Fig. 5 Cell-free membrane cyclase assay confirms that candidate compounds are specific antagonists of hNPR1. Cell-free membrane cyclase assay confirms.

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Fig. 5 Cell-free membrane cyclase assay confirms that candidate compounds are specific antagonists of hNPR1. Cell-free membrane cyclase assay confirms that candidate compounds are specific antagonists of hNPR1. (A) Schematic depicts our strategy to measure hNPR1 activity with an in vitro assay. A crude membrane fraction was prepared from HEK-hNPR1-cGMP sensor cells. Incubation of hNPR1 membranes with guanosine triphosphate (GTP) and NP results in production of cGMP, and cGMP was measured using an enzyme-linked immunosorbent assay (ELISA) test. (B) Quantifications of cGMP production by hNPR1 membranes, stimulated by hNPPA (blue) and SNP (red). (C) Quantification of inhibition of hNPPA-stimulated (1 nM) hNPR1 activity by JS-5 (blue), JS-8 (green), or JS-11 (purple). Data represent means ±SEM for triplicate (B, hNPPA), duplicate (B, SNP), and duplicate measurements (C). Hans Jürgen Solinski et al., Sci Transl Med 2019;11:eaav5464 Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works