Protein phosphatase inhibitory activity of tautomycin photoaffinity probes evaluated at femto-molar level

• Published in: Bioorganic & medicinal chemistry Vol. 16; no. 4; pp. 1747 - 1755
• Main Authors: Sydnes, Magne O., Kuse, Masaki, Kurono, Masakuni, Shimomura, Aya, Ohinata, Hiroshi, Takai, Akira, Isobe, Minoru
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 15.02.2008; Elsevier Science
• Subjects: Bacteriology; Biological and medical sciences; Enzyme Inhibitors; Firefly bioluminescence assay; Firefly Luciferin; Fundamental and applied biological sciences. Psychology; General aspects; Luciferine phosphate; Luminescent Measurements - methods; Luminescent Measurements - standards; Microbiology; Molecular Probes; Phosphoprotein Phosphatases - antagonists & inhibitors; Photoaffinity probe; Photochemistry; Protein phosphatase 1; Pyrans - pharmacology; Spiro Compounds - pharmacology; Tautomycin; Tautomycin; Luciferine phosphate; Protein phosphatase 1; Photoaffinity probe; Firefly bioluminescence assay; Bioluminescence; Enzyme; Phosphoprotein phosphatase; Phosphoric monoester hydrolases; Esterases; Photoaffinity labelling; Biological activity; Spiran; Acetal; Signal transduction; Hydrolases; Dicarboxylic acid; Secondary alcohol
• ISSN: 0968-0896; 1464-3391
• DOI: 10.1016/j.bmc.2007.11.034

Development of a highly sensitive firefly bioluminescence assay system was, in fact, applied to ten tautomycin analogs having photoaffinity probes to obtain the K i values in the range of 3.4–213 nM, and one of them showed slightly higher activity than the natural tautomycin diacid derivative. Herein we describe the further improvement of our in-house developed firefly bioluminescence assay system for the determination of inhibition of protein phosphatase (PP). The advantage with the new system is higher sensitivity as well as being time and sample efficient. The inhibition activity of tautomycin with PP1γ was determined using the upgraded test system and K i was found to be 4.5 nM, which compare favorably with the activity reported previously by others using different methods. The test system was then used in order to determine the activity of nine tautomycin (TTM) photoaffinity probes. One of the TTM photoaffinity probes ( anti- 10) was found to possess higher activity than the natural product itself with a K i of 3.4 nM, while the remaining photoaffinity probes were found to possess K i in the range of 8.0–213 nM.