20.500.12556/RUL-135754 Evaluation of firefly and Renilla luciferase inhibition in reporter-gene assays a case of isoflavonoids Firefly luciferase is susceptible to inhibition and stabilization by compounds under investigation for biological activity and toxicity. This can lead to false-positive results in in vitro cell-based assays. However, firefly luciferase remains one of the most commonly used reporter genes. Here, we evaluated isoflavonoids for inhibition of firefly luciferase. These natural compounds are often studied using luciferase reporter-gene assays. We used a quantitative structure–activity relationship (QSAR) model to compare the results of in silico predictions with a newly developed in vitro assay that enables concomitant detection of inhibition of firefly and Renilla luciferases. The QSAR model predicted a moderate to high likelihood of firefly luciferase inhibition for all of the 11 isoflavonoids investigated, and the in vitro assays confirmed this for seven of them: daidzein, genistein, glycitein, prunetin, biochanin A, calycosin, and formononetin. In contrast, none of the 11 isoflavonoids inhibited Renilla luciferase. Molecular docking calculations indicated that isoflavonoids interact favorably with the D-luciferin binding pocket of firefly luciferase. These data demonstrate the importance of reporter-enzyme inhibition when studying the effects of such compounds and suggest that this in vitro assay can be used to exclude false-positives due to firefly or Renilla luciferase inhibition, and to thus define the most appropriate reporter gene. isoflavonoids firefly luciferase Renilla luciferase inhibition quantitative structure–activity relationships molecular docking reporter-gene assay OECD test guidelines alternative methods izoflavonoidi zaviralci alternativne metode molekularno mapiranje farmacevtska kemija flavonoidi true false true Angleški jezik Slovenski jezik Članek v reviji 2022-03-30 13:37:15 2022-03-30 13:37:17 2023-03-15 08:51:15 0000-00-00 00:00:00 2021 0 0 14 str. iss. 13, art. 6927 Vol. 22 2021 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 547.972:615.4.:54 1422-0067 10.3390/ijms22136927 72671747 RAZ_Kenda_Masa_2021.pdf RAZ_Kenda_Masa_2021.pdf 1 43760E4B3AB91AE4AF533690EFCB9E4A 30f86e24806b2594e629b2a12f6ea334f1a846d13e2a5cce7a224ef0502fc92f 2a1e1c8f-b01e-11ec-8aca-00155dcfd717 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=155030 https://www.mdpi.com/1422-0067/22/13/6927 1 bb988917-b01d-11ec-8aca-00155dcfd717 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=155029 Fakulteta za farmacijo 0 0 0