title: A new class of rhomboid protease inhibitors discovered by activity-based fluorescence polarization.
abstract: Rhomboids are intramembrane serine proteases that play diverse biological roles, including some that are of potential therapeutical relevance. Up to date, rhomboid inhibitor assays are based on protein substrate cleavage. Although rhomboids have an overlapping substrate specificity, substrates cannot be used universally. To overcome the need for substrates, we developed a screening assay using fluorescence polarization activity-based protein profiling (FluoPol ABPP) that is compatible with membrane proteases. With FluoPol ABPP, we identified new inhibitors for the E. coli rhomboid GlpG. Among these was a structural class that has not yet been reported as rhomboid inhibitors: β-lactones. They form covalent and irreversible complexes with the active site serine of GlpG. The presence of alkyne handles on the β-lactones also allowed activity-based labeling. Overall, these molecules represent a new scaffold for future inhibitor and activity-based probe development, whereas the assay will allow inhibitor screening of ill-characterized membrane proteases.
authors.affiliation: Lehrstuhl für Chemie der Biopolymere, Technische Universität München, Freising, Germany.
authors.forename: Eliane V
authors.initials: EV
authors.lastname: Wolf
authors.forename: Annett
authors.initials: A
authors.lastname: Zeißler
authors.forename: Oliver
authors.initials: O
authors.lastname: Vosyka
authors.forename: Evelyn
authors.initials: E
authors.lastname: Zeiler
authors.forename: Stephan
authors.initials: S
authors.lastname: Sieber
authors.forename: Steven H L
authors.initials: SH
authors.lastname: Verhelst
mesh_terms: D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D055808:Drug Discovery; D010450:Endopeptidases / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D005454:Fluorescence Polarization / Q000379:methods*; D020128:Inhibitory Concentration 50; D008565:Membrane Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015335:Molecular Probes; D011480:Protease Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D013379:Substrate Specificity
chemical_list: D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C102898:GlpG protein, E coli; D008565:Membrane Proteins; D015335:Molecular Probes; D011480:Protease Inhibitors; D010450:Endopeptidases