Composition and Chemical Variability of the Essential Oil from Aerial Parts of Cladanthus scariosus,an Endemic Species to the Moroccan High Atlas

Cladanthus scariosus (Ball) Oberpr. & Vogt is endemic to Moroccan High Atlas. It is known under the vernacular names Irezghi or Irezgui. Three essential oil samples have been isolated from aerial parts and analyzed by combination of chromatographic and spectroscopic techniques [gas chromatography (GC) in combination with retention indices (RI), gas chromatography-mass spectrometry (GC/MS) and ¹³C-NMR spectroscopy]. The compositions of oil samples were dominated by monoterpenes: α-pinene sabinene, and terpinen-4-ol. Chamazulene and dihydrochamazulene isomers as well as various hemiterpene esters and analogs have been identified. To evidence a chemical variability, statistical analysis performed on 13 oil sample compositions allowed partitioning into three groups, mainly differentiated by their contents of sabinene, camphor, borneol, terpinen-4-ol, and germacrene D.     

Abscisic acid, indole-3-acetic acid and cytokinin changes in buds of Pseudotsuga menziesii during bud quiescence release

Bud quiescence release, considered as the ultimate dormancy breaking phase, was achieved in Pseudotsuga menziesii (Mirb.) Franco by a 9-week cold (5°C) treatment, under short daylength (9 h) followed by a transfer to mild temperature (22°C) under long daylength (16 h). Indole-3-acetic acid (IAA), abscisic acid (ABA), zeatin-type (Z) and isopentenyladenine-type (iPA) cytokinin (CK) levels were measured by means of an ELISA technique performed on HPLC-fractionated extracts of terminal and axillary buds. During the cold period, all hormones except IP-type CK levels decreased, whereas the opposite observation was made after transfer to mild temperature and long daylength, when buds started to grow. Some other immunoreactive compounds were also detected and quantified. The ABA-glucosyl ester (ABA-GE) level pattern was similar to that of ABA, but no accumulation occurred at mild temperatures. A putative IAA conjugate, more polar than IAA, was also detected. Its level increased transiently like IAA in terminal buds and, to a lesser extent, in axillary buds during the 10th week of the experiment. In terminal buds, isopentenyladenosine ([9R]-iP) was released by alkaline hydrolysis of a polar immunoreactive compound detected with anti-[9R]iP antibodies. This compound accumulated during the cold period and quickly dropped at 22°C. Relationships between environmental conditions and endogenous hormones are discussed.     

Possible role of a short extra loop of the long-chain flavodoxin from Azotobacter chroococcum in electron transfer to nitrogenase: Complete 1H, 15N and 13C backbone assignments and secondary solution structure of the flavodoxin

Summary The ¹H, ¹⁵N and ¹³C backbone and ¹H and ¹³C beta resonance assignments of the long-chain flavodoxin from Azotobacter chroococcum (the 20-kDa nifF product, flavodoxin-2) in its oxidized form were made at pH 6.5 and 30°C using heteronuclear multidimensional NMR spectroscopy. Analysis of the NOE connectivities, together with amide exchange rates, ³J_Hn_H coupling constants and secondary chemical shifts, provided extensive solution secondary structure information. The secondary structure consists of a five-stranded parallel -sheet and five -helices. One of the outer regions of the -sheet shows no regular extended conformation, whereas the outer strand 4/6 is interrupted by a loop, which is typically observed in long-chain flavodoxins. Two of the five -helices are nonregular at the N-terminus of the helix. Loop regions close to the FMN are identified. Negatively charged amino acid residues are found to be mainly clustered around the FMN, whereas a cluster of positively charged residues is located in one of the -helices. Titration of the flavodoxin with the Fe protein of the A. chroococcum nitrogenase enzyme complex revealed that residues Asn¹¹, Ser⁶⁸ and Asn⁷² are involved in complex formation between the flavodoxin and Fe protein. The interaction between the flavodoxin and the Fe protein is influenced by MgADP and is of electrostatic nature.Abbreviations SQ semiquinone - FMN riboflavin 5-monophosphate; nif, nitrogen fixation - TSP 3-(trimethylsilyl)propionate sodium salt - DSS 2,2-dimethyl-2-silapentane-5-sulfonate sodium salt Supplementary Material is available on request, comprising a Materials and Methods section for the expression and purification of the A. chroococcum flavodoxin, a Table S1 containing the parameters of the titration of A. chroococcum flavodoxin with the Fe protein, and a Table S2 containing the ¹⁵N, H^N, ¹³C, ¹H, ¹³C, ¹H and ¹³CO chemical shifts.To whom correspondence should be addressed.     

