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The structure of the phases obtained upon dehydration and decomposition of Friedel's salt [Ca2Al(OH)6]Cl·2H2O, also known as hydrocalumite, was investigated by several experimental techniques, in particular, high-temperature in situ XRD measurements, which allowed the detection of a metastable intermediate phase. Thermogravimetric analyses show that Friedel's salt, like most of the layered double hydroxides, undergoes a three-step decomposition on heating (dehydration, dehydroxylation, and anion expulsion) over the following temperature ranges: 25−280, 280−400, and >400 °C. Sharp phase transitions are observed as a result of the ordered distribution of Ca and Al atoms in the hydroxide layer and the well-ordered interlayer structure. Upon cooling to room temperature and exposure to the atmosphere, the dehydrated phase obtained by calcination between 80 and 280 °C was found to recover the basal spacing characteristic of hydrated galleries (7.81 Å) within a few minutes. The structural determination of this thermally metastable phase based on X-ray powder diffraction data recorded at 116 °C reveals a quasi-pillared layer structure with chloride anions situated midway in the interlamellar space at only 2.904(3) Å from Ca atoms of adjacent hydroxide layers. Friedel's salt becomes amorphous at ca. 400 °C, and above 750 °C, it crystallizes into a mixture of CaO and mayenite Ca12Al14O33. Exposure of the amorphous residue obtained at 400 °C to aqueous solutions of KCl led to reconstruction−intercalation phenomena.

Hydrocalumite, Ca2Al(OH)6(anion)·nH2O, belongs to the layered double-hydroxide family, with the particularity for both intralayered cations and interlayered anions and water molecules to be well-ordered. The monomer vinyl benzene sulfonate (VBS) and the syndiotactic polymer poly(styrene sulfonate) (PSS) were readily incorporated into its lamellar structure via a coprecipitation method. In situ thermal polymerization of the monomer VBS is simultaneously supported by the disappearance of the vinyl bond and the formation of −CH, −CH2 bonds. These facts are evidenced by the experiments performed by ¹³C solid-state NMR and FTIR spectroscopies. CaAl−VBS intercalate crystallizes in the C2/c space group, inducing a bilayer arrangement of VBS anions in the interlamellar space. Besides, distances between anionic sites allow an in situ zigzag polymerization of syndiotactic type along the a axis. Additionally, the contraction of the lamellar structure is found to be reversible up to 200 °C; the adjacent monomers then start to attach to each other via polymerization. The thermal stability is found to be highly dependent on the synthesis pathway. The decomposition is delayed up to 450 °C for the sample prepared via in situ polymerization while as-prepared PSS intercalate decomposes at lower temperature. Under a nitrogen atmosphere, CaS is formed at 700 °C.

The reaction of enantiopure (R)-(2-cyclohexa-1,4-dienyl)propan-1-ol with ruthenium chloride hydrate yields, without racemisation of the chiral ligand, the chloro-bridged dinuclear complex (S,S)-[RuCl2{C6H5[CH(CH3)CH2OH]}]2 (1). The dimer 1 reacts with triphenylphosphane to give the mononuclear complex (S)-[RuCl2{C6H5[CH(CH3)CH2OH]}(PPh3)] (2). A single-crystal X-ray structure analysis of 2 reveals the absolute configuration of the asymmetric carbon atom to have remained, the change from (R) to (S) being due to the priority change caused by coordination; in the solid state, onedimensional hydrogen-bonded chains are formed between the hydroxy functions and chloro ligands of neighbouring molecules. The mononuclear cationic complex (S)-[Ru{C6H5[CH(CH3)CH2OH]}(H2O)3]²⁺, formed in situ from 1 in aqueous solution, reacts with the dinuclear complex [H3Ru2(C6Me6)2]⁺ to give a chiral trinuclear arene-ruthenium cluster, the cation (S)-[H3Ru3{C6H5[CH(CH3)CH2OH]} (C6Me6)2(O)]⁺ (3). This enantiopure cation has been isolated and characterised as its tetrafluoroborate salt. The single-crystal X-ray structure analysis of (S)-[3][BF4] shows a strong intramolecular hydrogen bond between the μ3-oxo cap and the hydroxy function, which also persists in acetone solvent, as demonstrated by NMR spectroscopy. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003)

