ABSTRACT
Arginine-pyrimidine conjugates represent a novel class of compounds that exhibits therapeutic and prophylactic activity in lethal infections by Gram-positive and Gram-negative bacteria without showing antibacterial activity in vitro.
Subject(s)
Anti-Bacterial Agents/pharmacology , Arginine/pharmacology , Cross Infection/drug therapy , Microbial Sensitivity Tests , Pyrimidines/pharmacology , Animals , Anti-Bacterial Agents/therapeutic use , Arginine/chemistry , Bacterial Infections/drug therapy , Drug Interactions , Drug Resistance, Multiple, Bacterial , Gram-Negative Bacteria/drug effects , Methicillin Resistance , Mice , Mice, Inbred ICR , Pyrimidines/chemistry , Streptococcal Infections/drug therapyABSTRACT
Aminoacyl-tRNA synthetases (aa-RS) attracted interest as potential targets for new antibacterial compounds. Most organisms express 20 aa-RSs: one for each amino acid. Aa-RSs are essential proteins in all living organisms. When one aa-RS is inhibited, the corresponding tRNA is not charged and is therefore unavailable for translation. This leads to protein synthesis inhibition, which in turn causes cell growth arrest. Consequently, each compound that inhibits any of the aa-RS could be a potential antibacterial agent. Only one aa-RS inhibitor, the Ile-RS inhibitor mupirocin, is currently marketed as an antibacterial agent. We focused on phenylalanyl (Phe)-tRNA synthetase (Phe-RS), but the described methods are not restricted to Phe-RS and might be adapted to other aa-RS.
Subject(s)
Amino Acyl-tRNA Synthetases/antagonists & inhibitors , Bacteria/drug effects , Enzyme Inhibitors/analysis , Amino Acids/blood , Animals , Anti-Bacterial Agents/analysis , Anti-Bacterial Agents/pharmacology , Bacteria/enzymology , Bacteria/pathogenicity , Cell-Free System , Enzyme Inhibitors/pharmacology , Female , Humans , Mice , Microbial Sensitivity Tests , Phenylalanine-tRNA Ligase/antagonists & inhibitors , Sepsis/drug therapy , Sepsis/microbiologyABSTRACT
Here we show that a new class of antibiotics-acyldepsipeptides-has antibacterial activity against Gram-positive bacteria in vitro and in several rodent models of bacterial infection. The acyldepsipeptides are active against isolates that are resistant to antibiotics in clinical application, implying a new target, which we identify as ClpP, the core unit of a major bacterial protease complex. ClpP is usually tightly regulated and strictly requires a member of the family of Clp-ATPases and often further accessory proteins for proteolytic activation. Binding of acyldepsipeptides to ClpP eliminates these safeguards. The acyldepsipeptide-activated ClpP core is capable of proteolytic degradation in the absence of the regulatory Clp-ATPases. Such uncontrolled proteolysis leads to inhibition of bacterial cell division and eventually cell death.
Subject(s)
Anti-Bacterial Agents/classification , Anti-Bacterial Agents/pharmacology , Bacteria/drug effects , Bacteria/metabolism , Depsipeptides/pharmacology , Endopeptidase Clp/metabolism , Escherichia coli Proteins/metabolism , Animals , Anti-Bacterial Agents/pharmacokinetics , Anti-Bacterial Agents/toxicity , Bacillus subtilis/drug effects , Bacteria/enzymology , Depsipeptides/metabolism , Depsipeptides/pharmacokinetics , Depsipeptides/toxicity , Drug Resistance, Multiple, Bacterial , Escherichia coli/drug effects , Female , Mice , Molecular Structure , Mutation , Pneumococcal Infections/drug therapy , Pneumococcal Infections/microbiology , Protein Binding , Protein Processing, Post-Translational , Rats , Rats, Wistar , Sepsis/drug therapy , Sepsis/microbiologyABSTRACT
Phenylalanyl (Phe)-tRNA synthetase (Phe-RS) is an essential enzyme which catalyzes the transfer of phenylalanine to the Phe-specific transfer RNA (tRNA(Phe)), a key step in protein biosynthesis. Phenyl-thiazolylurea-sulfonamides were identified as a novel class of potent inhibitors of bacterial Phe-RS by high-throughput screening and chemical variation of the screening hit. The compounds inhibit Phe-RS of Escherichia coli, Haemophilus influenzae, Streptococcus pneumoniae, and Staphylococcus aureus, with 50% inhibitory concentrations in the nanomolar range. Enzyme kinetic measurements demonstrated that the compounds bind competitively with respect to the natural substrate Phe. All derivatives are highly selective for the bacterial Phe-RS versus the corresponding mammalian cytoplasmic and human mitochondrial enzymes. Phenyl-thiazolylurea-sulfonamides displayed good in vitro activity against Staphylococcus, Streptococcus, Haemophilus, and Moraxella strains, reaching MICs below 1 micro g/ml. The antibacterial activity was partly antagonized by increasing concentrations of Phe in the culture broth in accordance with the competitive binding mode. Further evidence that inhibition of tRNA(Phe) charging is the antibacterial principle of this compound class was obtained by proteome analysis of Bacillus subtilis. Here, the phenyl-thiazolylurea-sulfonamides induced a protein pattern indicative of the stringent response. In addition, an E. coli strain carrying a relA mutation and defective in stringent response was more susceptible than its isogenic relA(+) parent strain. In vivo efficacy was investigated in a murine S. aureus sepsis model and a S. pneumoniae sepsis model in rats. Treatment with the phenyl-thiazolylurea-sulfonamides reduced the bacterial titer in various organs by up to 3 log units, supporting the potential value of Phe-RS as a target in antibacterial therapy.
Subject(s)
Anti-Bacterial Agents/pharmacology , Bacteria/drug effects , Bacteria/enzymology , Enzyme Inhibitors/pharmacology , Animals , Bacillus subtilis/drug effects , Bacillus subtilis/genetics , CHO Cells , Colony Count, Microbial , Cricetinae , Drug Design , Escherichia coli/drug effects , Escherichia coli/enzymology , Female , Gram-Negative Bacteria/drug effects , Gram-Positive Bacteria/drug effects , Mice , Microbial Sensitivity Tests , Pneumococcal Infections/drug therapy , Pneumococcal Infections/microbiology , Proteome/genetics , Rats , Rats, Wistar , Staphylococcal Infections/drug therapy , Staphylococcal Infections/microbiology , Substrate SpecificityABSTRACT
Several types of furanomycin analogues were synthesized and investigated with respect to their antibacterial activity. Two different synthetic pathways were developed, based on aldol reactions/ring closing metathesis and an ester enolate Claisen rearrangement. Only the natural product and its desmethyl derivative showed antibacterial activity, pointing towards a narrow structure-activity relationship.