Computer calculation-based quantitative structure-activity relationships for the oxidation of phenol derivatives horseradish peroxidase compound II

 The second-order rate constants for the oxidation of a series of phenol derivatives by horseradish peroxidase compound II were compared to computer-calculated chemical parameters characteristic for this reaction step. The phenol derivatives studied were phenol, 4-chlorophenol, 3-hydroxyphenol, 3-methylphenol, 4-methylphenol, 4-hydroxybenzoate, 4-methoxyphenol and 4-hydroxybenzaldehyde. Assuming a reaction of the phenolic substrates in their non-dissociated, uncharged forms, clear correlations (r = 0.977 and r = 0.905) were obtained between the natural logarithm of the second-order rate constants (ln k _app and ln k ₂ respectively) for their oxidation by compound II and their calculated ionisation potential, i.e. minus the energy of their highest occupied molecular orbital [E(HOMO)]. In addition to this first approach in which the quantitative structure-activity relationship (QSAR) was based on a calculated frontier orbital parameter of the substrate, in a second and third approach the relative heat of formation (ΔΔHF) calculated for the process of one-electron abstraction and H^• abstraction from the phenol derivatives was used as a parameter. Plots of the natural logarithms of the second-order rate constants (k _app and k ₂) for the reaction and the calculated ΔΔHF values for the process of one-electron abstraction also provide clear QSARs with correlation coefficients of –0.968 and –0.926 respectively. Plots of the natural logarithms of the second-order rate constants (k _app and k ₂) for the reaction and the calculated ΔΔHF values for the process of H^• abstraction provide QSARs with correlation coefficients of –0.989 and –0.922 respectively. Since both mechanisms considered, i.e. initial electron abstraction versus initial H^• abstraction, provided clear QSARs, the results could not be used to discriminate between these two possible mechanisms for phenol oxidation by horseradish peroxidase compound II. The computer calculation-based QSARs thus obtained for the oxidation of the various phenol derivatives by compound II from horseradish peroxidase indicate the validity of the approaches investigated, i.e. both the frontier orbital approach and the approach in which the process is described by calculated relative heats of formation. The results also indicate that outcomes from computer calculations on relatively unrelated phenol derivatives can be reliably compared to one another. Furthermore, as the actual oxidation of peroxidase substrates by compound II is known to be the rate-limiting step in the overall catalysis by horseradish peroxidase, the QSARs of the present study may have implications for the differences in the overall rate of substrate oxidation of the phenol derivatives by horseradish peroxidase. Received: 29 March 1996 / Accepted: 17 July 1996     

Analysis of human V H gene repertoire expression in peripheral CD19+ B cells

Using CD19 B-cell selection and polymerase chain reaction-amplified cDNA libraries, we analyzed the peripheral immunoglobulin heavy chain variable repertoire of three healthy adult donors. Here we report that most of the CD19+ circulating B cells expressed unmutated V _H-D-J_H rearrangements. By specific V _H family hybridization, we show that V _H gene family utilization in the periphery roughly corresponds to the complexity of these families in the germline and appears to be relatively constant among the analyzed subjects. However, sequence data of clones picked at random from one IgM cDNA library reveals that in spite of this random utilization, the V _H gene expression in naive circulating B cells is highly biased towards the expression of a limited set of V _H genes. As previously reported by others, this restricted mechanism is also found for the D and J _H segments.The nucleotide sequence data reported in this paper have been submitted to the GenBank/EMBL nucleotide sequence database and have been assigned the accession numbers Z47213-Z47243 and Z47349     

Astroglial Cells Express Large Amounts of GABAA Receptor Proteins in Mature Brain

Abstract: GABA_A receptors were characterized in cellular fractions isolated from adult bovine brain. The fraction enriched in cortical astrocytes is very rich in high-affinity binding sites for [³H]flunitrazepam and other "central-type" benzodiazepine ligands. The amount of specific [³H]flunitrazepam binding was more than five times higher in the glial fraction than in synaptosomal and perikaryal fractions. [³H]Flunitrazepam was displaced by low concentrations of clonazepam and other specific ligands for central GABA_A receptors. Specific binding sites for GABA, flunitrazepam, barbiturates, and picrotoxin-like convulsants were characterized. Allosteric interactions between the different sites were typical of central-type GABA_A receptors. The presence of α-subunit(s), as revealed by [³H]flunitrazepam photoaffinity labeling, was demonstrated in all brain fractions at molecular mass 51–53 kDa. Photoaffinity labeling was highest in the glial fraction. However, in primary cultured astrocytes from neonate rat cortex, no photoaffinity labeling was detected. Information obtained from astrocytes in culture should thus be taken with caution when extrapolated to differentiated astroglial cells. Our results actually show that, in mature brain, most of the fully pharmacologically active GABA_A receptors are extrasynaptic and expressed in astroglia.     

A review of issues related to the quantification of bacteria from the phyllosphere

Abstract: Quantification of the size of epiphytic bacterial populations and characterization of their composition involves definition of a sampling strategy in time and space, the choice of methods for liberating the bacteria from the leaf surface and for recovering them for subsequent determination of the number of viable or culturable cells. This literature review focuses on some of the issues related to these choices. After briefly reviewing the different types of epiphytic colonizers we consider the biological, methodological, and statistical consequences of the choice of the sampling unit and of the spatial and temporal variability of population size and composition for epiphytic bacteria. The different methods available for the detection and enumeration of naturally occurring microorganisms in the phyllosphere are discussed. Advantages and drawbacks of each are described in this review designed as a 'hands-on' guide.