Abstract: Fumarate reductase was used as a model oxidoreductase to demonstrate continuous electrical cofactor reduction-oxidation during the bioelectrochemical synthesis and detection of chemicals. The enzyme preparation was immobilized onto a graphite felt electrode that was modified with carboxymethylcellulose (CMC). Nicotinamide adenine dinucleotide (NAD), neutral red, and fumarate reductase (which contained menaquinone) were covalently linked by peptide bonds to the CMC. The electron mediator neutral red allowed NAD and menaquinone to be recycled electrically during enzymatic chemical synthesis. Succinate detection by the bioelectrocatalyst was linear from 5 μM to 10 mM succinate. Fumarate synthesis using this bioelectrode was dependent on succinate utilization and resulted in proportional production of electricity and fumarate. Succinate synthesis using this bioelectrocatalyst was dependent on current and fumarate concentration. This bioelectrocatalyst system may enhance the utility of menaquinone- and/or pyridine nucleotide-linked oxidoreductases in diverse enzymatic fuel cells and sensors. It may also enhance the utility of oxidoreductase-based chemical synthesis systems because it eliminates the problem of cofactor recycling.

The Thermotoga neapolitana xylose isomerase (TNXI) is extremely thermostable and optimally active at 95°C. Its derivative, TNXI Val185Thr (V185T), is the most active type II xylose isomerase reported, with a catalytic efficiency of 25.1 s–1 mM–1 toward glucose at 80°C (pH 7.0). To further optimize TNXI’s potential industrial utility, two rounds of random mutagenesis and low temperature/low pH activity screening were performed using the TNXI V185T‐encoding gene as the template. Two highly active mutants were obtained, 3A2 (V185T/L282P) and 1F1 (V185T/L282P/F186S). 1F1 was more active than 3A2, which in turn was more active than TNXI V185T at all temperatures and pH values tested. 3A2 and 1F1’s high activities at low temperatures were due to significantly lower activation energies (57 and 44 kJ/mol, respectively) than that of TNXI and V185T (87 kJ/mol). Mutation L282P introduced a kink in helix α7 of 3A2’s (α/β)8 barrel. Surprisingly, this mutation kinetically destabilized 3A2 only at pH 5.5. 1F1 displayed kinetic stability slightly above that of TNXI V185T. In 1F1, mutation F186S creates a cavity that disrupts a four‐residue network of aromatic interactions. How the conformation of the neighboring residues is affected by this cavity and how these conformational changes increase 1F1’s stability still remain unclear.

The adenylate kinase (AK) gene from Thermotoga neapolitana, a hyperthermophilic bacterium, was cloned and overexpressed in Escherichia coli, and the recombinant enzyme was biochemically characterized. The T. neapolitana AK (TNAK) sequence indicates that this enzyme belongs to the long bacterial AKs. TNAK contains the four cysteine residues that bind Zn2+ in all Gram-positive AKs and in a few other Zn2+-containing bacterial AKs. Atomic emission spectroscopy and titration data indicate a content of 1 mol of Zn2+/mol of recombinant TNAK. The EDTA-treated enzyme has a melting temperature (Tm=93.5 °C) 6.2 °C below that of the holoenzyme (99.7 °C), identifying Zn2+ as a stabilizing feature in TNAK. TNAK is a monomeric enzyme with a molecular mass of approx. 25 kDa. TNAK displays Vmax and Km values at 30 °C identical with those of the E. coli AK at 30 °C, and displays very high activity at 80 °C, with a specific activity above 8000 units/mg. The unusually high activity of TNAK at 30 °C makes it an interesting model to test the role of enzyme flexibility in activity.

In situ high-pressure NMR spectroscopy of the hydrogenation of benzene to give cyclohexane, catalysed by the cluster cation [(η6-C6H6) (η6-C6Me6)2Ru3(μ3-O)(μ2-OH)(μ2-H)2]+2, supports a mechanism involving a supramolecular host-guest complex of the substrate molecule in the hydrophobic pocket of the intact cluster molecule.

