<p>Fluorescence enzymatic assay for bacterial polyphosphate kinase 1 (PPK1) as a platform for screening antivirulence molecules</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.

Discovery and antibacterial study of potential PPK1 inhibitors against uropathogenic E. coli

“…Since PPK1 is essential for virulence of bacteria [21][22][23] and CFT073 is a uropathogenic strain, we further investigated the efficacy of 8 and 17 in the treatment and/or prevention of UTIs. To examine the therapeutic potential, 8 and 17 were determined for their ability to reduce bacterial burdens in the urinary tracts of C57BL/6J mice during acute UTIs.…”