title: Substrate profiling of Finegoldia magna SufA protease, inhibitor screening and application to prevent human fibrinogen degradation and bacteria growth in vitro.
abstract: SufA, which belongs to the subtilisin-like serine protease family, contains a non-canonical Asp-His-Ser catalytic triad. Under in vitro conditions, SufA is capable of human fibrinogen hydrolysis leading to inhibition of fibrin network formation, thus suggesting its important role in the development and progression of Finegoldia magna infections. In addition, it has been demonstrated that SufA can hydrolyze antibacterial peptides such as LL-37 and the chemokine MIG/CXCL 9, hence evading host defence mechanisms. Although the SufA protease from F. magna was discovered several years ago, its optimal substrate preference has not yet been identified. Considering the role of SufA, we have focused on the profiling of its substrate sequence preference spanning S1-S3 binding pockets using the FRET (fluorescence resonance energy transfer) approach. Next, based on the structure of the P1 residue of the developed substrate, we narrowed the inhibitor screening to the phosphonic analogues of amino acids containing an arginine-like side chain. Among all the compounds tested, only Cbz-6-AmNphth(P)(OPh)2 showed any inhibitory activity against SufA displaying k2/Ki value of 10,800 M(-1) s(-1). In addition, it prevented SufA-mediated human fibrinogen hydrolysis in vitro and exhibited potent antibacterial activity against F. magna, Staphylococcus aureus and Escherichia coli. Herein, we report on the substrate specificity, synthesis and kinetic evaluation of phosphonic inhibitors of SufA protease from F. magna which could help to establish its function in pathogenesis development and may lead to the elaboration of new antibacterial drugs.
authors.affiliation: Division of Medicinal Chemistry and Microbiology, Faculty of Chemistry, Wroclaw University of Technology, Wybrzeze Wyspianskiego 27, 50-370 Wroclaw, Poland. Electronic address: ewa.burchacka@pwr.wroc.pl.
authors.forename: Ewa
authors.initials: E
authors.lastname: Burchacka
authors.affiliation: Division of Medicinal Chemistry and Microbiology, Faculty of Chemistry, Wroclaw University of Technology, Wybrzeze Wyspianskiego 27, 50-370 Wroclaw, Poland.
authors.forename: Marcin
authors.initials: M
authors.lastname: Sieńczyk
authors.affiliation: Division of Infection Medicine, Department of Clinical Sciences, Lund University, SE-221 84 Lund, Sweden.
authors.forename: Inga-Maria
authors.initials: IM
authors.lastname: Frick
authors.affiliation: Faculty of Chemistry, Gdansk University, Wita Stwosza 63, 80-952 Gdansk, Poland.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Wysocka
authors.affiliation: Faculty of Chemistry, Gdansk University, Wita Stwosza 63, 80-952 Gdansk, Poland.
authors.forename: Adam
authors.initials: A
authors.lastname: Lesner
authors.affiliation: Division of Medicinal Chemistry and Microbiology, Faculty of Chemistry, Wroclaw University of Technology, Wybrzeze Wyspianskiego 27, 50-370 Wroclaw, Poland.
authors.forename: Józef
authors.initials: J
authors.lastname: Oleksyszyn
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D004353:Drug Evaluation, Preclinical; D005340:Fibrinogen / Q000378:metabolism*; D006094:Gram-Positive Bacteria / Q000187:drug effects / Q000201:enzymology* / Q000254:growth & development*; D006801:Humans; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D010447:Peptide Hydrolases / Q000378:metabolism*; D010757:Phosphorous Acids / Q000737:chemistry / Q000494:pharmacology; D011480:Protease Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D059748:Proteolysis / Q000187:drug effects*; D013379:Substrate Specificity
chemical_list: D000900:Anti-Bacterial Agents; D010757:Phosphorous Acids; D011480:Protease Inhibitors; C570063:phosphonic acid; D005340:Fibrinogen; D010447:Peptide Hydrolases
keywords: Finegoldia magna; Phosphonic inhibitors; Substrate mapping; SufA protease