The conversion of glucose to fructose at elevated temperatures, as catalyzed by soluble and immobilized xylose (glucose) isomerases from the hyperthermophiles Thermotoga maritima (TMGI) and Thermotoga neapolitana 5068 (TNGI) and from the mesophile Streptomyces murinus (SMGI), was examined. At pH 7.0 in the presence of Mg²⁺, the temperature optima for the three soluble enzymes were 85°C (SMGI), 95° to 100°C (TNGI), and >100°C (TMGI). Under certain conditions, soluble forms of the three enzymes exhibited an unusual, multiphasic inactivation behavior in which the decay rate slowed considerably after an initial rapid decline. However, the inactivation of the enzymes covalently immobilized to glass beads, monophasic in most cases, was characterized by a first-order decay rate intermediate between those of the initial rapid and slower phases for the soluble enzymes. Enzyme productivities for the three immobilized GIs were determined experimentally in the presence of Mg²⁺. The highest productivities measured were 750 and 760 kg fructose per kilogram SMGI at 60°C and 70°C, respectively. The highest productivity for both TMGI and TNGI in the presence of Mg²⁺ occurred at 70°C, pH 7.0, with approximately 230 and 200 kg fructose per kilogram enzyme for TNGI and TMGI, respectively. At 80°C and in the presence of Mg²⁺, productivities for the three enzymes ranged from 31 to 273. A simple mathematical model, which accounted for thermal effects on kinetics, glucose–fructose equilibrium, and enzyme inactivation, was used to examine the potential for high-fructose corn syrup (HFCS) production at 80°C and above using TNGI and SMGI under optimal conditions, which included the presence of both Co²⁺ and Mg²⁺. In the presence of both cations, these enzymes showed the potential to catalyze glucose-to-fructose conversion at 80°C with estimated lifetime productivities on the order of 2000 kg fructose per kilogram enzyme, a value competitive with enzymes currently used at 55° to 65°C, but with the additional advantage of higher fructose concentrations. At 90°C, the estimated productivity for SMGI dropped to 200, whereas, for TNGI, lifetime productivities on the order of 1000 were estimated. Assuming that the most favorable biocatalytic and thermostability features of these enzymes can be captured in immobilized form and the chemical lability of substrates and products can be minimized, HFCS production at high temperatures could be used to achieve higher fructose concentrations as well as create alternative processing strategies. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 80: 185–194, 2002.

ABSTRACTPrion infection relies on a continuous chain of PrPc-expressing tissues to spread from peripheral sites to the central nervous system (CNS). Direct neuroinvasion via peripheral nerves has long been considered likely. However, the speed of axonal flow is incompatible with the lengthy delay prior to the detection of PrPScin the brain. We hypothesized that Schwann cells could be the candidate implicated in this mechanism; for that, it has to express PrPcand to allow PrPScconversion. We investigated in vivo localization of PrPcin sciatic nerve samples from different strains of mice. We demonstrated that PrPcis mainly localized at the cell membrane of the Schwann cell. We also studied in vitro expression of PrPcin the Schwann cell line MSC-80 and demonstrated that it expresses PrPcat the same location. More specifically, we demonstrated that this glial cell line, when infected in vitro with the mouse Chandler prion strain, both produces the PrPSctill after 18 passages and is able to transmit disease to mice, which then develop the typical signs of prion diseases. It is the first time that infection and replication of PrPScare shown in a peripheral glial cell line.

AbstractHistorical factors are rarely taken into account despite their high potential for the understanding of vegetation determinism. This study aims to propose a method to stress the importance of historical factors on the current vegetation. It consists in a comparative socio-ecological analysis of archaeological sites versuscontrol sites, both being in the same ecological conditions for climate, substrate and biotic pressure at present. This approach has been applied to a set of ten feudal mottes located in some forests of the Picardie district (France). Our results show that 1) the feudal mottes support a neutrophilous, eutrophic and semi-heliotropic vegetation though the surrounding forest vegetation is markly acidic, mesotrophic and sciaphilous, 2) specific richness trends to be higher on the mottes, 3) a few species for which the status of « feudal plants » is questioned were also observed.

