title: Design of small molecule inhibitors of type III secretion system ATPase EscN from enteropathogenic Escherichia coli.
abstract: Enteropathogenic E. coli (EPEC) is a human pathogen using type III secretion system for delivery of proteins directly into the human host. The system contains a single ATPase, EscN, which is essential for uncoupling of proteins from their complexes with chaperones before the delivery. The structure of EscN ATPase (PDB code: 2obm) was used to screen computationally for small molecule inhibitors blocking its active site. Two lead candidates were examined but only one, Compound 54, was selected for further optimization. After extended QSAR optimization, two derivatives were found to be competitive inhibitors of EscN capable of blocking ATPase activity with a Kibelow 50 µM. One candidate, WEN05-03, with a Ki=16±2 µM, was also minimally toxic to mammalian cells as determined by other assays. In the cell infection model of HeLa cells with EPEC, Compound WEN05-03 completely blocked actin cluster formation at 100 µM concentration, when analyzed by confocal microscopy. The second best inhibitor of EscN ATPase activity was WEN04-34 with a Ki=46±2 µM. However, the compound was highly toxic to the BALB/3T3 cell line. In summary, the work identifies a compound blocking bacterial ATPase in its active site without causing cellular toxicity to the host cells. It is the first report showing feasibility of using bacterial virulence system ATPase as a target for safe, non-toxic compounds and offering a proof-of-concept for non-antibiotic alternatives.
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.
authors.forename: Lukasz
authors.initials: L
authors.lastname: Bzdzion
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.
authors.forename: Hanna
authors.initials: H
authors.lastname: Krezel
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.
authors.forename: Karol
authors.initials: K
authors.lastname: Wrzeszcz
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.
authors.forename: Irmina
authors.initials: I
authors.lastname: Grzegorek
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Nowinska
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Chodaczek
authors.affiliation: Department of Biotechnology, Wroclaw Research Centre EIT+ Ltd., Wrocław, Poland.|Institute of Immunology and Experimental Therapy of the Polish Academy of Sciences, Wrocław, Poland.
authors.forename: Wieslaw
authors.initials: W
authors.lastname: Swietnicki
mesh_terms: D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000494:pharmacology; D041702:BALB 3T3 Cells; D020134:Catalytic Domain; D015195:Drug Design; D054308:Enteropathogenic Escherichia coli / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D029968:Escherichia coli Proteins / Q000037:antagonists & inhibitors*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D021281:Quantitative Structure-Activity Relationship; D000069296:Type III Secretion Systems / Q000737:chemistry*
chemical_list: D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D029968:Escherichia coli Proteins; D000069296:Type III Secretion Systems; D000251:Adenosine Triphosphatases; C000593614:EscN protein, E coli
keywords: Escherichia coli (E. coli); enzyme inhibitor; small molecule; type III secretion system (T3SS)