scholarly journals Discovery and Characterization of QPT-1, the Progenitor of a New Class of Bacterial Topoisomerase Inhibitors

2008 ◽  
Vol 52 (8) ◽  
pp. 2806-2812 ◽  
Author(s):  
Alita A. Miller ◽  
Gordon L. Bundy ◽  
John E. Mott ◽  
Jill E. Skepner ◽  
Timothy P. Boyle ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Resistant Bacteria ◽  
Infection Model ◽  
Antibacterial Drug ◽  
Synthetic Compound ◽  
Antibiotic Resistant ◽  
New Class ◽  
Mechanism Of Inhibition ◽  
Barbituric Acid Derivative

ABSTRACT QPT-1 was discovered in a compound library by high-throughput screening and triage for substances with whole-cell antibacterial activity. This totally synthetic compound is an unusual barbituric acid derivative whose activity resides in the (−)-enantiomer. QPT-1 had activity against a broad spectrum of pathogenic, antibiotic-resistant bacteria, was nontoxic to eukaryotic cells, and showed oral efficacy in a murine infection model, all before any medicinal chemistry optimization. Biochemical and genetic characterization showed that the QPT-1 targets the β subunit of bacterial type II topoisomerases via a mechanism of inhibition distinct from the mechanisms of fluoroquinolones and novobiocin. Given these attributes, this compound represents a promising new class of antibacterial agents. The success of this reverse genomics effort demonstrates the utility of exploring strategies that are alternatives to target-based screens in antibacterial drug discovery.

scholarly journals An improved and highly selective fluorescence assay for measuring phosphatidylserine decarboxylase activity

2020 ◽  
Vol 295 (27) ◽  
pp. 9211-9222
Author(s):  
Jae-Yeon Choi ◽  
Raymond Black ◽  
HeeJung Lee ◽  
James Di Giovanni ◽  
Robert C. Murphy ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Mammalian Cells ◽  
Large Scale ◽  
Pathogenic Fungi ◽  
Fluorescence Yield ◽  
Resistant Bacteria ◽  
Fluorescence Signal ◽  
Decarboxylase Activity ◽  
Antibiotic Resistant ◽  
Anti Cancer

Phosphatidylserine decarboxylases (PSDs) catalyze the conversion of phosphatidylserine (PS) to phosphatidylethanolamine (PE), a critical step in membrane biogenesis and a potential target for development of antimicrobial and anti-cancer drugs. PSD activity has typically been quantified using radioactive substrates and products. Recently, we described a fluorescence-based assay that measures the PSD reaction using distyrylbenzene-bis-aldehyde (DSB-3), whose reaction with PE produces a fluorescence signal. However, DSB-3 is not widely available and also reacts with PSD's substrate, PS, producing an adduct with lower fluorescence yield than that of PE. Here, we report a new fluorescence-based assay that is specific for PSD and in which the presence of PS causes only negligible background. This new assay uses 1,2-diacetyl benzene/β-mercaptoethanol, which forms a fluorescent iso-indole-mercaptide conjugate with PE. PE detection with this method is very sensitive and comparable with detection by radiochemical methods. Model reactions examining adduct formation with ethanolamine produced stable products of exact masses (m/z) of 342.119 and 264.105. The assay is robust, with a signal/background ratio of 24, and can readily detect formation of 100 pmol of PE produced from Escherichia coli membranes, Candida albicans mitochondria, or HeLa cell mitochondria. PSD activity can easily be quantified by sequential reagent additions in 96- or 384-well plates, making it readily adaptable to high-throughput screening for PSD inhibitors. This new assay now enables straightforward large-scale screening for PSD inhibitors against pathogenic fungi, antibiotic-resistant bacteria, and neoplastic mammalian cells.

scholarly journals Bioprospecting Staphylococcus Phages with Therapeutic and Bio-Control Potential

Viruses ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 133 ◽  
Author(s):  
Joseph M. Ochieng’ Oduor ◽  
Ermir Kadija ◽  
Atunga Nyachieo ◽  
Marianne W. Mureithi ◽  
Mikael Skurnik
Keyword(s):  
Phage Particle ◽  
Burst Size ◽  
Radiation Energy ◽  
Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Rna Ligase ◽  
The Public ◽  
Ph Range ◽  
Host Metabolism

Emergence of antibiotic-resistant bacteria is a serious threat to the public health. This is also true for Staphylococcus aureus and other staphylococci. Staphylococcus phages Stab20, Stab21, Stab22, and Stab23, were isolated in Albania. Based on genomic and phylogenetic analysis, they were classified to genus Kayvirus of the subfamily Twortvirinae. In this work, we describe the in-depth characterization of the phages that electron microscopy confirmed to be myoviruses. These phages showed tolerance to pH range of 5.4 to 9.4, to maximum UV radiation energy of 25 µJ/cm2, to temperatures up to 45 °C, and to ethanol concentrations up to 25%, and complete resistance to chloroform. The adsorption rate constants of the phages ranged between 1.0 × 10−9 mL/min and 4.7 × 10−9 mL/min, and the burst size was from 42 to 130 plaque-forming units. The phages Stab20, 21, 22, and 23, originally isolated using Staphylococcus xylosus as a host, demonstrated varied host ranges among different Staphylococcus strains suggesting that they could be included in cocktail formulations for therapeutic or bio-control purpose. Phage particle proteomes, consisting on average of ca 60–70 gene products, revealed, in addition to straight-forward structural proteins, also the presence of enzymes such DNA polymerase, helicases, recombinases, exonucleases, and RNA ligase polymer. They are likely to be injected into the bacteria along with the genomic DNA to take over the host metabolism as soon as possible after infection.

