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Barrera Quinn posted an update 6 months, 2 weeks ago
the physiological and metabolic characteristics is critical. Caldicellulosiruptor bescii and other species in this genus have untapped potential for conversion of unpretreated plant biomass into industrial fuels and chemicals. The highly interactive and complex machinery used by C. bescii to acquire and process complex carbohydrates contained in lignocellulose was elucidated here to complement related efforts to develop a metabolic engineering platform with this bacterium. Guided by the findings here, a clearer picture of how C. bescii natively drives carbohydrate utilization is provided and strategies to engineer this bacterium for optimal conversion of lignocellulose to commercial products emerge.Prophage integration, release, and dissemination exert various effects on host bacteria. In the genus Lactobacillus, they may cause bacteriophage contamination during fermentation and even regulate bacterial populations in the gut. However, little is known about their distribution, genetic architecture, and relationships with their hosts. Here, we conducted prophage prediction analysis on 1,472 genomes from 16 different Lactobacillus species and found prophage fragments in almost all lactobacilli (99.8%), with 1,459 predicted intact prophages identified in 64.1% of the strains. We present an uneven prophage distribution among Lactobacillus species; multihabitat species retained more prophages in their genomes than restricted-habitat species. Characterization of the genome features, average nucleotide identity, and landscape visualization presented a high genome diversity of Lactobacillus prophages. We detected antibiotic resistance genes in more than 10% of Lactobacillus prophages and validated that the occurn genome feature, integration site, and genomic organization. Our data of the prophage-mediated antibiotic resistance genes (ARGs) and the resistance phenotype of lactobacilli provide evidence for deciphering the putative role of prophages as vectors of the ARGs. Furthermore, understanding the association between prophages and CRISPR-Cas systems is crucial to appreciate the coevolution of phages and Lactobacillus.Proteins are major contributors to the composition and the functions in the cell. They often assemble into larger structures, macromolecular machines, to carry out intricate essential functions. Although huge progress in understanding how macromolecular machines function has been made by reconstituting them in vitro, the role of the intracellular environment is still emerging. The development of fluorescence microscopy techniques in the last 2 decades has allowed us to obtain an increased understanding of proteins and macromolecular machines in cells. Here, we describe how proteins move by diffusion, how they search for their targets, and how they are affected by the intracellular environment. We also describe how proteins assemble into macromolecular machines and provide examples of how frequent subunit turnover is used for them to function and to respond to changes in the intracellular conditions. This review emphasizes the constant movement of molecules in cells, the stochastic nature of reactions, and the dynamic nature of macromolecular machines.Bacteriophage serine integrases catalyze highly specific recombination reactions between defined DNA segments called att sites. These reactions are reversible depending upon the presence of a second phage-encoded directionality factor. The bipartite C-terminal DNA-binding region of integrases includes a recombinase domain (RD) connected to a zinc-binding domain (ZD), which contains a long flexible coiled-coil (CC) motif that extends away from the bound DNA. We directly show that the identities of the phage A118 integrase att sites are specified by the DNA spacing between the RD and ZD DNA recognition determinants, which in turn directs the relative trajectories of the CC motifs on each subunit of the att-bound integrase dimer. Saracatinib in vivo Recombination between compatible dimer-bound att sites requires minimal-length CC motifs and 14 residues surrounding the tip where the pairing of CC motifs between synapsing dimers occurs. Our alanine-scanning data suggest that molecular interactions between CC motif tips may differ in minants regulating synaptic complex formation between correct DNA sites, including the DNA architecture responsible for specifying the identity of recombination sites, features of the unique coiled-coil structure on the integrase that are required to initiate synapsis, and amino acid residues on the integrase oligomerization helix that control the remodeling of synapsing dimers into a tetramer active for DNA strand exchange.Resistance in VanA-type vancomycin-resistant Enterococcus faecium (VREfm) is due to an inducible gene cassette encoding seven proteins (vanRSHAXYZ). This provides for an alternative peptidoglycan (PG) biosynthesis pathway whereby D-Ala-D-Ala is replaced by D-Ala-d-lactate (Lac), to which vancomycin cannot bind effectively. This study aimed to quantify cytoplasmic levels of normal and alternative pathway PG intermediates in VanA-type VREfm by liquid chromatography-tandem mass spectrometry before and after vancomycin exposure and to correlate these changes with changes in vanA operon mRNA levels measured by real-time quantitative PCR (RT-qPCR). Normal pathway intermediates predominated in the absence of vancomycin, with low levels of alternative pathway intermediates. Extended (18-h) vancomycin exposure resulted in a mixture of the terminal normal (UDP-N-acetylmuramic acid -l-Ala-D-Glu-l-Lys-D-Ala-D-Ala ) and alternative (UDP-NAM-l-Ala-γ-D-Glu-l-Lys-D-Ala-D-Lac ) pathway intermedways. IMPORTANCE VREfm is highly resistant to vancomycin due to the presence of a vancomycin resistance gene cassette. Exposure to vancomycin induces the expression of genes in this cassette, which encode enzymes that provide for an alternative PG biosynthesis pathway. In VanA-type resistance, these alternative pathway enzymes replace the D-Ala-D-Ala terminus of normal PG intermediates with D-Ala-D-Lac terminated intermediates, to which vancomycin cannot bind. While the general features of this resistance mechanism are well known, the details of the choreography between vancomycin exposure, vanA gene induction, and changes in the normal and alternative pathway intermediate levels have not been described previously. This study comprehensively explores how VREfm responds to vancomycin exposure at the mRNA and PG intermediate levels.
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