tailieunhanh - Báo cáo khoa học: Ecdysteroid ligand–receptor selectivity – exploring trends to design orthogonal gene switches

A set of thirty-two natural and ten semisynthetic ecdysteroids was assayed in murine 3T3 cells across ten different ecdysteroid receptor (EcR) ligand-binding domains derived from nine arthropod species in an engineered gene switch format. Among the ecdysteroids tested, the most biologically wide-spread ecdysteroid, 20-hydroxyecdysone (20E), was moderately and consis-tently potent across the tested EcRs. | ỊFEBS Journal Ecdysteroid ligand-receptor selectivity - exploring trends to design orthogonal gene switches Silvia Lapenna1 Jennifer Friz2 1 Anna Barlow1 í Subba R. Palli3 Laurence Dinan1 and Robert E. Hormann2 1 Department of BiologicalSciences University of Exeter UK 2 Intrexon Corp. Norristown PA USA 3 Department of Entomology College of Agriculture University of Kentucky Lexington KY USA Keywords canescensterone EcR gene switch orthogonality ponasterone A Correspondence R. E. Hormann Intrexon Corp. 2650 Eisenhower Ave. Norristown PA 19027 USA Fax 1 610 650 8755 Tel 1 540 808 2623 E-mail rhormann@ Present address CROM - Oncology Research Centre of Mercogliano Fondazione F. Lo Vuolo Mercogliano AV Italy Bioprocess and BioanalyticalResearch Merck Co. Inc. West Point PA USA Cadbury Confectionary Ltd. Dunedin New Zealand Laboratoire de Biochimie Structurale et Fonctionnelle des Proteines CNRS FRE 2852 Universite Pierre et Marie Curie Paris France Received 16 June 2008 revised 12 August 2008 accepted 17 September 2008 doi A set of thirty-two natural and ten semisynthetic ecdysteroids was assayed in murine 3T3 cells across ten different ecdysteroid receptor EcR ligandbinding domains derived from nine arthropod species in an engineered gene switch format. Among the ecdysteroids tested the most biologically widespread ecdysteroid 20-hydroxyecdysone 20E was moderately and consistently potent across the tested EcRs. The most potent ligand-receptor combination EC50 nM was ponasterone A PoA actuating the Nephotettix cincticeps EcR switch. The most robust ligand-receptor combination as measured by potency and efficacy was PoA actuating either the Bombyx mori EcR or a VY E274V V390I Y410E mutant of Choristone-ura fumiferana EcR. Parallel ecdysteroid structure-activity relationships were observed across species addition of hydroxyl groups at positions 2 3 14 20 and 22 incrementally enhanced potency whereas hydroxylation at .