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- Title
Fluorescence enzymatic assay for bacterial polyphosphate kinase 1 (PPK1) as a platform for screening antivirulence molecules.
- Authors
Campos, Francisca; Álvarez, Javiera A; Ortiz-Severín, Javiera; Varas, Macarena A; Lagos, Carlos F; Cabrera, Ricardo; Álvarez, Sergio A; Chávez, Francisco P
- Abstract
Inorganic polyphosphate (polyP) and its metabolic enzymes are important in several cellular processes related with virulence and antibiotic susceptibility. Accordingly, bacterial polyP synthesis has been proposed as a good target for designing novel antivirulence molecules as alternative to conventional antibiotics. In most pathogenic bacteria, polyphosphate kinase 1 (PPK1), in charge of polyP synthesis from ATP, is widely conserved. Current colorimetric and radioactive polyP synthesis enzymatic assays are not suitable for high-throughput screening of PPK1 inhibitors. Given the ability of polyP to modify the excitation-emission spectra of DAPI (4ʹ-6-diamidino-2-phenylindole), a fluorescence assay was previously developed by using a purified recombinant PPK1 enzyme from Escherichia coli. In this work we have developed a suitable methodology for high-throughput measurement of E. coli PPK1 activity. This platform can be used for the screening putative antimicrobial molecules for related enteropathogenic bacteria.
- Subjects
FLUORESCENCE; MOLECULES; PEPTIDE antibiotics
- Publication
Infection & Drug Resistance, 2019, Vol 12, p2237
- ISSN
1178-6973
- Publication type
Article
- DOI
10.2147/IDR.S181906