RESUMEN
Tetracycline resistance by antibiotic inactivation was first identified in commensal organisms but has since been reported in environmental and pathogenic microbes. Here, we identify and characterize an expanded pool of tet(X)-like genes in environmental and human commensal metagenomes via inactivation by antibiotic selection of metagenomic libraries. These genes formed two distinct clades according to habitat of origin, and resistance phenotypes were similarly correlated. Each gene isolated from the human gut encodes resistance to all tetracyclines tested, including eravacycline and omadacycline. We report a biochemical and structural characterization of one enzyme, Tet(X7). Further, we identify Tet(X7) in a clinical Pseudomonas aeruginosa isolate and demonstrate its contribution to tetracycline resistance. Lastly, we show anhydrotetracycline and semi-synthetic analogues inhibit Tet(X7) to prevent enzymatic tetracycline degradation and increase tetracycline efficacy against strains expressing tet(X7). This work improves our understanding of resistance by tetracycline-inactivation and provides the foundation for an inhibition-based strategy for countering resistance.
Asunto(s)
Antibacterianos/farmacología , Pseudomonas aeruginosa/enzimología , Resistencia a la Tetraciclina/genética , Tetraciclinas/antagonistas & inhibidores , Interacciones Huésped-Patógeno , Humanos , Pruebas de Sensibilidad Microbiana , Pseudomonas aeruginosa/efectos de los fármacos , SimbiosisRESUMEN
The synthesis and biological evaluation of semisynthetic anhydrotetracycline analogues as small molecule inhibitors of tetracycline-inactivating enzymes are reported. Inhibitor potency was found to vary as a function of enzyme (major) and substrate-inhibitor pair (minor), and anhydrotetracycline analogue stability to enzymatic and nonenzymatic degradation in solution contributes to their ability to rescue tetracycline activity in whole cell Escherichia coli expressing tetracycline destructase enzymes. Taken collectively, these results provide the framework for the rational design of next-generation inhibitor libraries en route to a viable and proactive adjuvant approach to combat the enzymatic degradation of tetracycline antibiotics.
Asunto(s)
Antibacterianos/química , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Proteínas de Escherichia coli/antagonistas & inhibidores , Escherichia coli/enzimología , Tetraciclina/metabolismo , Tetraciclinas/química , Tetraciclinas/farmacología , Antibacterianos/síntesis química , Antibacterianos/farmacología , Inhibidores Enzimáticos/síntesis química , Escherichia coli/química , Escherichia coli/efectos de los fármacos , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Tetraciclinas/síntesis químicaRESUMEN
Although tetracyclines are an important class of antibiotics for use in agriculture and the clinic, their efficacy is threatened by increasing resistance. Resistance to tetracyclines can occur through efflux, ribosomal protection, or enzymatic inactivation. Surprisingly, tetracycline enzymatic inactivation has remained largely unexplored, despite providing the distinct advantage of antibiotic clearance. The tetracycline destructases are a recently discovered family of tetracycline-inactivating flavoenzymes from pathogens and soil metagenomes that have a high potential for broad dissemination. Here, we show that tetracycline destructases accommodate tetracycline-class antibiotics in diverse and novel orientations for catalysis, and antibiotic binding drives unprecedented structural dynamics facilitating tetracycline inactivation. We identify a key inhibitor binding mode that locks the flavin adenine dinucleotide cofactor in an inactive state, functionally rescuing tetracycline activity. Our results reveal the potential of a new tetracycline and tetracycline destructase inhibitor combination therapy strategy to overcome resistance by enzymatic inactivation and restore the use of an important class of antibiotics.
Asunto(s)
Antibacterianos/metabolismo , Inhibidores Enzimáticos/farmacología , Legionella longbeachae/efectos de los fármacos , Legionella longbeachae/enzimología , Resistencia a la Tetraciclina/efectos de los fármacos , Tetraciclina/metabolismo , Antibacterianos/química , Antibacterianos/farmacología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/química , Flavina-Adenina Dinucleótido/metabolismo , Legionella longbeachae/metabolismo , Modelos Moleculares , Conformación Molecular , Relación Estructura-Actividad , Tetraciclina/química , Tetraciclina/farmacologíaRESUMEN
Lithium-air (O2) batteries have shown great promise because of their high gravimetric energy density-an order of magnitude greater than Li-ion-but challenges such as electrolyte and electrode instability have led to poor capacity retention and low cycle life. Positive electrodes such as carbon and inorganic metal oxides have been heavily explored, but the degradation of carbon and the limited surface area of the metal oxides limit their practical use. In this work, we study the electron-conducting polymer poly(3,4-ethylenedioxythiophene) (PEDOT) and show it can support oxygen reduction to form Li2O2 in a nonaqueous environment. We also propose a degradation mechanism and show that the formation of sulfone functionalities on the PEDOT surface and cleavage of the polymer repeat unit impairs electron conductivity and leads to poor cycling. Our findings are important in the search for new Li-O2 electrodes, and the physical insights provided are significant and timely.