- Volume 141, Issue 3, 1995
Volume 141, Issue 3, 1995
- Physiology And Growth
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Carbon-arsenic bond cleavage by a newly isolated Gram-negative bacterium, strain ASV2
More LessSummary: Strain ASV2, an unidentified Gram-negative bacterium newly isolated from activated sludge, was found to utilize arsonoacetate at concentrations up to at least 30 mM as sole carbon and energy source, with essentially quantitative extracellular release of arsenate. Cell-free conversion of arsonoacetate could not be obtained, but resting-cell studies indicated that the carbon-arsenic bond cleavage activity was inducible in the presence of arsonoacetate and was of limited substrate specificity, also breaking down arsonochloroacetate. The inorganic product of the reaction may be arsenite since an inducible arsenite-oxidizing activity was also found in arsonoacetate-metabolizing cells. This is the first report of a micro-organism capable of utilizing a compound containing the carbon-arsenic bond. The results indicate that the ability of bacteria to degrade arsonoacetate is not fortuitous and may be found in environments not previously exposed to organoarsenicals.
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23Na NMR spectroscopy of free Na+ in the halotolerant bacterium Brevibacterium sp. and Escherichia coli
More LessSummary:23Na NMR spectroscopy was used to determine free Na+ concentrations in a halotolerant bacterium, Brevibacterium sp., and Escherichia coli. The internal Na+ concentration of both strains depended little on the growth phases and was unchanged after 5 d storage at 2°C. In Brevibacterium sp. the level of intracellular sodium increased gradually at higher extracellular NaCI concentrations in both the presence and absence of yeast extract in the growth medium. E. coli cells accumulated a higher concentration of free Na+ than those of Brevibacterium sp. The change of Na+ concentration in both strains was inverse to that of growth rate. When appropriate amounts of osmoprotectants (proline, glycine betaine, or γ-aminobutyrate) were added with the NaCI, internal free Na+ levels in Brevibacterium sp. were lowered, but those of E. coli were unchanged. While addition of KCl to medium containing NaCI increased the intracellular level of free Na+, the total sodium concentration in the cells remained unchanged, indicating that sodium that had been bound or attached was made free in the cytosol. In Brevibacterium sp. grown in the presence of 0.5 M NaCI, free and bound sodium concentrations in the cytosol were estimated to be 0.14 and 0.23 μmol (mg protein)-1, respectively. As a result, visibility by 23Na NMR was 38%.
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Hemicellulolytic enzymes in P- and S-strains of Heterobasidion annosum
More LessSummary: The secretion of several hemicellulolytic enzymes and endoglucanase was studied in S- and P-group strains of a wood-rotting pathogen, Heterobasidio annosum, using enzyme activity assays and isoelectric focusing. In liquid cultures supplemented with birch sawdust or pure xylan, both strains of the fungus produced xylanases, mannanases, endoglucanases and α-galactosidases. Low activities were detected in cultures containing only glucose as a carbon source. When S-strain cultures were provided with a cart matrix, protein secretion to the culture medium started earlier and was significantly increased compared to free cell cultures. This was not the case with the P-strain. In both strains of H. annosum the pH of the birch sawdust culture medium fell within 12 d to below 3.5, and then started to increase towards the end of the cultivation period. The increase in pH appeared to corresprelate with the formation of hemicellulolytic and endoglucanase activitie. The isoenzyme patterns of xylanase, mannanase and endoglucanase were typical for P- and S-strains. In P-strain cultures containing xylan, a xylanase with an alkaline pl value was induced which, however, was absent when the strain was grown with the birch sawdust substrate. In the P-strain the mannanase pattern at the early stage of growth was comparable to that of t S-strain at the later stages of growth on birch sawdust medium. Later in the growth of the P-strain, this pattern was replaced by mannanases with more acid pl values. The relative intensities of the individual endoglucanases varie between the P- and S-strains.
