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. Author manuscript; available in PMC: 2019 May 12.
Published in final edited form as: Nat Chem Biol. 2018 Nov 12;14(12):1099–1108. doi: 10.1038/s41589-018-0155-8

Figure 1.

Figure 1.

Development of a high-throughput assay (HTS) screen for ABHD12 inhibitors. (a) An enzyme-coupled assay that begins with ABHD12-mediated hydrolysis of 17:0 lyso-PA, followed by glycerol-3-phosphate oxidase (GPO)-mediated generation of H2O2, and culminating in the horseradish peroxidase (HRP)-mediated production of the fluorescent compound resorufin. (b) Kinetic performance of the enzyme-coupled HTS assay with membrane lysates from ABHD12- or mock-transfected HEK293T cells. The Z’ and S/B values at 45 min were 0.87 and 3.0, respectively. THL (10 μM) was used as a control ABHD12 inhibitor. (c) Screening data for the Maybridge HitFinder™ collection of 16,0000 compounds. Compounds showing> 50% inhibition are marked in red (198 total). The screen was performed once. (d) Structure of the hit compound DO130. (e) IC50 value for inhibition of lyso-PS hydrolysis activity of ABHD12 by DO130 measured using ABHD12-transfected cell lysates with 17:1 lyso-PS substrate (100 μM, 20 min, 37 °C). Data represent average values ± SD (n = 3 independent experiments).