Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 8 de 8
Filter
Add more filters










Database
Language
Publication year range
1.
Antimicrob Agents Chemother ; 54(9): 3659-70, 2010 Sep.
Article in English | MEDLINE | ID: mdl-20547796

ABSTRACT

The widespread emergence of antibiotic-resistant bacteria and a lack of new pharmaceutical development have catalyzed a need for new and innovative approaches for antibiotic drug discovery. One bottleneck in antibiotic discovery is the lack of a rapid and comprehensive method to identify compound mode of action (MOA). Since a hallmark of antibiotic action is as an inhibitor of essential cellular targets and processes, we identify a set of 308 essential genes in the clinically important pathogen Staphylococcus aureus. A total of 446 strains differentially expressing these genes were constructed in a comprehensive platform of sensitized and resistant strains. A subset of strains allows either target underexpression or target overexpression by heterologous promoter replacements with a suite of tetracycline-regulatable promoters. A further subset of 236 antisense RNA-expressing clones allows knockdown expression of cognate targets. Knockdown expression confers selective antibiotic hypersensitivity, while target overexpression confers resistance. The antisense strains were configured into a TargetArray in which pools of sensitized strains were challenged in fitness tests. A rapid detection method measures strain responses toward antibiotics. The TargetArray antibiotic fitness test results show mechanistically informative biological fingerprints that allow MOA elucidation.


Subject(s)
Anti-Bacterial Agents/pharmacology , Gene Expression Regulation, Bacterial/genetics , Genes, Essential/genetics , Staphylococcus aureus/genetics , Gene Expression Regulation, Bacterial/drug effects , RNA, Antisense/genetics , Staphylococcus aureus/drug effects
4.
Bioorg Med Chem Lett ; 16(20): 5451-6, 2006 Oct 15.
Article in English | MEDLINE | ID: mdl-16890435

ABSTRACT

Structure-activity relationships of the 3,5-diamino-piperidinyl triazine series, a novel class of bacterial translation inhibitors, are described. Optimization was focused on the triazine C-4 position in which aromatic substituents that contained electron-withdrawing groups led to potent inhibitors. The initial lack of antibacterial activity was correlated with poor cellular penetration. Whole cell antibacterial activity was achieved by linking additional aromatic moieties at the triazine C-4 position.


Subject(s)
Anti-Bacterial Agents/pharmacology , Escherichia coli/drug effects , Piperidines/pharmacology , Protein Biosynthesis/drug effects , Staphylococcus aureus/drug effects , Triazines/pharmacology , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , Microbial Sensitivity Tests , Molecular Structure , Piperidines/chemical synthesis , Piperidines/chemistry , Stereoisomerism , Structure-Activity Relationship , Triazines/chemical synthesis , Triazines/chemistry
5.
J Bacteriol ; 188(3): 1180-3, 2006 Feb.
Article in English | MEDLINE | ID: mdl-16428424

ABSTRACT

An Escherichia coli mutant was isolated and shown to be polymyxin B resistant. Mapping and sequence analysis revealed a missense mutation at codon 53 within the pmrA (basR) gene that results in a G-to-V substitution. Fusions of promoters from the pmrC, yibD, and pmrH genes with the lacZ reporter showed that they were constitutively expressed in pmrA53 cells. In pmrA+ strains, these promoters were induced by iron and zinc, while a DeltapmrA mutation blocked induction. The PmrA regulon regulates genes whose products remodel the composition and charge of lipid A and hence the barrier properties of the outer membrane. Along these lines, the pmrA53 mutant was also found to be hypersensitive to the anionic bile detergent deoxycholic acid.


Subject(s)
Bacterial Proteins/genetics , Deoxycholic Acid/pharmacology , Escherichia coli/drug effects , Polymyxin B/pharmacology , Bacterial Proteins/metabolism , Drug Resistance, Bacterial , Escherichia coli/genetics , Escherichia coli/pathogenicity , Gene Expression Regulation, Bacterial , Lipid A/chemistry , Mutation
6.
Antimicrob Agents Chemother ; 49(12): 4942-9, 2005 Dec.
Article in English | MEDLINE | ID: mdl-16304156

ABSTRACT

We report the structure-guided discovery, synthesis, and initial characterization of 3,5-diamino-piperidinyl triazines (DAPT), a novel translation inhibitor class that targets bacterial rRNA and exhibits broad-spectrum antibacterial activity. DAPT compounds were designed as structural mimetics of aminoglycoside antibiotics which bind to the bacterial ribosomal decoding site and thereby interfere with translational fidelity. We found that DAPT compounds bind to oligonucleotide models of decoding-site RNA, inhibit translation in vitro, and induce translation misincorporation in vivo, in agreement with a mechanism of action at the ribosomal decoding site. The novel DAPT antibacterials inhibit growth of gram-positive and gram-negative bacteria, including the respiratory pathogen Pseudomonas aeruginosa, and display low toxicity to human cell lines. In a mouse protection model, an advanced DAPT compound demonstrated efficacy against an Escherichia coli infection at a 50% protective dose of 2.4 mg/kg of body weight by single-dose intravenous administration.


Subject(s)
Aminoglycosides/pharmacology , Protein Biosynthesis/drug effects , Anti-Bacterial Agents/pharmacology , Biomimetic Materials/chemistry , Biomimetic Materials/pharmacology , Drug Design , Gram-Negative Bacteria/drug effects , Gram-Positive Bacteria/drug effects , Piperidines/pharmacology , Protein Conformation , Ribosomes/drug effects , Structure-Activity Relationship , Triazines/pharmacology
8.
Mol Microbiol ; 43(6): 1387-400, 2002 Mar.
Article in English | MEDLINE | ID: mdl-11952893

ABSTRACT

To address the need for new approaches to antibiotic drug development, we have identified a large number of essential genes for the bacterial pathogen, Staphylococcus aureus, using a rapid shotgun antisense RNA method. Staphylococcus aureus chromosomal DNA fragments were cloned into a xylose-inducible expression plasmid and transformed into S. aureus. Homology comparisons between 658 S. aureus genes identified in this particular antisense screen and the Mycoplasma genitalium genome, which contains 517 genes in total, yielded 168 conserved genes, many of which appear to be essential in M. genitalium and other bacteria. Examples are presented in which expression of an antisense RNA specifically reduces its cognate mRNA. A cell-based, drug-screening assay is also described, wherein expression of an antisense RNA confers specific sensitivity to compounds targeting that gene product. This approach enables facile assay development for high throughput screening for any essential gene, independent of its biochemical function, thereby greatly facilitating the search for new antibiotics.


Subject(s)
Bacterial Proteins/genetics , Gene Targeting , Genes, Essential , Genome, Bacterial , RNA, Antisense , Staphylococcus aureus/genetics , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/metabolism , Cloning, Molecular , Computational Biology/methods , Mycoplasma/genetics , Mycoplasma/metabolism , Plasmids , RNA, Messenger/genetics , Staphylococcus aureus/drug effects , Staphylococcus aureus/growth & development , Transformation, Bacterial , Xylose/pharmacology
SELECTION OF CITATIONS
SEARCH DETAIL
...