Experimental treatment of Staphylococcus aureus bovine intramammary infection using a guanine riboswitch ligand analog

Ster, Céline; Allard, Marie-Julie; Boulanger, Simon; Boulet, Maxime Lamontagne; Mulhbacher, Jérôme; Lafontaine, Daniel A.; Marsault, Éric; Lacasse, P.; Malouin, François

doi:10.3168/jds.2012-5890

Cited by 41 publications

(36 citation statements)

References 24 publications

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“…Based on the crystal structure of hypoxanthine bound to a guanine riboswitch (22), it was proposed that purine and pyrimidine analogs could also form a network of hydrogen bonds for proper complex formation between ligand and aptamer (7). A pyrimidine analog 2,5,6-triaminopyrimidin-4-one (PC1) was reported to kill S. aureus but not bacteria that lack guanine riboswitch control of guaA (7,8). We tested the antibiotic potential of PC1 against S. aureus, Staphylococcus epidermidis, Bacillus subtilis, Klebsiella pneumoniae, Enterococcus faecalis, and Escherichia coli and confirmed that PC1 indeed kills Staphylococcus but not the other Gram-positive and -negative bacteria examined (see Fig.…”

Section: Figmentioning

confidence: 99%

“…The identity and purity of PC1 were verified in-house by liquid chromatography (LC) and mass spectrometry (MS). PC1 stock solutions (2 mg/ml) required 0.03 N NaOH and 1.5 mM dithiothreitol (DTT) to promote solubility and prevent oxidative self-condensation (7,8). All other compounds were made as 100 mM stocks in 0.1 N NaOH.…”

Section: Methodsmentioning

confidence: 99%

“…In Staphylococcus aureus, the riboswitch at the 5=-UTR of the mRNA consisting of the four-gene operon xptpbuX-guaB-guaA (henceforth referred to as xpt-riboswitch) has been reported to regulate the expression of these genes in response to guanine (7,8). Both guaB and guaA, which encode inosine-5=-monophosphate dehydrogenase and GMP synthetase, respectively, are required for de novo biosynthesis of guanine nucleotides.…”

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De Novo Guanine Biosynthesis but Not the Riboswitch-Regulated Purine Salvage Pathway Is Required for Staphylococcus aureus Infection In Vivo

Kofoed¹,

Yan²,

Katakam³

et al. 2016

J Bacteriol

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De novo guanine biosynthesis is an evolutionarily conserved pathway that creates sufficient nucleotides to support DNA replication, transcription, and translation. Bacteria can also salvage nutrients from the environment to supplement the de novo pathway, but the relative importance of either pathway during Staphylococcus aureus infection is not known. In S. aureus, genes important for both de novo and salvage pathways are regulated by a guanine riboswitch. Bacterial riboswitches have attracted attention as a novel class of antibacterial drug targets because they have high affinity for small molecules, are absent in humans, and regulate the expression of multiple genes, including those essential for cell viability. Genetic and biophysical methods confirm the existence of a bona fide guanine riboswitch upstream of an operon encoding xanthine phosphoribosyltransferase (xpt), xanthine permease (pbuX), inosine-5=-monophosphate dehydrogenase (guaB), and GMP synthetase (guaA) that represses the expression of these genes in response to guanine. We found that S. aureus guaB and guaA are also transcribed independently of riboswitch control by alternative promoter elements. Deletion of xpt-pbuX-guaB-guaA genes resulted in guanine auxotrophy, failure to grow in human serum, profound abnormalities in cell morphology, and avirulence in mouse infection models, whereas deletion of the purine salvage genes xpt-pbuX had none of these effects. Disruption of guaB or guaA recapitulates the xpt-pbuXguaB-guaA deletion in vivo. In total, the data demonstrate that targeting the guanine riboswitch alone is insufficient to treat S. aureus infections but that inhibition of guaA or guaB could have therapeutic utility. IMPORTANCEDe novo guanine biosynthesis and purine salvage genes were reported to be regulated by a guanine riboswitch in Staphylococcus aureus. We demonstrate here that this is not true, because alternative promoter elements that uncouple the de novo pathway from riboswitch regulation were identified. We found that in animal models of infection, the purine salvage pathway is insufficient for S. aureus survival in the absence of de novo guanine biosynthesis. These data suggest targeting the de novo guanine biosynthesis pathway may have therapeutic utility in the treatment of S. aureus infections.

