Wolfgang Zimmer

Physiological and molecular biological characterization of ammonia oxidation of the heterotrophic nitrifier Pseudomonas putida.

Abstract. The heterotrophic nitrifier Pseudomonas putida aerobically oxidized ammonia to hydroxylamine, nitrite, and nitrate. Product formation was accompanied by a small but significant release of NO, whereas N2O evolution could not be detected under the assay conditions employed. The isolate reduced nitrate to nitrite and partially further to NO under anaerobic conditions. Aerobically grown cells utilized γ-aminobutyrate as a carbon source and as a N-source by ammonification. The physiological experiments, in particular the inhibition pattern by C2H2, indicated that P. putida expressed an ammonia monooxigenase. DNA-hybridization with an amoA gene probe coding for the smaller subunit of the ammonia monooxigenase of Nitrosomonas europaea allowed us to identify, to clone, and to sequence a region with an open reading frame showing distinct sequence similarities to the amoA gene of autotrophic ammonia oxidizers.

Identification and isolation of the indole-3-pyruvate decarboxylase gene from Azospirillum brasilense Sp7: sequencing and functional analysis of the gene locus.

Abstract. The root-associated bacterium Azospirillum brasilense Sp7 produces the growth-stimulating phytohormone indole-3-acetic acid (=IAA) via the indole-3-pyruvate pathway. The DNA region containing ipdC, the structural gene for indole-3-pyruvate decarboxylase, was identified in a cosmid gene library of strain Sp7 by hybridization and has been sequenced. Upstream of the gene, two other ORF homologous to gltX and cysS were sequenced that are transcribed in the opposite direction. A functional analysis of the cloned ipdC region has been performed. To test the expression of the gene, a lacZ-Km cartridge was introduced into the gene. By this construct, tryptophan-dependent stimulation of gene expression in A. brasilense Sp7 was observed. Evidences for the existence of another copy of the ipdC gene in the Azospirillum genome are also reported.

Identification of the nifJ gene coding for pyruvate: ferredoxin oxidoreductase in dinitrogen-fixing cyanobacteria

A 329 bp DNA segment from both Anabaena variabilis and Anabaena PCC 7119 was amplified using the polymerase chain reaction (PCR). The sequences from the two cyanobacteria showed strong similarities to the corresponding part of the nifJ gene from Klebsiella pneumoniae and Enterobacter agglomerans. The present findings underline earlier results of enzymatic studies that heterocystous cyanobacteria possess a pyruvate: ferredoxin (flavodoxin) oxidoreductase (PFO). The nifJ gene segment could not be detected in the non-dinitrogenfixing, unicellular cyanobacterium Anacystis nidulans which is also in accord with previous findings from enzyme assays.

Amperometric method for determining nitrous oxide in denitrification and in nitrogenase-catalyzed nitrous oxide reduction

A conventional Clark-type O2 probe was used to determine N2O concentrations in suspensions. At a polarizing voltage of−0.95 V versus the reference Ag/AgCl electrode, the probe is almost half as sensitive for N2O as for O2, and the detection limit is less than 1 μM N2O. The probe can also be used to determine NO for which the suitable polarizing voltage is−0.7 V. The method was successfully applied for continuously recording dissimilatory formation or utilization of N2O by intactAzospirillum brasilense Sp 7, NO production by extracts from this bacterium, and N2O reduction catalyzed by nitrogenase in intactKlebsiella pneumoniae. It is concluded that the probe is useful for measuring N2O or NO contents in bacterial suspensions when the O2 level is zero or kept constant during the assays.

Energy transduction efficiencies in nitrogenous oxide respirations of Azospirillum brasilense Sp7

AbstractFor Azospirillum brasilense Sp7, the energy transformation efficiencies were measured in anaerobic respirations with either nitrate, nitrite or nitrous oxide as respiratory electron acceptors by determining the maximal molar growth yields and the H+-translocations using the oxidant pulse method. In continuous cultures grown with malate limiting, the maximal molar growth yields (Ysmax-values) were essentially the same with O2 or N2O but were 1/3 and 2/3 lower with NO2-or NO3-, respectively, as respiratory electron acceptors. Both the maximal molar growth yields and the maintenance energy coefficients were surprisingly high when Azospirillum was grown with nitrite as the sole electron acceptor and source for N-assimilation. Growth under N2-fixing conditions drastically reduced the Ysmax-values in the N2O and O2-respiring cells. In the H+-translocation measurements, the

Auxin biosynthesis and denitrification in plant growth promoting bacteria

\vec H^ +

Isolation and functional analysis of a cDNA encoding a myrcene synthase from holm oak (Quercus ilex L.).

/oxidant ratios were 5.6 for O2→H2O, 2.5–2.8 for NO3-→NO2-, 2.2 for NO2-→N2O and 3.1 for N2O→N2 respirations when the cells were preincubated with valinomycin and K+. All the values were enhanced when the experiments were performed with valinomycin plus methyltriphenylphosphonium (=TPMP+) cation. The uncoupler carbonyl cyanide-m-chlorophenyl-hydrazone diminished the H+-excretion indicating that this translocation was due to vectorial flow across the membrane. In the absence of any ionophore, nitrate and nitrite respirations were accompanied by a H+-uptake