S^Nitrosoglutathione (GSNO), an adduct of nitric oxide (NO) with glutathione, is known as a biological NO reservoir. Heterologous expression in Escherichia coli of a cDNA encoding a glutathione-dependent formaldehyde dehydrogenase of Arabidopsis thaliana showed that the recombinant protein reduces GSNO. The identity of the cDNA was further confirmed by functional complementation of the hypersensitivity to GSNO of a yeast mutant with impaired GSNO metabolism. This is the first demonstration of a plant GSNO reductase, suggesting that plants possess the enzymatic pathway that modulates the bioactivity and toxicity of NO. ß
HIROMICHI MORIKAWA and ROBERT H. MARCHESSAULT. Can. J. Chem. 59,2306(1981. Pyrolysis products from bacterial polyesters, poly-P-hydroxybutyrate (PHB), and froma heteropolyester(P-hydroxyvalerate and P-hydroxybutyrate) were identified. Different physical forms of PHB were studied: PHB purified by dissolution in chloroform, native granules of PHB, and PHB in bacterial cells. The products were characterized by gc, ms, and nmr analysis. The yield of crotonic acid obtained by the pyrolysis of purified PHB was 60 to 65%. Pyrolysis of PHB native granules yielded crotonic acid as well as oligomers of PHB with a terminal crotonate. Upon direct pyrolysis of dry bacterial cells, the yield of crotonic acid was 20 to 25% of the PHB in the cells. From the heteropolymer, 2-pentenoic acid and terminally unsaturated oligomers of polyhydroxyvalerate were obtained. The usefulness of the pyrolysis method for obtaining vinyl compounds from bacterial cultures containing polyalkanoates is discussed.HIROMICHI MORIKAWA et ROBERT H. MARCHESSAULT. Can. J. Chem. 59,2306(1981. On a identifie les produits provenant de la pyrolyse des polyesters bactiriens poly-P-hydroxybutyrate (PHB) et du polyester heterogene (P-hydroxyvalirate e t P-hydroxybutyrate). On a Ctudie les differentes formes physiques du PHB: le PHB purifie par dissolution dans le chloroforme, les granules naturels de PHB et le PHB des cellules bacteriennes. On acaracterise les produits par chromatographie en phase gazeuse, par spectrometrie de masse et par la rmn. Le rendement en acide crotonique provenant de la pyrolyse du PHB purifie est de 60 a 65%. L a pyrolyse des granules naturels de PHB conduit aussi bien 5 I'acide crotonique qu'a des oligomkres du PHB avec un crotonate terminal. La pyrolyse directe des cellules bacteriennes s6ches conduit a un rendement en acide crotonique qui represente 20 a 25% du PHB contenu dans les cellules. On a obtenu du polymere hkterogkne I'acide penten-2 I oique et des oligomkres du polyhydroxyvalerate insaturk en position terminale. On discute de I'utilite de la methode de pyrolyse en I vue d'obtenir des composes vinyliques a partir des cultures bacteriennes contenant des polyalcanoates.
. Can. J. Chem. 59,38 (1981).The solid state properties of poly-0-hydroxybutyrate (PHB) were investigated for samples with degrees of polymerization (m) from 4 to 994. The observed melting points ranged from 47°C to 18O0C. Electron diffractograms on carefully prepared single crystals of a high molecular weight sample provided data which confirmed the reported a and b parameters from X-ray fiber diffraction and provided clear justification for the P2,2,2, space group. The observed intensities in the electron diffractogram, corresponding to (hM) reflections, were compared with the predicted intensities for two proposed structures in the literature. The observed and calculated structure factors for both structures were in good agreement. Small angle X-ray diffraction ofthe meridional maximum for annealed "cold drawn" and "hot drawn" fibers showed a distinctly different dependence on temperature than the maximumm stacks of lamellar single crystals. For oligomers a long spacing was observed which was about twice the length of the sample DPn multiplied by the crystalline advance per monomer. Les points de fusion vont de 47°C a 180°C. Les absences systematiques observees en diffraction electronique sur monocristaux de forte masse confirment le groupe spatiale P2,2,2, propose a partir du diagramme a fibre (rayons-X). Les intensites observies des reflections (hkO) du cliche de diffraction tlectronique sont en accord avec celles predites a partir de deux structures crystallines proposies dans la literature et basies sur les donnees du diagramme a fibre. La diffusion meridionale des rayons-X aux petlts angles pour des fibres ttirtes "ifroid" et "achaud" a e t t enregistree et la variation des maximade diffusion, enfonction de la temperature, differe sensiblement de ce qui est observe pour la variation de l'kpaisseur de lamelles monocristallines. Pour des oligomtres cristallins I'observation d'une tache de diffraction dont I'espacement est deux fois la longueur de la chaine dans sa conformation crystalline mtne proposer une maille avec deux chaines bout bout selon I'axe c.
Transgenic plants of Arabidopsis bearing the spinach (Spinacia oleracea) nitrite reductase (NiR, EC 1.7.7.1) gene that catalyzes the six-electron reduction of nitrite to ammonium in the second step of the nitrate assimilation pathway were produced by use of the cauliflower mosaic virus 35S promoter and nopaline synthase terminator. Integration of the gene was confirmed by a genomic polymerase chain reaction (PCR) and Southern-blot analysis; its expression by a reverse transcriptase-PCR and two-dimensional polyacrylamide gel electrophoresis western-blot analysis; total (spinach ϩ Arabidopsis) NiR mRNA content by a competitive reverse transcriptase-PCR; localization of NiR activity (NiRA) in the chloroplast by fractionation analysis; and NO 2 assimilation by analysis of the reduced nitrogen derived from NO 2 (NO 2 -RN). Twelve independent transgenic plant lines were characterized in depth. Three positive correlations were found for NiR gene expression; between the total NiR mRNA and total NiR protein contents (r ϭ 0.74), between the total NiR protein and NiRA (r ϭ 0.71), and between NiRA and NO 2 -RN (r ϭ 0.65). Of these twelve lines, four had significantly higher NiRA than the wild-type control (P Ͻ 0.01), and three had significantly higher NO 2 -RN (P Ͻ 0.01). Each of the latter three had one to two copies of spinach NiR cDNA per haploid genome. The NiR flux control coefficient for NO 2 assimilation was estimated to be about 0.4. A similar value was obtained for an NiR antisense tobacco (Nicotiana tabacum cv Xanthi XHFD8). The flux control coefficients of nitrate reductase and glutamine synthetase were much smaller than this value. Together, these findings indicate that NiR is a controlling enzyme in NO 2 assimilation by plants.
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