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Section: Figmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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confidence: 99%

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De Novo Guanine Biosynthesis but Not the Riboswitch-Regulated Purine Salvage Pathway Is Required for Staphylococcus aureus Infection In Vivo

Kofoed¹,

Yan²,

Katakam³

et al. 2016

J Bacteriol

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“…Thus, they contain a pocket that can be targeted for developing new antibiotics. Indeed, it was shown that compounds binding to the guanine riboswitch in vitro have activity in an in vivo infection model [9][10][11]. Further, it was demonstrated that the antibiotic compounds pyrithiamine and roseoflavin most likely act via the thiamine pyrophosphate (TPP) and flavin mononucleotide (FMN) riboswitches, resp.…”

Section: Introductionmentioning

confidence: 99%

Structure-Based Discovery of Small Molecules Binding to RNA

Wehler

Brenk

2017

Topics in Medicinal Chemistry

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Ribonucleic acids (RNAs) constitute attractive drug targets. The wealth of structural information about RNAs is steadily increasing making it possible to use this information for the design of new ligands. Two methods that make heavy use of structural knowledge for ligand discovery are molecular docking and fragment screening. In molecular docking the structure of the binding site is used as a template for the design of new ligands using computational methods whereas in fragment screening biophysical methods are used for the detection of weak binding ligands which are subsequently elaborated into tighter binding molecules. In this chapter, we give an overview of both methods in the context of ligand discovery for RNA targets and illustrate their applications for hit discovery.

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“…Three of these riboswitch classes have been revealed as important cellular targets of antibacterial small molecules whose mechanism of action had not been previously defined (9)(10)(11)(12)(13). More recently, several publications have demonstrated that novel small molecules that bind to selected riboswitch aptamers with affinities comparable to that of the cognate ligand can be rationally identified and optimized (14)(15)(16)(17)(18)(19)(20)(21).…”

mentioning

confidence: 99%

Novel Riboswitch-Binding Flavin Analog That Protects Mice against Clostridium difficile Infection without Inhibiting Cecal Flora

Blount

Megyola

Plummer

et al. 2015

Antimicrob Agents Chemother

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Despite nearly 85 years of successful antibacterial chemotherapy, bacterial infections still pose a serious threat to human health. Continually emerging drug-resistant bacteria make existing agents less effective, and a paucity of new agents and decreased development effort from the pharmaceutical industry provide little hope of replenishing the arsenal (1, 2). Complications and mortality due to bacterial infections continue to increase, already reaching epidemic proportions in some areas of the world. If humans are to regain the upper hand in fighting bacterial infections, then innovation, investment, and new antibacterial agents will be needed. Although some of the barriers to the discovery and approval of new compounds are economic or policy related, there is also a desperate need for new targets and new mechanisms of action. A renewed effort to expand our knowledge of bacterial physiology and to translate discoveries into the clinic will be needed to address these challenges and to reinvigorate antibiotic development pipelines.Recent advances in our understanding of how bacteria maintain physiological homeostasis revealed a promising class of potential antibiotic targets called riboswitches-noncoding mRNAs that form a structured receptor (or aptamer) which can directly bind to a specific small-molecule ligand or ion and thereby regulate gene expression (3-5). Ligand binding to a riboswitch aptamer stabilizes a conformationally distinct architecture in the mRNA that modulates the expression of the adjacent coding region(s) (4-8). To date, more than 35 riboswitch classes have been discovered and characterized (7). Three of these riboswitch classes have been revealed as important cellular targets of antibacterial small molecules whose mechanism of action had not been previously defined (9-13). More recently, several publications have demonstrated that novel small molecules that bind to selected riboswitch aptamers with affinities comparable to that of the cognate ligand can be rationally identified and optimized (14-21).In some cases, synthetic or natural riboswitch ligand analogs have demonstrated potent antibacterial activity (12-15, 20, 22). For example, the phosphorylated form of roseoflavin (RoF) (

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Experimental treatment of Staphylococcus aureus bovine intramammary infection using a guanine riboswitch ligand analog

Cited by 41 publications

References 24 publications

De Novo Guanine Biosynthesis but Not the Riboswitch-Regulated Purine Salvage Pathway Is Required for Staphylococcus aureus Infection In Vivo

De Novo Guanine Biosynthesis but Not the Riboswitch-Regulated Purine Salvage Pathway Is Required for Staphylococcus aureus Infection In Vivo

Structure-Based Discovery of Small Molecules Binding to RNA

Novel Riboswitch-Binding Flavin Analog That Protects Mice against Clostridium difficile Infection without Inhibiting Cecal Flora

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