Xylose isomerases (XIs) from Thermoanaerobacterium thermosulfurigenes (TTXI) and Thermotoga neapolitana (TNXI) are 70.4% identical in their amino acid sequences and have a nearly superimposable crystal structure. Nonetheless, TNXI is much more thermostable than TTXI. Except for a few additional prolines and fewer Asn and Gln residues in TNXI, no other obvious differences in the enzyme structures can explain the differences in their stabilities. TNXI has two additional prolines in the Phe59 loop (Pro58 and Pro62). Mutations Gln58Pro, Ala62Pro and Gln58Pro/Ala62Pro in TTXI and their reverse counterpart mutations in TNXI were constructed by site-directed mutagenesis. Surprisingly, only the Gln58Pro mutation stabilized TTXI. The Ala62Pro and Gln58Pro/Ala62Pro mutations both dramatically destabilized TTXI. Analysis of the three-dimensional (3D) structures of TTXI and its Ala62Pro mutant derivative showed a close van der Waal's contact between Pro62-Cδ and atom Lys61-Cβ (2.92 Å) thus destabilizing TTXI. All the reverse counterpart mutations destabilized TNXI thus confirming that these two prolines play important roles in TNXI's thermostability. TTXI's active site has been previously engineered to improve its catalytic efficiency toward glucose and increase its thermostability. The same mutations were introduced into TNXI, and similar trends were observed, but to different extents. Val185Thr mutation in TNXI is the most efficient mutant derivative with a 3.1-fold increase in its catalytic efficiency toward glucose. With a maximal activity at 97°C of 45.4 U/mg on glucose, this TNXI mutant derivative is the most active type II XI ever reported. This `true' glucose isomerase engineered from a native xylose isomerase has now comparable kinetic properties on glucose and xylose.

Epidemiological surveillance, namely the continuous monitoring of diseases and health determinants in a population, has developed over the past fifteen years, in the sphere of human health as well as in animal health. All epidemiological surveillance networks include the following four stages: data collection, data transmission, data processing and dissemination of information. However, despite this basic similarity, the very many networks existing in France are extremely varied in nature. At the national level, the bodies involved in epidemiological surveillance for infectious animal diseases are the Direction générale de l'alimentation, the Agence française de sécurité sanitaire des aliments and, to a lesser degree, the Institut français de recherche pour l'exploitation de la mer. In the field, the networks rely on the Direction des services vétérinaires, veterinary practitioners, laboratories in each département, and livestock producers' groups (especially animal health protection groups). Some twenty French networks currently in operation are presented in this article according to a classification based on published criteria. In the case of human infectious diseases, epidemiological surveillance is carried out almost entirely by the Direction générale de la santé and the Directions départementales d'action sanitaire et sociale, the Institut de veille sanitaire and the various Centres nationaux de référence (CNRs). Most human infectious diseases are monitored by one or more of the following broad categories of networks: reporting of notifiable diseases, the CNRs, the network of sentinel doctors, the network of hospital laboratories and departments, and medical causes of death. An example where surveillance is covered by several networks is also presented, namely surveillance for salmonellosis and Salmonella. Lastly, methods for evaluating networks are discussed.

This study compares the clinical efficacy and safety of a thermal uterine balloon system with hysteroscopic endometrial resection in the treatment of dysfunctional uterine bleeding. In all, 147 women were treated by two experienced gynaecological surgeons: one performed 73 thermal balloon ablations and the other 74 endometrial resections between November 1994 and April 1998. The inclusion criteria were similar in both groups. The operative time was reduced significantly with the uterine balloon technique. There were no intra-operative complications in either group and postoperative morbidities were minimal and not statistically different. Multivariate analysis noted two prognostic factors associated with failures: retroverted uterus with thermal balloon ablation and age under 43 years with endometrial resection. The overall success rate did not differ significantly between the two groups 83.0 ± 5% for balloon ablation and 76.3 ± 6% for endometrial resection. Uterine balloon ablation appears to be as efficacious as endometrial resection. The former is much easier to perform, making the technique readily reproducible, especially by those with limited expertise in hysteroscopic surgery, and thus more widely applicable and safer.

