Figure 5. The S248W GUCY2D activated by GCAP1 binds human RD3 in vitro. Fractional guanylyl cyclase activity was measured in human embryonic kidney cells (HEK293) membranes containing recombinant wild-type (open square) or S248W (black circle) GUCY2D reconstituted with 1.5 µM guanylyl cyclase activating protein 1 (GCAP1) and variable concentrations of recombinant human retinal degeneration 3 (RD3) in the presence of 2 mM EGTA. The EC₅₀ for RD3-dependent inhibition from the fit assuming the Hill function was 1.9 ± 0.3 nM (n = 5) and 1.4 ± 0.1 nM (n = 3), respectively (p = 0.07). Error bars = standard deviation.