title: Far-red fluorogenic probes for esterase and lipase detection.
abstract: Fluorogenic enzyme probes go from a dark to a bright state following hydrolysis and can provide a sensitive, real-time readout of enzyme activity. They are useful for examining enzymatic activity in bacteria, including the human pathogen Mycobacterium tuberculosis. Herein, we describe two fluorogenic esterase probes derived from the far-red fluorophore 7-hydroxy-9H-(1,3-dichloro-9,9-dimethylacridin-2-one) (DDAO). These probes offer enhanced optical properties compared to existing esterase probes because the hydrolysis product, DDAO, excites above 600 nm while retaining a good quantum yield (ϕ=0.40). We validated both probes with a panel of commercially available enzymes alongside known resorufin- and fluorescein-derived esterase substrates. Furthermore, we used these probes to reveal esterase activity in protein gel-resolved mycobacterial lysates. These probes represent new tools for esterase detection and characterization and should find use in a variety of applications.
authors.affiliation: Department of Physiology and Pharmacology, Department of Biomedical Engineering, Oregon Health & Science University, 2730 SW Moody Avenue, CL3B, Portland, OR 97201 (USA).
authors.forename: Katie R
authors.initials: KR
authors.lastname: Tallman
authors.forename: Kimberly E
authors.initials: KE
authors.lastname: Beatty
mesh_terms: D000166:Acridines / Q000138:chemical synthesis / Q000737:chemistry*; D001426:Bacterial Proteins / Q000032:analysis* / Q000737:chemistry; D001692:Biological Transport; D004950:Esterases / Q000032:analysis* / Q000737:chemistry; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D005656:Fungal Proteins / Q000032:analysis* / Q000737:chemistry; D005658:Fungi / Q000201:enzymology; D006090:Gram-Negative Bacteria / Q000201:enzymology; D006094:Gram-Positive Bacteria / Q000201:enzymology; D006868:Hydrolysis; D057230:Limit of Detection; D008049:Lipase / Q000032:analysis* / Q000737:chemistry; D015394:Molecular Structure; D013050:Spectrometry, Fluorescence; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: C416725:9H-(1,3-dichloro-9,9-dimethylacridine-2-on-7-yl)phosphate; D000166:Acridines; D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D005656:Fungal Proteins; D004950:Esterases; D008049:Lipase
keywords: esterases; fluorescence; fluorogenic probes; lipases; tuberculosis