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  • 1
    Publication Date: 2017-01-19
    Description: Although nitric oxide (NO) is an important signaling molecule in bacteria and higher organisms, excessive intracellular NO is highly reactive and dangerous. Therefore, living cells need strict regulation systems for cellular NO homeostasis. Recently, we discovered that Streptomyces coelicolor A3(2) retains the nitrogen oxide cycle (NO 3 – -〉NO 2 – -〉NO-〉NO 3 – ) and nitrite removal system. The nitrogen oxide cycle regulates cellular NO levels, thereby controlling secondary metabolism initiation (red-pigmented antibiotic, RED production) and morphological differentiation. Nitrite induces gene expression in neighboring cells, suggesting another role for this cycle as a producer of transmittable intercellular communication molecules. Here, we demonstrated that ammonium-producing nitrite reductase (NirBD) is involved in regulating NO homeostasis in S. coelicolor A3(2). NirBD was constitutively produced in culture independently of GlnR, a known transcriptional factor. NirBD cleared the accumulated nitrite from the medium. Nir deletion mutants showed increased NO-dependent gene expression at later culture stages, whereas the wild-type M145 showed decreased expression, suggesting that high NO concentration was maintained in the mutant. Moreover, the nir deletion mutant produced more RED than that produced by the wild-type M145. These results suggest that NO 2 – removal by NirBD is important to regulate NO homeostasis and to complete NO signaling in S. coelicolor .
    Keywords: Physiology & Biochemistry
    Print ISSN: 0378-1097
    Electronic ISSN: 1574-6968
    Topics: Biology
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  • 2
    Publication Date: 2017-01-13
    Description: Earlier, vitamin C was demonstrated to sterilize Mycobacterium tuberculosis culture via Fenton's reaction at high concentration. It alters the regulatory pathways associated with stress response and dormancy. Since (p)ppGpp is considered to be the master regulator of stress response and is responsible for bacterial survival under stress, we tested the effect of vitamin C on the formation of (p)ppGpp. In vivo estimation of (p)ppGpp showed a decrease in (p)ppGpp levels in vitamin C-treated M. smegmatis cells in comparison to the untreated cells. Furthermore, in vitro (p)ppGpp synthesis using Rel MSM enzyme was conducted in order to confirm the specificity of the inhibition in the presence of variable concentrations of vitamin C. We observed that vitamin C at high concentration can inhibit the synthesis of (p)ppGpp. We illustrated binding of vitamin C to Rel MSM by isothermal titration calorimetry. Enzyme kinetics was followed where K 0.5 was found to be increased with the concomitant reduction of V max value suggesting mixed inhibition. Both long-term survival and biofilm formation were inhibited by vitamin C. The experiments suggest that vitamin C has the potential to be developed as the inhibitor of (p)ppGpp synthesis and stress response, at least in the concentration range used here.
    Keywords: Physiology & Biochemistry
    Print ISSN: 0378-1097
    Electronic ISSN: 1574-6968
    Topics: Biology
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  • 3
    Publication Date: 2016-12-23
    Description: The culturability of Escherichia coli , Ralstonia eutropha and Bacillus subtilis after incubation in phosphate-buffered saline at either 5°C or 30°C was determined. The culturability of B. subtilis showed little dependence on temperature. The culturability of E. coli rapidly decreased at 30°C but remained almost constant at 5°C. In contrast, the culturability of R. eutropha decreased by three orders of magnitude at 5°C within 24 h but only moderately decreased (one order of magnitude) at 30°C. Remarkably, prolonged incubation of R. eutropha at 30°C resulted in a full recovery of colony forming units in contrast to only a partial recovery at 5°C. Ralstonia eutropha cells at 30°C remained culturable for 3 weeks while culturability at 5°C constantly decreased. The effect of temperature was significantly stronger in a polyhydroxybutyrate-negative mutant. Our data show that accumulated polyhydroxybutyrate has a cold-protective function and can prevent R. eutropha entering the viable but not culturable state.
    Keywords: Physiology & Biochemistry
    Print ISSN: 0378-1097
    Electronic ISSN: 1574-6968
    Topics: Biology
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  • 4
    Publication Date: 2016-12-16
    Description: Pseudomonas plecoglossicida is a facultative fish pathogen. Recent studies showed that P. plecoglossicida infection in fish was associated with temperature. The aim of this study was to compare the secretomes of P. plecoglossicida cultured in vitro at representative temperatures for pathogenic (20°C) and less pathogenic (30°C) phenotypes. Thirteen proteins in the culture supernatants of P. plecoglossicida showed significant difference in abundance at 20 vs. 30°C. Four proteins were strongly increased at 20°C, including two hemolysin co-regulated proteins (Hcp) that are part of the bacterial type VI secretion system (T6SS), flagellin and an unknown protein. Immunoblot analysis verified the induced secretion of Hcps at 20°C. Furthermore, the upregulation of Hcps at 20°C was confirmed at transcriptional level by RT-qPCR analysis, which also demonstrated the induction of expression of other T6SS-related genes at 20°C. Taken together, we demonstrate the presence of two functionally active T6SS proteins in fish pathogenic P. plecoglossicida strains, as evidenced by the secretion of the T6SS substrate Hcp, the production of which were found to be controlled by temperature. Our findings also support efforts to develop vaccines targeting secreted virulence factors as prophylactic strategies for diseases in fish caused by P. plecoglossicida .
    Keywords: Physiology & Biochemistry
    Print ISSN: 0378-1097
    Electronic ISSN: 1574-6968
    Topics: Biology
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  • 5
    Publication Date: 2016-10-22
    Description: The hydrocarbonoclastic bacterium Acinetobacter venetianus RAG-1 has attracted substantial attention due to its powerful oil-degrading capabilities and its potential to play an important ecological role in the cleanup of alkanes. In this study, we compare the transcriptome of the strain RAG-1 grown in dodecane, the corresponding alkanol (dodecanol), and sodium acetate for the characterization of genes involved in dodecane uptake and utilization. Comparison of the transcriptional responses of RAG-1 grown on dodecane led to the identification of 1074 genes that were differentially expressed relative to sodium acetate. Of these, 622 genes were upregulated when grown in dodecane. The highly upregulated genes were involved in alkane catabolism, along with stress response. Our data suggest AlkMb to be primarily involved in dodecane oxidation. Transcriptional response of RAG-1 grown on dodecane relative to dodecanol also led to the identification of permease, outer membrane protein and thin fimbriae coding genes potentially involved in dodecane uptake. This study provides the first model for key genes involved in alkane uptake and metabolism in A. venetianus RAG-1.
    Keywords: Physiology & Biochemistry
    Print ISSN: 0378-1097
    Electronic ISSN: 1574-6968
    Topics: Biology
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