Fig. 3. Bumped ligand ET binds to engineered L/A bromodomains with high selectivity.
(A) Thermal stabilization (ΔTm, in degrees Celsius) of WT Brd2 bromodomains and their L/A mutants by I-BET, ME, and ET measured by means of DSF. The data shown are mean ± SD of three measurements. (B) Dissociation constants (Kd, in nanomolar) determined by means of ITC for I-BET binding to WT and for ME and ET binding to WT and L/A mutant Brd2 bromodomains. The error bars reflect the quality of the fit between the nonlinear least-squares curve and the experimental points. (C and D) Cocrystal structures of Brd2(2)L383A (green, surface representation) in complex with ME [(C) stick, yellow carbons] and ET [(D) stick, pink carbons].The L/A mutation is shown in red, and A383 is shown as stick, green carbons. (E and F) Affinity profile (Kd, in nanomolar) of ET binding to (E) wild-type and (F) L/A mutant BET bromodomains measured by means of ITC.