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  • American Society for Microbiology  (6)
  • Weidenbach, Katrin  (6)
  • 1
    In: Journal of Bacteriology, American Society for Microbiology, Vol. 202, No. 8 ( 2020-03-26)
    Abstract: Archaeosine (G + ) is a structurally complex modified nucleoside found quasi-universally in the tRNA of Archaea and located at position 15 in the dihydrouridine loop, a site not modified in any tRNA outside the Archaea . G + is characterized by an unusual 7-deazaguanosine core structure with a formamidine group at the 7-position. The location of G + at position 15, coupled with its novel molecular structure, led to a hypothesis that G + stabilizes tRNA tertiary structure through several distinct mechanisms. To test whether G + contributes to tRNA stability and define the biological role of G + , we investigated the consequences of introducing targeted mutations that disrupt the biosynthesis of G + into the genome of the hyperthermophilic archaeon Thermococcus kodakarensis and the mesophilic archaeon Methanosarcina mazei , resulting in modification of the tRNA with the G + precursor 7-cyano-7-deazaguansine (preQ 0 ) (deletion of arcS ) or no modification at position 15 (deletion of tgtA ). Assays of tRNA stability from in vitro -prepared and enzymatically modified tRNA transcripts, as well as tRNA isolated from the T. kodakarensis mutant strains, demonstrate that G + at position 15 imparts stability to tRNAs that varies depending on the overall modification state of the tRNA and the concentration of magnesium chloride and that when absent results in profound deficiencies in the thermophily of T. kodakarensis . IMPORTANCE Archaeosine is ubiquitous in archaeal tRNA, where it is located at position 15. Based on its molecular structure, it was proposed to stabilize tRNA, and we show that loss of archaeosine in Thermococcus kodakarensis results in a strong temperature-sensitive phenotype, while there is no detectable phenotype when it is lost in Methanosarcina mazei . Measurements of tRNA stability show that archaeosine stabilizes the tRNA structure but that this effect is much greater when it is present in otherwise unmodified tRNA transcripts than in the context of fully modified tRNA, suggesting that it may be especially important during the early stages of tRNA processing and maturation in thermophiles. Our results demonstrate how small changes in the stability of structural RNAs can be manifested in significant biological-fitness changes.
    Type of Medium: Online Resource
    ISSN: 0021-9193 , 1098-5530
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2020
    detail.hit.zdb_id: 1481988-0
    SSG: 12
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  • 2
    Online Resource
    Online Resource
    American Society for Microbiology ; 2011
    In:  Applied and Environmental Microbiology Vol. 77, No. 24 ( 2011-12-15), p. 8669-8675
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 77, No. 24 ( 2011-12-15), p. 8669-8675
    Abstract: In spite of the significant impact of biomethylation on the mobility and toxicity of metals and metalloids in the environment, little is known about the biological formation of these methylated metal(loid) compounds. While element-specific methyltransferases have been isolated for arsenic, the striking versatility of methanoarchaea to methylate numerous metal(loid)s, including rare elements like bismuth, is still not understood. Here, we demonstrate that the same metal(loid)s (arsenic, selenium, antimony, tellurium, and bismuth) that are methylated by Methanosarcina mazei in vivo are also methylated by in vitro assays with purified recombinant MtaA, a methyltransferase catalyzing the methyl transfer from methylcobalamin [CH 3 Cob(III)] to 2-mercaptoethanesulfonic acid (CoM) in methylotrophic methanogenesis. Detailed studies revealed that cob(I)alamin [Cob(I)] , formed by MtaA-catalyzed demethylation of CH 3 Cob(III), is the causative agent for the multimetal(loid) methylation observed. Moreover, Cob(I) is also capable of metal(loid) hydride generation. Global transcriptome profiling of M. mazei cultures exposed to bismuth did not reveal induced methyltransferase systems but upregulated regeneration of methanogenic cofactors in the presence of bismuth. Thus, we conclude that the multimetal(loid) methylation in vivo is attributed to side reactions of CH 3 Cob(III) with reduced cofactors formed in methanogenesis. The close connection between metal(loid) methylation and methanogenesis explains the general capability of methanoarchaea to methylate metal(loid)s.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2011
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 3
    In: Journal of Virology, American Society for Microbiology, Vol. 91, No. 22 ( 2017-11-15)
    Abstract: A novel archaeal lytic virus targeting species of the genus Methanosarcina was isolated using Methanosarcina mazei strain Gö1 as the host. Due to its spherical morphology, the virus was designated Met hanosarcina s pherical v irus (MetSV). Molecular analysis demonstrated that MetSV contains double-stranded linear DNA with a genome size of 10,567 bp containing 22 open reading frames (ORFs), all oriented in the same direction. Functions were predicted for some of these ORFs, i.e., such as DNA polymerase, ATPase, and DNA-binding protein as well as envelope (structural) protein. MetSV-derived spacers in CRISPR loci were detected in several published Methanosarcina draft genomes using bioinformatic tools, revealing a potential protospacer-adjacent motif (PAM) motif (TTA/T). Transcription and expression of several predicted viral ORFs were validated by reverse transcription-PCR (RT-PCR), PAGE analysis, and liquid chromatography-mass spectrometry (LC-MS)-based proteomics. Analysis of core lipids by atmospheric pressure chemical ionization (APCI) mass spectrometry showed that MetSV and Methanosarcina mazei both contain archaeol and glycerol dialkyl glycerol tetraether without a cyclopentane moiety (GDGT-0). The MetSV host range is limited to Methanosarcina strains growing as single cells ( M. mazei , Methanosarcina barkeri and Methanosarcina soligelidi ). In contrast, strains growing as sarcina-like aggregates were apparently protected from infection. Heterogeneity related to morphology phases in M. mazei cultures allowed acquisition of resistance to MetSV after challenge by growing cultures as sarcina-like aggregates. CRISPR/Cas-mediated resistance was excluded since neither of the two CRISPR arrays showed MetSV-derived spacer acquisition. Based on these findings, we propose that changing the morphology from single cells to sarcina-like aggregates upon rearrangement of the envelope structure prevents infection and subsequent lysis by MetSV. IMPORTANCE Methanoarchaea are among the most abundant organisms on the planet since they are present in high numbers in major anaerobic environments. They convert various carbon sources, e.g., acetate, methylamines, or methanol, to methane and carbon dioxide; thus, they have a significant impact on the emission of major greenhouse gases. Today, very little is known about viruses specifically infecting methanoarchaea that most probably impact the abundance of methanoarchaea in microbial consortia. Here, we characterize the first identified Methanosarcina -infecting virus (MetSV) and show a mechanism for acquiring resistance against MetSV. Based on our results, we propose that growth as sarcina-like aggregates prevents infection and subsequent lysis. These findings allow new insights into the virus-host relationship in methanogenic community structures, their dynamics, and their phase heterogeneity. Moreover, the availability of a specific virus provides new possibilities to deepen our knowledge of the defense mechanisms of potential hosts and offers tools for genetic manipulation.
    Type of Medium: Online Resource
    ISSN: 0022-538X , 1098-5514
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2017
    detail.hit.zdb_id: 1495529-5
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  • 4
    In: mSystems, American Society for Microbiology, Vol. 7, No. 2 ( 2022-04-26)
    Type of Medium: Online Resource
    ISSN: 2379-5077
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 2844333-0
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  • 5
    In: mSystems, American Society for Microbiology, Vol. 7, No. 1 ( 2022-02-22)
    Abstract: Raman microspectroscopy has been used to thoroughly assess growth dynamics and heterogeneity of prokaryotic cells, yet little is known about how the chemistry of individual cells changes during infection with virulent viruses, resulting in so-called virocells. Here, we investigate biochemical changes of bacterial and archaeal cells of three different species in laboratory cultures before and after addition of their respective viruses using single-cell Raman microspectroscopy. By applying multivariate statistics, we identified significant differences in the spectra of single cells with/without addition of virulent dsRNA phage ( phi6 ) for Pseudomonas syringae . A general ratio of wavenumbers that contributed the greatest differences in the recorded spectra was defined as an indicator for virocells. Based on reference spectra, this difference is likely attributable to an increase in nucleic acid versus protein ratio of virocells. This method also proved successful for identification of Bacillus subtilis cells infected with the double-stranded DNA (dsDNA) phage phi29 , displaying a decrease in respective ratio, but failed for archaeal virocells ( Methanosarcina mazei with the dsDNA methanosarcina spherical virus) due to autofluorescence. Multivariate and univariate analyses suggest that Raman spectral data of infected cells can also be used to explore the complex biology behind viral infections of bacteria. Using this method, we confirmed the previously described two-stage infection of P. syringae ’s phi6 and that infection of B. subtilis with phi29 results in a stress response within single cells. We conclude that Raman microspectroscopy is a promising tool for chemical identification of Gram-positive and Gram-negative virocells undergoing infection with virulent DNA or RNA viruses. IMPORTANCE Viruses are highly diverse biological entities shaping many ecosystems across Earth. However, understanding the infection of individual microbial cells and the related biochemical changes remains limited. Using Raman microspectroscopy in conjunction with univariate and multivariate statistics, we established a marker for identification of infected Gram-positive and Gram-negative bacteria. This nondestructive, label-free analytical method at single-cell resolution paves the way for future studies geared towards analyzing virus-host systems of prokaryotes to further understand the complex chemistry and function of virocells.
    Type of Medium: Online Resource
    ISSN: 2379-5077
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 2844333-0
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  • 6
    Online Resource
    Online Resource
    American Society for Microbiology ; 2022
    In:  Journal of Bacteriology Vol. 204, No. 1 ( 2022-01-18)
    In: Journal of Bacteriology, American Society for Microbiology, Vol. 204, No. 1 ( 2022-01-18)
    Abstract: In recent years, increasing numbers of small proteins have moved into the focus of science. Small proteins have been identified and characterized in all three domains of life, but the majority remains functionally uncharacterized, lack secondary structure, and exhibit limited evolutionary conservation. While quite a few have already been described for bacteria and eukaryotic organisms, the amount of known and functionally analyzed archaeal small proteins is still very limited. In this review, we compile the current state of research, show strategies for systematic approaches for global identification of small archaeal proteins, and address selected functionally characterized examples. Besides, we document exemplarily for one archaeon the tool development and optimization to identify small proteins using genome-wide approaches.
    Type of Medium: Online Resource
    ISSN: 0021-9193 , 1098-5530
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 1481988-0
    SSG: 12
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