Characterisation of utrophin modulator SMT C1100 as a non-competitive inhibitor of firefly luciferase.

Isabel V L Wilkinson, Jessica K Reynolds, Sébastien R G Galan, Aini Vuorinen, Adam J Sills, Elisabete Pires, Graham M Wynne, Francis X Wilson, Angela J Russell

Bioorganic chemistry 2020 Jan

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Firefly luciferase (FLuc) is a powerful tool for molecular and cellular biology, and popular in high-throughput screening and drug discovery. However, FLuc assays have been plagued with positive and negative artefacts due to stabilisation and inhibition by small molecules from a range of chemical classes. Here we disclose Phase II clinical compound SMT C1100 for the treatment of Duchenne muscular dystrophy as an FLuc inhibitor (KD of 0.40 ± 0.15 µM). Enzyme kinetic studies using SMT C1100 and other non-competitive inhibitors including resveratrol and NFκBAI4 identified previously undescribed modes of inhibition with respect to FLuc's luciferyl adenylate intermediate. Employing a photoaffinity strategy to identify SMT C1100's binding site, a photolabelled SMT C1100 probe instead underwent FLuc-dependent photooxidation. Our findings support novel binding sites on FLuc for non-competitive inhibitors. Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.

Citation

Isabel V L Wilkinson, Jessica K Reynolds, Sébastien R G Galan, Aini Vuorinen, Adam J Sills, Elisabete Pires, Graham M Wynne, Francis X Wilson, Angela J Russell. Characterisation of utrophin modulator SMT C1100 as a non-competitive inhibitor of firefly luciferase. Bioorganic chemistry. 2020 Jan;94:103395