This study compares the clinical efficacy and safety of a thermal uterine balloon system with hysteroscopic endometrial resection in the treatment of dysfunctional uterine bleeding. In all, 147 women were treated by two experienced gynaecological surgeons: one performed 73 thermal balloon ablations and the other 74 endometrial resections between November 1994 and April 1998. The inclusion criteria were similar in both groups. The operative time was reduced significantly with the uterine balloon technique. There were no intra-operative complications in either group and postoperative morbidities were minimal and not statistically different. Multivariate analysis noted two prognostic factors associated with failures: retroverted uterus with thermal balloon ablation and age under 43 years with endometrial resection. The overall success rate did not differ significantly between the two groups 83.0 ± 5% for balloon ablation and 76.3 ± 6% for endometrial resection. Uterine balloon ablation appears to be as efficacious as endometrial resection. The former is much easier to perform, making the technique readily reproducible, especially by those with limited expertise in hysteroscopic surgery, and thus more widely applicable and safer.

The human HOXB2 gene is a member of the vertebrate Hox gene family that contains genes coding for specific developmental stage DNA-binding proteins. Remarkably, within the hematopoietic compartment, genes of the HOXB complex are expressed specifically in erythromegakaryocytic cell lines and, for some of them, in hematopoietic progenitors. Here, we report the study of HOXB2 gene transcriptional regulation in hematopoietic cells, an initial step in understanding the lineage-specific expression of the whole HOXB complex in these cells. We have isolated the HOXB2 5'-flanking sequence and have characterized a promoter fragment extending 323 base pairs upstream from the transcriptional start site, which, in transfection experiments, was sufficient to direct the tissue-specific expression of HOXB2 in the erythroid cell line K562. In this fragment, we have identified a potential GATA-binding site that is essential to the promoter activity as demonstrated by point mutation experiments. Gel shift analysis revealed the formation of a specific complex in both erythroleukemic lines K562 and HEL that could be prevented by the addition of a specific antiserum raised against GATA-1 protein. These findings suggest a regulatory hierarchy in which GATA-1 is upstream of the HOXB2 gene in erythroid cells.

Members of theAntennapediaclass of homeobox genes, known as Hox genes, are believed to be pivotal in vertebrate craniofacial development. Here we show that eight members of paralogous groups 1, 2, 3, and 4 are expressed in the human embryonic hindbrain and branchial arches at 4 weeks of development. The combinatorial patterns of expression of genes representing the first three paralogous groups parallel the patterns described for their homologues in various animal models, demonstrating a high degree of conservation of the branchial Hox code. Arch expression of group 4 genes is defined for the first time in any vertebrate. Furthermore, as development proceeds, individual paralogues of a single paralogous group (group 3), which initially share a common expression domain, are differentially down-regulated in a tissue-, organ-, or site-specific fashion.

AbstractObjective:To determine the rates and routes of Acinetobacter baumaniicolonization and pneumonia among ventilated patients in a surgical intensive-care unit (SICU) before and after architectural modifications.Design:A nonsequential study comparing two groups of patients. All isolates from systematic and clinical samples were genotyped by pulsed-field gel electrophoresis (PFGE). Records of patients hospitalized during the first and second periods were reviewed and findings were compared. Between the two periods, the SICU was remodeled from enclosed isolation rooms and open rooms to only enclosed isolation rooms with handwashing facilities in each room.Setting and Patients:All patients hospitalized and mechanically ventilated for more than 48 hours in the 15-bed SICU of the University Hospital of Besançon (France).Results:For the first and second periods, the rates of colonization were, respectively, 28.1% and 5.0% of patients (P<10−7; relative risk [RR], 2.23; 95% confidence interval [CI95], 1.8-2.75) and the specific rates of bronchopulmonary (BP) colonization were, respectively, 9.1 and 0.5 per 1,000 days of mechanical ventilation (P<10−5). Seven major PFGE isolate types were identified, 4 of which were isolated from 44 of the 47 colonized or infected patients. Logistic regression analysis showed that colonization was not associated with patient characteristics.Conclusion:Conversion from open rooms to isolation rooms may help control nosocomial BP tract acquisition of A baumaniiin mechanically ventilated patients hospitalized in an SICU.