scholarly journals Characterization of ESBL-Producing Enterobacteria from Fruit Bats in an Unprotected Area of Makokou, Gabon

2020 ◽  
Vol 8 (1) ◽  
pp. 138 ◽  
Author(s):  
Pierre Philippe Mbehang Nguema ◽  
Richard Onanga ◽  
Guy Roger Ndong Atome ◽  
Jean Constant Obague Mbeang ◽  
Arsène Mabika Mabika ◽  
...  
Keyword(s):  
Resistant Bacteria ◽  
Fruit Bats ◽  
Antibiotic Resistant ◽  
Disk Diffusion Test ◽  
Beta Lactamases ◽  
E Coli ◽  
Terrestrial Mammals ◽  
Extended Spectrum Beta Lactamases ◽  
First Time

In Gabon, terrestrial mammals of protected areas have been identified as a possible source of antibiotic-resistant bacteria. Some studies on antibiotic resistance in bats have already been carried out. The main goal of our study was to detect extended-spectrum beta-lactamases (ESBLs) that are produced by enterobacteria from bats in the Makokou region in Gabon. Sixty-eight fecal samples were obtained from 68 bats caught in the forests located 1 km from the little town of Makokou. After culture and isolation, 66 Gram-negative bacterial colonies were obtained. The double-disk diffusion test confirmed the presence of ESBLs in six (20.69%) Escherichia coli isolates, four (13.79%) Klebsiella pneumoniae isolates, and one (3.45%) Enterobacter cloacae isolate. The analysis based on the nucleotide sequences of the ESBL resistance genes showed that all cefotaximase-Munichs (CTX-Ms) were CTX-M-15 and that all sulfhydryl variables (SHVs) were SHV-11: 41.67% CTX-M-15-producing E. coli, 16.67% CTX-M-15+SHV-11-producing E. coli, 8.33% CTX-M-15-producing K. pneumoniae, 25% CTX-M-15+SHV-11-producing K. pneumoniae, and 8.33% CTX-M-15-produced E. cloacae. This study shows for the first time the presence of multiresistant ESBL-producing enterobacteria in fruit bats in Makokou.

Isolation and characterization of antibiotic-resistant bacteria from pharmaceutical industrial wastewaters

2015 ◽  
Vol 89 ◽  
pp. 54-61 ◽  
Author(s):  
Leyla Tahrani ◽  
Leila Soufi ◽  
Ines Mehri ◽  
Afef Najjari ◽  
Abdenaceur Hassan ◽  
...  
Keyword(s):  
Resistant Bacteria ◽  
Antibiotic Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Industrial Wastewaters ◽  
Isolation And Characterization

scholarly journals Single Ribosomal Protein Mutations in Antibiotic-Resistant Bacteria Analyzed by Mass Spectrometry

2001 ◽  
Vol 45 (11) ◽  
pp. 3046-3055 ◽  
Author(s):  
Sheri K. Wilcox ◽  
Gregory S. Cavey ◽  
James D. Pearson
Keyword(s):  
Mass Spectrometry ◽  
Antibiotic Resistance ◽  
Ribosomal Proteins ◽  
Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Mutant Proteins ◽  
E Coli ◽  
Maldi Tof ◽  
Molecular Masses

ABSTRACT Mutations in several ribosomal proteins are known to be related to antibiotic resistance. For several strains of Escherichia coli, the mutated protein is known but the amino acid actually altered has not been documented. Characterization of these determinants for antibiotic resistance in proteins will further the understanding of the precise mechanism of the antibiotic action as well as provide markers for resistance. Mass spectrometry can be used as a valuable tool to rapidly locate and characterize mutant proteins by using a small amount of material. We have used electrospray and matrix-assisted laser desorption ionization–time of flight (MALDI–TOF) mass spectrometry to map out all 56 ribosomal proteins in E. coli based on intact molecular masses. We used this fingerprinting approach to locate variants of ribosomal proteins displaying a change in mass. In particular we have studied proteins responsible for streptomycin, erythromycin, and spectinomycin resistance in three strains of E. coli, and then we characterized each mutation responsible for resistance by analyzing tryptic peptides of these proteins by using MALDI-TOF and nanoelectrospray tandem mass spectrometry. The results provided markers for antibiotic resistance and demonstrated that mass spectrometry can be used to rapidly investigate changes in individual proteins from a complex with picomole amounts of protein.

Sign in / Sign up

Export Citation Format

Share Document