scholarly journals Discovery of nicoyamycin A, an inhibitor of uropathogenic Escherichia coli growth in low iron environments

2017 ◽  
Vol 53 (95) ◽  
pp. 12778-12781 ◽  
Author(s):  
Laura A. Mike ◽  
Ashootosh Tripathi ◽  
Connor M. Blankenship ◽  
Alyssa Saluk ◽  
Pamela J. Schultz ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Natural Product ◽  
High Throughput ◽  
High Throughput Screening ◽  
Uropathogenic Escherichia Coli ◽  
Coli Growth

High-throughput screening and activity-guided purification resulted in the identification of a novel natural product that inhibits uropathogenic Escherichia coli growth.

scholarly journals A High-Throughput Approach To Identify Compounds That Impair Envelope Integrity in Escherichia coli

2016 ◽  
Vol 60 (10) ◽  
pp. 5995-6002 ◽  
Author(s):  
Kristin R. Baker ◽  
Bimal Jana ◽  
Henrik Franzyk ◽  
Luca Guardabassi
Keyword(s):  
Escherichia Coli ◽  
High Throughput ◽  
High Throughput Screening ◽  
Gram Negative Bacteria ◽  
Chromogenic Substrate ◽  
Intrinsic Resistance ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Screening Platform

ABSTRACTThe envelope of Gram-negative bacteria constitutes an impenetrable barrier to numerous classes of antimicrobials. This intrinsic resistance, coupled with acquired multidrug resistance, has drastically limited the treatment options against Gram-negative pathogens. The aim of the present study was to develop and validate an assay for identifying compounds that increase envelope permeability, thereby conferring antimicrobial susceptibility by weakening of the cell envelope barrier in Gram-negative bacteria. A high-throughput whole-cell screening platform was developed to measureEscherichia colienvelope permeability to a β-galactosidase chromogenic substrate. The signal produced by cytoplasmic β-galactosidase-dependent cleavage of the chromogenic substrate was used to determine the degree of envelope permeabilization. The assay was optimized by using known envelope-permeabilizing compounds andE. coligene deletion mutants with impaired envelope integrity. As a proof of concept, a compound library comprising 36 peptides and 45 peptidomimetics was screened, leading to identification of two peptides that substantially increased envelope permeability. Compound 79 reduced significantly (from 8- to 125-fold) the MICs of erythromycin, fusidic acid, novobiocin and rifampin and displayed synergy (fractional inhibitory concentration index, <0.2) with these antibiotics by checkerboard assays in two genetically distinctE. colistrains, including the high-risk multidrug-resistant, CTX-M-15-producing sequence type 131 clone. Notably, in the presence of 0.25 μM of this peptide, both strains were susceptible to rifampin according to the resistance breakpoints (R> 0.5 μg/ml) for Gram-positive bacterial pathogens. The high-throughput screening platform developed in this study can be applied to accelerate the discovery of antimicrobial helper drug candidates and targets that enhance the delivery of existing antibiotics by impairing envelope integrity in Gram-negative bacteria.

The Search for Novel Human Pancreatic α-Amylase Inhibitors: High-Throughput Screening of Terrestrial and Marine Natural Product Extracts

ChemBioChem ◽  
2008 ◽  
Vol 9 (3) ◽  
pp. 433-438 ◽  
Author(s):  
Chris A. Tarling ◽  
Kate Woods ◽  
Ran Zhang ◽  
Harry C. Brastianos ◽  
Gary D. Brayer ◽  
...  
Keyword(s):  
Natural Product ◽  
High Throughput ◽  
High Throughput Screening ◽  
Marine Natural Product ◽  
Amylase Inhibitors

scholarly journals Development of a Robust Cytopathic Effect-Based High-Throughput Screening Assay To Identify Novel Inhibitors of Dengue Virus

2012 ◽  
Vol 56 (6) ◽  
pp. 3399-3401 ◽  
Author(s):  
Kevin D. McCormick ◽  
Shufeng Liu ◽  
Jana L. Jacobs ◽  
Ernesto T. A. Marques ◽  
Nicolas Sluis-Cremer ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Natural Product ◽  
High Throughput ◽  
High Throughput Screening ◽  
Cytopathic Effect ◽  
Complex Molecule ◽  
Denv Infection ◽  
Screening Assay ◽  
High Throughput Screening Assay ◽  
Natural Product Library

ABSTRACTWe have developed a robust cytopathic effect-based high-throughput screening assay to identify inhibitors of dengue virus (DENV) infection. Screening of a small natural product library yielded 11 hits. Four of these were found to be potent inhibitors of DENV, although serotype differences were noted. Taken together, these data suggest that screening of larger and more complex molecule libraries may result in the identification of more potent and specific DENV inhibitors.

scholarly journals Development of a Simple Assay Protocol for High-Throughput Screening of Mycobacterium tuberculosis Glutamine Synthetase for the Identification of Novel Inhibitors

