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. Author manuscript; available in PMC: 2018 Dec 1.
Published in final edited form as: DNA Repair (Amst). 2017 Oct 10;60:29–39. doi: 10.1016/j.dnarep.2017.10.002

Figure 4. L82-G17 is a selective uncompetitive inhibitor of LigI.

Figure 4

(A) LigI (500 fmol), LigIII (500 fmol) and LigIV (2 pmol) were pre-incubated with 200 µM of either L82 (gray) or L82-G17 (black) for 30 min at 25°C prior to incubation with the fluorescent nicked DNA substrate (1 pmol) for 5 min at 25°C as described in Materials and Methods. The results of three independent experiments are shown graphically. The effect of L82 (B) and L82-G17 (C) on Michaelis-Menten saturation curves of reaction velocity (pmol/min) versus DNA substrate concentration (µM) derived from at least three independent fluorescence-based ligation assays are shown. Lineweaver-Burk double reciprocal plots of the same data for L82 (D) and L82-G17 (E) calculated using the standard error of mean and showing the extrapolated trendline. Solid line, no inhibitor; long dashes, 20 µM inhibitor; short dashes, 200 µM. Curve fitting was performed with Graphpad PRISM analysis software.