Figure 5.
Identification of the binding sites for the interaction between PHD3 and CYP33 RRM. (A) Plot of chemical shift changes (Δδtot) of the backbone HN and N resonances of PHD3 upon CYP33 RRM titration. Asterisks indicate the residues whose resonances are broadened out. (B) Residues whose backbone resonance were broadened out (yellow) or significantly perturbed (red) in panel A are mapped on the structure of PHD3. The side chain of V1617 is colored green. (C) ITC measurements of the binding of CYP33 RRM (left) to wild-type PHD3 and to the V1617A mutant (right). Wild-type PHD3 (62 μM) was titrated with 755 μM CYP33 RRM, and 98 μM V1617A was titrated with 1 mM CYP33 RRM. (D) Plot of chemical shift changes (Δδtot) of the backbone HN and N resonances of CYP33 RRM upon PHD3 titration. (E) Residues whose backbone resonances were significantly perturbed (red) in panel D are mapped on the structure of CYP33 RRM. The lowest-energy structure from calculation is shown as a ribbon-and-arrow diagram.