The adhB gene encoding Thermoanaerobacter ethanolicus 39E secondary-alcohol dehydrogenase (S-ADH) was cloned, sequenced and expressed in Escherichia coli. The 1056 bp gene encodes a homotetrameric recombinant enzyme consisting of 37.7 kDa subunits. The purified recombinant enzyme is optimally active above 90 °C with a half-life of approx. 1.7 h at 90 °C. An NADP(H)-dependent enzyme, the recombinant S-ADH has 1400-fold greater catalytic efficiency in propan-2-ol oxidation than in ethanol oxidation. The enzyme was inactivated by chemical modification with dithionitrobenzoate (DTNB) and diethylpyrocarbonate, indicating that Cys and His residues are involved in catalysis. Zinc was the only metal enhancing S-ADH reactivation after DTNB modification, implicating the involvement of bound zinc in catalysis. Arrhenius plots for the oxidation of propan-2-ol by the native and recombinant S-ADHs were linear from 25 to 90 °C when the enzymes were incubated at 55 °C before assay. Discontinuities in the Arrhenius plots for propan-2-ol and ethanol oxidations were observed, however, when the enzymes were preincubated at 0 or 25 °C. The observed Arrhenius discontinuity therefore resulted from a temperature-dependent, catalytically significant S-ADH structural change. Hydrophobic cluster analysis comparisons of both mesophilic and thermophilic S-ADH and primary- versus S-ADH amino acid sequences were performed. These comparisons predicted that specific proline residues might contribute to S-ADH thermostability and thermophilicity, and that the catalytic Zn ligands are different in primary-alcohol dehydrogenases (two Cys and a His) and S-ADHs (Cys, His, and Asp).

Regional and national legislation mandates the disclosure of “priority” allergens when present as an ingredient in foods, but this does not extend to the unintended presence of allergens due to shared production facilities. This has resulted in a proliferation of precautionary allergen (“may contain”) labels (PAL) that are frequently ignored by food-allergic consumers. Attempts have been made to improve allergen risk management to better inform the use of PAL, but a lack of consensus has led to variety of regulatory approaches and nonuniformity in the use of PAL by food businesses. One potential solution would be to establish internationally agreed “reference doses,” below which no PAL would be needed. However, if reference doses are to be used to inform the need for PAL, then it is essential to characterize the hazard associated with these low-level exposures. For peanut, there are now published data relating to over 3000 double-blind, placebo-controlled challenges in allergic individuals, but a similar level of evidence is lacking for other priority allergens. We present the results of a rapid evidence assessment and meta-analysis for the risk of anaphylaxis to a low-level allergen exposure for priority allergens. On the basis of this analysis, we propose that peanut can and should be considered an exemplar allergen for the hazard characterization at a low-level allergen exposure.

L’absence unilatérale d’artère pulmonaire (UAPA) résulte d’une anomalie de développement du 6earc aortique lors de l’embryogenèse. Il s’agit d’une malformation rare, dont la prévalence est estimée à 1 cas sur 200 000, fréquemment associée à des malformations cardiaques congénitales lorsqu’elle est découverte dans l’enfance.

This paper provides an overview of the latest scientific data related to the safety of uncontaminated oats (<20 ppm of gluten) in the diet of individuals with celiac disease (CD). It updates the previous Health Canada position posted on the Health Canada website in 2007 and a related paper published in 2009. It considers a number of recent studies published between January 2008 and January 2015. While recognizing that a few people with celiac disease seem to be clinically intolerant to oats, this review concludes that oats uncontaminated by gluten-containing cereals (wheat, rye, and barley) can be safely ingested by most patients with celiac disease and that there is no conclusive evidence that the consumption of uncontaminated or specially produced oats containing no greater than 20 ppm gluten by patients with celiac disease should be limited to a specific daily amount. However, individuals with CD should observe a stabilization phase before introducing uncontaminated oats to the gluten-free diet (GFD). Oats uncontaminated with gluten should only be introduced after all symptoms of celiac disease have resolved and the individual has been on a GFD for a minimum of 6 months. Long-term regular medical follow-up of these patients is recommended but this is no different recommendation to celiac individuals on a GFD without oats.