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- Plant-Microbe Interactions
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Regulation of pectin methylesterase and polygalacturonate lyase activity during differentiation of infection structures in Uromyces viciae-fabae
More LessSummaryThe broad bean rust fungus Uromyces viciae-fabae differentiates infection structures up to the haustorial mother cell stage on thigmotropically inductive membranes in the absence of its host plant. Formation of pectin methylesterase (PME) and polygalacturonate lyase (PL), potentially involved in host cell wall degradation, was studied during infection structure differentiation by this biotrophic fungus. PME was first detectable when substomatal vesicles were formed and reached a maximum when infection hyphae and haustorial mother cells were differentiated. Four isoenzymes, exhibiting pls of 8·2, 5·6, 5·2 and 4·5, were separated by chromatofocusing, and the kinetics of their synthesis and the K ms of the three major isoenzymes were determined. The enzyme activity was formed independently of the presence of its substrate and its regulation was thus differentiation-specific. A single PL was induced when haustorial mother cells were formed and its synthesis appeared to be controlled by both the developmental stage of infection structures and the availability of its substrate. Polygalacturonate concentrations lower than 0·025 mg ml−1 induced enzyme synthesis, and at 0·25 mg ml-1 the induction process appeared to be saturated. Enzyme formation in the presence of 50 mM glucose, fructose or sucrose suggested that neither pectic enzyme was subject to catabolite repression. Significant proportions of PME (approx. 57 %) and PL (approx. 76 %) activity were located extracellularly in 24-h-old differentiated infection structures and could contribute to the establishment of the parasite. Physico-chemical and kinetic properties of the enzymes and associated alterations of the apoplastic pH of infected host plants appeared to be important factors in the success of infection and could explain the restriction of cell wall damage at the penetration site usually observed in interactions involving obligately biotrophic fungi.
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Surface polysaccharide mutants of Rhizobium sp. ( Acacia ) strain GRH2: major requirement of lipopolysaccharide for successful invasion of Acacia nodules and host range determination
More LessSummary: Two transposon Tn5-induced mutants of wild-type broad-host-range Rhizobium sp. GRH2 were isolated and found to harbour different alterations in surface polysaccharides. These mutants, designated GRH2-14 and GRH2-50, induced a few, empty nodules on Acacia and lost the ability to nodulate most host herbaceous legumes. Whereas mutant GRH2-14 produces an acidic exopolysaccharide (EPS) similar to the wild-type, the acidic EPS of mutant GRH2-50 lacks galactose and the pyruvyl and 3-hydroxybutyryl substituents attached to this sugar moiety. In addition, both mutants GRH2-50 and GRH2-14 were altered in smooth lipopolysaccharides (LPS). DNA sequence analyses of the correspresponding Tn5 insertions revealed that strain GRH2-50 was mutated in a DNA locus homologous to galE , and in vitro enzyme assays indicated that the UDPglucose 4-epimerase (GalE) activity was missing in this mutant strain. DNA hybridization studies showed that the GRH2-50 mutant DNA has homologous sequences within the different biovars of Rhizobium leguminosarum. However, no DNA homology to GRH2-14 altered DNA was found in those rhizobial strains, indicating that it represents a new chromosomal lps locus in Rhizobium sp. ( Acacia) involved in symbiotic development.
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- Genome Analysis
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The Bacillus subtilis dnaC gene encodes a protein homologous to the DnaB helicase of Escherichia coli
More LessSummary: Within the region of the Bacillus subtilis chromosome assigned to us in the genome sequencing project, we found a gene, the product of which is similar to the DnaB protein (replicative DNA helicase) of Escherichia coli. Three B. subtilis dna gene mutations, dnaC30 and ts56 causing defects in elongation and ts199 causing a defect in the initiation of replication, were mapped in the gene by transformation and DNA sequencing. Both dnaC30 and ts56 have been located near the amino-termina-l end of the B. subtilis DnaC protein. In contrast, ts199 has been located near the carboxy-terminal of the protein. Our results indicate that the B. subtilis dnaC gene encodes a counterpart of the E. coli dnaB helicase.
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A putative new peptide synthase operon in Bacillus subtilis : partial characterization
Summary:A large operon-type structure has been located between the gltA and citB loci on the Bacillus subtilis chromosome. On the basis of the analysis of the 25 kb sequenced so far, it potentially encodes at least three large proteins which contain structural motifs associated with the subunits of all characterized peptide synthases. The amino acid recognition specificity of this new peptide synthase is discussed in the light of sequence homology with other synthases.
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Mini-chromosomes in Fusarium sporotrichioides are mosaics of dispersed repeats and unique sequences
More LessSummary:Variations in trichothecene patterns of 26 Fusarium sporotrichioides isolates from different plant and geographic origins showed no corresprelation with electrophoretic karyotype polymorphisms. When intact chromosomes were examined, interisolate karyotype differences were observed only in the mini-chromosome range. Further polymorphisms were revealed in Notl-digested samples. By summing the Notl fragments the average genome size of F. sporotrichioides was estimated to be 20.4 Mb. Mini-chromosomes shared common sequences with the larger ones; however, clones (RMS-1 and RMS-2) specific to these structures have also been found. These clones contained no coding region and no promising similarities were observed when they were compared to sequences held at GenBank. Mini-chromosomes in F. sporotrichioides constitute a mosaic composed of dispersed repeats and unique sequences. This mosaic structure was maintained in all non-interbreeding, genetically isolated strains examined.
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