Baceridin, a cyclic hexapeptide from an epiphytic bacillus strain, inhibits the proteasome.

A new cyclic hexapeptide, baceridin (1), was isolated from the culture medium of a plant-associated Bacillus strain. The structure of 1 was elucidated by HR-HPLC-MS and 1D and 2D NMR experiments and confirmed by ESI MS/MS sequence analysis of the corresponding linear hexapeptide 2. The absolute configurations of the amino acid residues were determined after derivatization by GC-MS and Marfey's method.

Homologous expression of the nrdF gene of Corynebacterium ammoniagenes strain ATCC 6872 generates a manganese-metallocofactor (R2F) and a stable tyrosyl radical (Y˙) involved in ribonucleotide reduction.

Ribonucleotide reduction, the unique step in the pathway to DNA synthesis, is catalyzed by enzymes via radical-dependent redox chemistry involving an array of diverse metallocofactors. The nucleotide reduction gene (nrdF) encoding the metallocofactor containing small subunit (R2F) of the Corynebacterium ammoniagenes ribonucleotide reductase was reintroduced into strain C. ammoniagenes ATCC 6872.

A tyrosyl-dimanganese coupled spin system is the native metalloradical cofactor of the R2F subunit of the ribonucleotide reductase of Corynebacterium ammoniagenes.

The X-ray crystallographic structure of the native R2F subunit of the ribonucleotide reductase (RNR) of Corynebacterium ammoniagenes ATCC 6872 is reported, with a resolution of 1.36 A. The metal site contains an oxo/hydroxo-bridged manganese dimer, located near a tyrosine residue (Y115).

Crystallization and preliminary X-ray analysis of the small subunit (R2F) of native ribonucleotide reductase from Corynebacterium ammoniagenes.

Ribonucleotide reduction, the unique step in DNA-precursor biosynthesis, involves radical-dependent redox chemistry and diverse metallo-cofactors. The metallo-cofactor (R2F) encoded by the nrdF (nucleotide reduction) gene in Corynebacterium ammoniagenes ATCC 6872 was isolated after homologous expression and a new crystal form of ribonucleotide reductase R2F was obtained. R2F was crystallized at 277 K using the vapour-diffusion method with PEG as the precipitating agent.

Electron paramagnetic resonance (EPR) spectroscopy of the stable-free radical in the native metallo-cofactor of the manganese-ribonucleotide reductase (Mn-RNR) of Corynebacterium glutamicum.

Ribonucleotide reductases (RNR; EC 1.17.4.

A polyphasic approach assigns the pathogenic Erwinia strains from diseased pear trees in Japan to Erwinia pyrifoliae.

Aims: Bacterial shoot blight of pear in Japan (BSBP) is caused by Erwinia strains which were formerly associated with the species Erwinia amylovora, the causative agent of fire blight. The description of Erwinia pyrifoliae as a pear pathogen in Korea renewed a possible connection of the pear pathogens in both countries.

Methods And Results: Nucleotide sequence analysis of the 16S rRNA, the house keeping genes gpd and recA, as well as DNA-DNA hybridization kinetics and microbiological assays place the pear pathogens from Japan into the species E.

Aquincola tertiaricarbonis gen. nov., sp. nov., a tertiary butyl moiety-degrading bacterium.

Strains L10(T), L108 and CIP I-2052 were originally obtained from methyl tert-butyl ether (MTBE)-contaminated groundwater and from a wastewater treatment plant, respectively. All share the ability to grow on tert-butanol, an intermediate of MTBE degradation. Cells are strictly aerobic, motile by a polar flagellum and exhibit strong pili formation.

High-level resistance to cobalt and nickel but probably no transenvelope efflux: Metal resistance in the Cuban Serratia marcescens strain C-1.

Molecular mechanisms underlying inducible cobalt and nickel resistance of a bacterial strain isolated from a Cuban serpentine deposit were investigated. This strain C-1 was assigned to Serratia marcescens by 16S rDNA analysis and DNA/DNA hybridization. Genes involved in metal resistance were identified by transposon mutagenesis followed by selection for cobalt- and nickel-sensitive derivatives.

Erwinia tasmaniensis sp. nov., a non-phytopathogenic bacterium from apple and pear trees.

Bacteria were isolated from flowers and bark of apple and pear trees at three places in Australia. In Victoria, Tasmania and Queensland, strains with white colonies on nutrient agar were screened for dome-shaped colony morphology on agar with sucrose and were found to be closely related by several criteria. The isolates were not pathogenic on apples or pears.

S434F in NrdE generates the thermosensitive phenotype of corynebacterium ammoniagenes CH31 and enhances thermolability by increasing the surface hydrophobicity of the NrdE(Ts) protein.

