Inhibition of D-Ala:D-Ala ligase through a phosphorylated form of the antibiotic D-cycloserine
Sarah Batson, Cesira de Chiara, Vita Majce, Adrian J Lloyd, Stanislav Gobec, Dean Rea, Vilmos Fülöp, Christopher W Thoroughgood, Katie J Simmons, Christopher G Dowson, Colin WG Fishwick, Luiz Pedro S de Carvalho & David I Roper
D-cycloserine is an antibiotic which targets sequential bacterial cell wall peptidoglycan biosynthesis enzymes: alanine racemase and D-alanine:D-alanine ligase. By a combination of structural, chemical and mechanistic studies here we show that the inhibition of D-alanine:D-alanine ligase by the antibiotic D-cycloserine proceeds via a distinct phosphorylated form of the drug. This mechanistic insight reveals a bimodal mechanism of action for a single antibiotic on different enzyme targets and has significance for the design of future inhibitor molecules based on this chemical structure.
Targeted treatment of yaws with contact tracing : how much do we miss?
Louise Dyson, Michael Marks, Oliver M Crook, Oliver Sokana, Anthony W Solomon, Alex Bishop, David CW Mabey, T Deirdre Hollingsworth
Yaws is a disabling bacterial infection primarily found in warm and humid tropical areas. The World Health Organisation strategy mandates an initial round of total community treatment (TCT) with single-dose azithromycin followed either by further TCT or active case finding and treatment of cases and their contacts (the Morges strategy). We wish to investigate the effectiveness of the Morges strategy. Our analysis demonstrates that at all prevalences present in the dataset, up to 90% of (active and asymptomatic) infections would not be treated under household-based contact tracing.
Engineering of biomolecules by bacteriophage directed evolution
Andreas K Brödel, Mark Isalan, Alfonso Jaramillo
Conventional in vivo directed evolution methods have primarily linked the biomolecule's activity to bacterial cell growth. Recent developments instead rely on the conditional growth of bacteriophages (phages), viruses that infect and replicate within bacteria. Here we review recent phage-based selection systems for in vivo directed evolution. These approaches have been applied to evolve a wide range of proteins including transcription factors, polymerases, proteases, DNA-binding proteins, and protein–protein interactions. Advances in this field expand the possible applications of protein and RNA engineering. This will ultimately result in new biomolecules with tailor-made properties, as well as giving us a better understanding of basic evolutionary processes.
Development and validation of a multiple locus variable number tandem repeat analysis (MLVA) scheme for Fusobacterium necrophorum
Fusobacterium necrophorum is associated with various diseases in humans and animals. The aim of this study was to develop multiple locus variable number tandem repeat analysis (MLVA) as a strain typing technique for F. necrophorum, and to test the use of this scheme to analyse both isolates and mixed communities of bacteriaThe results demonstrate the potential for this method to elucidate reservoirs of F. necrophorum.
Lost, but found with Nile red; a novel method to detect and quantify small microplastics (20 µm–1 mm) in environmental samples
Gabriel Erni-Cassola, Matthew I. Gibson, Richard C. Thompson, and Joseph Christie-Oleza
Marine plastic debris is a global environmental problem. Surveys have shown that plastic particles <5 mm in size, known as microplastics, are significantly more abundant in surface seawater and on shorelines than larger plastic particles. Nevertheless, quantification of microplastics in the environment is hampered by a lack of adequate high throughput methods to distinguish and quantify smaller size fractions (<1 mm), and this has probably resulted in an underestimation of actual microplastic concentrations. Here we present a protocol that allows high throughput detection and automated quantification of small microplastic particles (20–1000 µm) using the dye Nile red, fluorescence microscopy and image analysis software. We consider that this method presents a step change in the ability to detect small microplastics by substituting the subjectivity of human visual sorting with a sensitive and semi-automated procedure.
In vitro characterization of the antivirulence target of Gram-positive pathogens, peptidoglycan O-acetyltransferase A (OatA)
The O-acetylation of the essential cell wall polymer peptidoglycan occurs in most Gram-positive bacterial pathogens, including species of Staphylococcus, Streptococcus and Enterococcus. This modification to peptidoglycan protects these pathogens from the lytic action of the lysozymes of innate immunity systems and, as such, is recognized as a virulence factor. The key enzyme involved, peptidoglycan O-acetyltransferase A (OatA) represents a particular challenge to biochemical study since it is a membrane associated protein whose substrate is the insoluble peptidoglycan cell wall polymerThis study on the structure-function relationship of OatA provides a molecular and mechanistic understanding of this bacterial resistance mechanism opening the prospect for novel chemotherapeutic exploration to enhance innate immunity protection against Gram-positive pathogens.