2005 ◽  
Vol 10 (7) ◽  
pp. 725-729 ◽  
Author(s):  
Upasana Singh ◽  
Vinita Panchanadikar ◽  
Dhiman Sarkar
Keyword(s):  
Escherichia Coli ◽  
Mycobacterium Tuberculosis ◽  
Glutamine Synthetase ◽  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Coli Strain ◽  
Compound Library ◽  
Essential Enzyme ◽  
Assay Protocol

Mycobacterium tuberculosis glutamine synthetase (GS) is an essential enzyme involved in the pathogenicity of the organism. The screening of a compound library using a robust high-throughput screening (HTS) assay is currently thought to be the most efficient way of getting lead molecules, which are potent inhibitors for this enzyme. The authors have purified the enzyme to a >90% level from the recombinant Escherichia coli strain YMC21E, and it was used for partial characterization as well as standardization experiments. The results indicated that the Kmof the enzyme for L-glutamine and hydroxylamine were 60 mM and 8.3 mM, respectively. The Km for ADP, arsenate, and Mn2+ were 2 [.proportional]M, 5 [.proportional]M, and 25 [.proportional]M, respectively. When the components were adjusted according to their Km values, the activity remained constant for at least 3 h at both 25° C and 37° C. The Z′ factor determined in microplate format indicated robustness of the assay. When the signal/noise ratios were determined for different assay volumes, it was observed that the 200-[.proportional]l volume was found to be optimum. The DMSO tolerance of the enzyme was checked up to 10%, with minimal inhibition. The IC50 value determined for L-methionine S-sulfoximine on the enzyme activity was 3 mM. Approximately 18,000 small molecules could be screened per day using this protocol by a Beckman Coulter HTS setup.

Logistic Considerations to Deliver Natural Product Libraries for High-­Throughput Screening

2015 ◽  
pp. 91-104
Author(s):  
Ngoc Pham ◽  
Stephen Toms ◽  
David Camp ◽  
Ronald Quinn
Keyword(s):  
Natural Product ◽  
High Throughput ◽  
High Throughput Screening

scholarly journals Fluorescence Detection-Based Functional Assay for High-Throughput Screening for MraY

2004 ◽  
Vol 48 (3) ◽  
pp. 897-902 ◽  
Author(s):  
Thérèse Stachyra ◽  
Christophe Dini ◽  
Paul Ferrari ◽  
Ahmed Bouhss ◽  
Jean van Heijenoort ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Pressure ◽  
Cell Wall ◽  
High Throughput ◽  
High Throughput Screening ◽  
Reaction Medium ◽  
The Other ◽  
Functional Assay ◽  
Liquid Chromatography Separation ◽  
Fluorescent Derivative

ABSTRACT We have developed a novel assay specific to MraY, which catalyzes the first membrane step in the biosynthesis of bacterial cell wall peptidoglycan. This was accomplished by using UDP-MurNAc-Nε -dansylpentapeptide, a fluorescent derivative of the MraY nucleotide substrate, and a partially purified preparation of MraY solubilized from membranes of an Escherichia coli overproducing strain. Two versions of the assay were developed, one consisting of the high-pressure liquid chromatography separation of the substrate and product (dansylated lipid I) and the other, without separation and adapted to the high-throughput format, taking advantage of the different fluorescence properties of the nucleotide and lipid I in the reaction medium. The latter assay was validated with a set of natural and synthetic MraY inhibitors.

scholarly journals High-Throughput Screening Platform for Natural Product–Based Drug Discovery Against 3 Neglected Tropical Diseases

2014 ◽  
Vol 20 (1) ◽  
pp. 82-91 ◽  
Author(s):  
F. Annang ◽  
G. Pérez-Moreno ◽  
R. García-Hernández ◽  
C. Cordon-Obras ◽  
J. Martín ◽  
...  
Keyword(s):  
Natural Products ◽  
Drug Discovery ◽  
Natural Product ◽  
High Throughput ◽  
High Throughput Screening ◽  
Neglected Tropical Diseases ◽  
Chemical Diversity ◽  
Tropical Diseases ◽  
Isolation And Characterization ◽  
Microbial Extracts

African trypanosomiasis, leishmaniasis, and Chagas disease are 3 neglected tropical diseases for which current therapeutic interventions are inadequate or toxic. There is an urgent need to find new lead compounds against these diseases. Most drug discovery strategies rely on high-throughput screening (HTS) of synthetic chemical libraries using phenotypic and target-based approaches. Combinatorial chemistry libraries contain hundreds of thousands of compounds; however, they lack the structural diversity required to find entirely novel chemotypes. Natural products, in contrast, are a highly underexplored pool of unique chemical diversity that can serve as excellent templates for the synthesis of novel, biologically active molecules. We report here a validated HTS platform for the screening of microbial extracts against the 3 diseases. We have used this platform in a pilot project to screen a subset (5976) of microbial extracts from the MEDINA Natural Products library. Tandem liquid chromatography–mass spectrometry showed that 48 extracts contain potentially new compounds that are currently undergoing de-replication for future isolation and characterization. Known active components included actinomycin D, bafilomycin B1, chromomycin A3, echinomycin, hygrolidin, and nonactins, among others. The report here is, to our knowledge, the first HTS of microbial natural product extracts against the above-mentioned kinetoplastid parasites.

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