The thermosensitive phenotype of strain CH31, a derivative of Corynebacterium ammoniagenes ATCC 6872, was allocated by cloning, sequencing, and genetic complementation to a single C-->T exchange in the nrdE (nucleotide reduction) gene at nucleotide 1301. Protein modeling indicates the impaired surface hydrophobicity of NrdE(Ts) due to the S434F transition.

Plant-microbe interactions: identification of epiphytic bacteria and their ability to alter leaf surface permeability.

Bacteria were either isolated from leaf surfaces of Hedera helix or obtained from a culture collection in order to analyse their effect on barrier properties of isolated Hedera and Prunus laurocerasus cuticles. On the basis of the 16S rDNA sequences the genera of the six bacterial isolates from Hedera were identified as Pseudomonas sp., Stenotrophomonas sp.

Overproduction of NAD+ and 5'-inosine monophosphate in the presence of 10 microM Mn2+ by a mutant of Corynebacterium ammoniagenes with thermosensitive nucleotide reduction (nrd(ts)) after temperature shift.

Corynebacterium ammoniagenes strain CH31 is thermosensitive due to a mutation in nucleotide reduction ( nrd(ts)). The strain was examined for nucleotide overproduction upon shifting the culture temperature to a range of elevated temperatures. No overproduction of NAD(+) was detected in the control maintained at 27 degrees C whereas NAD(+) was accumulated extracellularily by strain CH31 at 37 degrees C and at 40 degrees C.

Arrest of cell cycle by inhibition of ribonucleotide reductase induces accumulation of NAD+ by Mn2+-supplemented growth of Corynebacterium ammoniagenes.

Cell division of the wild type strain Corynebacterium (formerly Brevibacterium) ammoniagenes ATCC 6872 which requires 1 microM Mn2+ for balanced growth was inhibited by addition of 20 mM hydroxyurea (HU) or 10 mM p-methoxyphenol (MP) to a Mn2+-supplemented fermentation medium at an appropriate time. Scanning electron microscopy (SEM) showed a restricted elongation characteristic of arrest of the cell cycle in coryneform bacteria. The cultures treated with HU or MP had, respectively, a fourfold or sixfold enhanced accumulation of NAD+ by a salvage biosynthetic pathway.

Isolation of toxigenic Nocardiopsis strains from indoor environments and description of two new Nocardiopsis Species, N. exhalans sp. nov. and N. umidischolae sp. nov.

Nocardiopsis strains were isolated from water-damaged indoor environments. Two strains (N. alba subsp.

Diversity of polyamine patterns in soft rot pathogens and other plant-associated members of the Enterobacteriaceae.

Polyamine profiles of 91 pectolytic and other plant-associated strains from 30 taxa of the Enterobacteriaceae were obtained by gradient high performance liquid chromatography (HPLC). Pectobacterium carotovorum, basonym Erwinia carotovora, contained a high amount of putrescine and less diaminopropane. Diaminopropane was absent in Pectobacterium chrysanthemi, basonym E.

Isolation and characterization of highly thermophilic xylanolytic Thermus thermophilus strains from hot composts.

This is the first detailed report of xylanolytic activity in Thermus strains. Two highly thermophilic xylanolytic bacteria, very closely related to non-xylanolytic T. thermophilus strains, have been isolated from the hottest zones of compost piles.

Ribonucleotide reductase (RNR) of Corynebacterium glutamicum ATCC 13032--genetic characterization of a second class IV enzyme.

Ribonucleotide reductases (RNRs) encoded by nrd (nucleotide reduction) genes are unique enzymes providing the DNA precursors in all living organisms and several viruses. The designation of four classes of RNRs reflects their use of diverse metallo-cofactors. Using oligonucleotide primers derived from conserved domains of the primary structure of known NrdA and NrdE proteins, an internal 938 bp fragment of the nrdE gene was amplified from genomic DNA of Corynebacterium glutamicum.

Thauera mechernichensis sp. nov., an aerobic denitrifier from a leachate treatment plant.

A heterotrophic bacterial strain TL1T capable of aerobic denitrification was previously enriched in continuous culture from a landfill leachate treatment plant and isolated as a pure culture. The taxonomic position of this isolate within the beta-subclass of the Proteobacteria was determined by 16S rDNA sequence analysis and by conventional taxonomy including substrate spectrum, quinone type (ubiquinone Q-8) and cellular fatty acid composition. Detection of the specific polyamine 2-hydroxyputrescine supports the membership of strain TL1T in the beta-subclass of the Proteobacteria.

Defluvibacter lusatiae gen. nov., sp. nov., a new chlorohenol-degrading member of the alpha-2 subgroup of proteobacteria.

The two Gram-negative bacterial strains S1 and S4 were isolated from activated sludge of an industrial waste water treatment plant and exhibited a stable capability to degrade 2,4-dichlorophenol, 4-chloro-2-methylphenol, 4-chlorophenol and phenol. The cells were short rods with a polar flagellum, being mesophilic, strictly aerobic, oxidase-positive, and chemoorganotrophic. They utilized a range of amino acids, but only a restricted number of carbohydrates.

