Markerless Escherichia coli rrn Deletion Strains for Genetic Determination of Ribosomal Binding Sites

Single-copy rrn strains facilitate genetic ribosomal studies in Escherichia coli. Consecutive markerless deletion of rrn operons resulted in slower growth upon inactivation of the fourth copy, which was reversed by supplying transfer RNA genes encoded in rrn operons in trans. Removal of the sixth, penultimate rrn copy led to a reduced growth rate due to limited rrn gene dosage. Whole-genome sequencing of variants of single-copy rrn strains revealed duplications of large stretches of genomic DNA. The combination of selective pressure, resulting from the decreased growth rate, and the six identical remaining scar sequences, facilitating homologous recombination events, presumably leads to elevated genomic instability

The role of magnesium and calcium ions in the glucose dehydrogenase activity of <i>Klebsiella pneumoniae</i> NCTC 418

Magnesium-limited chemostat cultures of Klebsiella pneumoniae NCTC 418 with 20 μM CaCl2 in the medium showed a low rate of gluconate plus 2-ketogluconate production relative to potassium- or phosphate-limited cultures. However, when the medium concentration of CaCl2 was increased to 1 mM, the glucose dehydrogenase (GDH) activities also increased and became similar to those observed in potassium- or phosphate limited cultures. It is concluded that this is due to Mg2+ and Ca2+ ions being involved in the binding of pyrroloquinoline quinone (PQQ) to the GDH apoenzyme. There seems to be an absolute requirement of divalent cations for proper enzyme functioning and in this respect Ca2+ ions could replace Mg2+ ions. The high GDH activity which has been found in cells grown under Mg2−-limited conditions in the presence of higher concentrations of Ca2+ ions, is compatible with the earlier proposal that GDH functions as an auxiliary energy generating system involved in the maintenance of high transmembrane ion gradients.Centro de Investigación y Desarrollo en Fermentaciones Industriale

Immune sensing of Candida albicans requires cooperative recognition of mannans and glucans by lectin and Toll-like receptors

Peer reviewedPublisher PD

Different Modes of Action of Naphthyridones in Gram-Positive and Gram-Negative Bacteria

Naphthyridones that were recently described as a class of translation inhibitors in gram-positive bacteria mediate their mode of action via GyrA in Haemophilus influenzae and Escherichia coli. These are the first examples of compounds in which modes of action in different bacterial pathogens are mediated through widely different targets

Utilization of Target-Specific, Hypersensitive Strains of Saccharomyces cerevisiae To Determine the Mode of Action of Antifungal Compounds

Target-specific hypersusceptible strains of Saccharomyces cerevisiae were used to screen antifungal compounds. Two novel Erg7p inhibitors were identified, providing proof of principle of the approach taken. However, observed hypersensitivities to antifungals acting via other targets imply that use of this tool to identify the mode of action requires significant deconvolution

Antimicrobial Activity of Adenine-Based Inhibitors of NAD⁺-Dependent DNA Ligase

The relationship between enzyme inhibition and antimicrobial potency of adenine-based NAD⁺-dependent DNA ligase (LigA) inhibitors was investigated using a strain of the Gram-negative pathogen <i>Haemophilus influenzae</i> lacking its major AcrAB-TolC efflux pump and the Gram-positive pathogen <i>Streptococcus pneumoniae</i>. To this end, biochemical inhibitors not mediating their antibacterial mode of action (MOA) via LigA were removed from the analysis. In doing so, a significant number of compounds were identified that acted via inhibition of LigA in <i>S. pneumoniae</i> but not in <i>H. influenzae</i>, despite being inhibitors of both isozymes. Deviations from the line correlating antimicrobial and biochemical potencies of LigA inhibitors with the correct MOA were observed for both species. These deviations, usually corresponding to higher MIC/IC₅₀ ratios, were attributed to varying compound permeance into the cell