, the optical spectra, and the redox activities of the [Fe(L^R)Cl₂] compounds.