Description of Pseudaminobacter gen. nov. with two new species, Pseudaminobacter salicylatoxidans sp. nov. and Pseudaminobacter defluvii sp. nov.

An aerobic bacterium, strain BN12T, which degrades substituted naphthalenesulfonates and substituted salicylates was isolated from a 6-aminonaphthalene-2-sulfonate-degrading microbial consortium originating from the River Elbe, Germany. Chemotaxonomic investigations of quinones, polyamines and polar lipids allowed allocation of this strain to the alpha-subclass of the Proteobacteria and revealed similarity to species of the genera Aminobacter, Chelatobacter and Mesorhizobium. This was confirmed by typing with 16S rRNA-targeted oligonucleotide probes and 16S rDNA sequencing and phylogenetic analysis, indicating that BN12T clusters most closely with a strain 'Thiobacillus' THI 051T and with the above genera but comprising a separate branch.

Phylogenetic diversity, polyamine pattern and DNA base composition of members of the order Planctomycetales.

The 16S rDNA sequences of 20 novel isolates of members of the order Planctomycetales were compared to those of the type strains of described planctomycete species and 22 planctomycete isolates for which the 16S rDNA sequences had been previously determined. The novel isolates could be assigned to several phylogenetically broad groups, four of which are defined by the genera Gemmata, Isosphaera, Planctomyces and Pirellula. To evaluate polyamines as a chemotaxonomic marker within this order, the polyamine pool was determined for six planctomycete reference species and for 20 planctomycete isolates.

Ribonucleotide reductase in Bacillus subtilis--evidence for a Mn-dependent enzyme.

The reduction of 2'-ribonucleotides to 2'-deoxyribonucleotides, a unique step in DNA formation, is catalyzed by ribonucleotide reductase (RRase), an allosterically regulated, cell cycle-dependent enzyme. This work reports a reversible impairment of DNA formation and ribonucleotide reduction upon manganese depletion in Bacillus subtilis demonstrated through in vivo labeling with necleic acid precursors and enzyme assays with ether-permeabilized cells. No deoxyadenosylcobalamin-dependent reduction of ribonucleotides was detected in the cytosol, and the properties of a partially purified enzyme fraction, i.

A divalent metal site in the small subunit of the manganese-dependent ribonucleotide reductase of Corynebacterium ammoniagenes.

Based on its metallo-cofactor, the manganese-dependent ribonucleotide reductase (Mn-RRase) responsible for delivery of DNA precursors in the Mn-requiring Gram-positive bacterium Corynebacterium (formerly Brevibacterium) ammoniagenes ATCC 6872 is no longer considered as a simple analogue of the aerobic Fe-RRase of Escherichia coli but as the prototype of the class IV enzymes (1). Deliberate dissociation of the Mn-RRase holoenzyme and an improved sample preparation of the dimeric CA2 protein allowed further characterization of the inherent metallo-cofactor by Q-band electron paramagnetic resonance (EPR) spectroscopy. At 40 K, a distinct hyperfine sextet (I = 5/2,55Mn) pattern with a weak zero-field splitting was detected in the CA2 protein prepared from manganese-sufficient cells displaying high RRase activity as expected.

Temperature-sensitive mutants of Corynebacterium ammoniagenes ATCC 6872 with a defective large subunit of the manganese-containing ribonucleotide reductase.

Chemical mutagenesis of the nucleotide-producing strain Corynebacterium ammoniagenes ATCC 6872 with N-methyl-N-nitro-N-nitrosoguanidine followed by an enrichment protocol yielded 46 temperature-sensitive (ts) clones. A rapid assay for the allosterically regulated Mn-ribonucleotide reductase (RRase) was developed with nucleotide-permeable cells of C. ammoniagenes in order to screen for possible defects in DNA precursor biosynthesis at elevated temperature.

Detection of a stable free radical in the B2 subunit of the manganese ribonucleotide reductase (Mn-RRase) of Corynebacterium ammoniagenes.

Ribonucleotide reductases catalyze the irreversible reductive formation of 2'-deoxyribonucleotides required for DNA replication and cell proliferation, and a radical mechanism was assumed to be involved in this reaction. In order to search for a radical in the aerobic manganese ribonucleotide reductase (Mn-RRase) by electron paramagnetic resonance (EPR) the native metal-containing 100 kDa B2 subunit was deliberately prepared from the wild type strain Corynebacterium ammoniagenes ATCC 6872. Enrichment by 2'5'-ADP Sepharose 4B affinity chromatography, fast protein liquid chromatography (FPLC) with SuperoseTM12 and concentration by vacuum evaporation allowed for the first time the detection of a stable free radical by EPR spectroscopy at 77 K.