Sickle cell disease (SCD) is a monogenic disorder caused by the substitution of polar glutamic acid with hydrophobic valine (E6V) in β-globin chain to form sickle hemoglobin (HbS). The new hydrophobic contact created by this mutation facilitates HbS polymerization when deoxygenated. Peptide-based inhibitors have been proposed to block HbS polymerization by targeting hydrophobic pocket interacting with V6. One promising cyclic pentapeptide, c-VEVFV, was proposed to interact with this pocket based on computational studies; however, those studies focused on hydrophobic pocket and did not examine other sites on HbS. We studied this peptide using experimental approaches and docking studies including tetrameric HbS. Isothermal titration calorimetry demonstrated that c-VEVFV binds HbS with high affinity (Kd = 4.16 × 10 −8 M); however, despite strong binding, it failed to inhibit sickling in a red blood cell assay. To elucidate the binding mechanism, we performed computational blind docking to identify potential binding pockets across HbS, followed by targeted docking on most promising cavities. The results revealed that c-VEVFV preferentially binds to a deep pocket in central core of tetrameric HbS with a calculated affinity similar to experimentally measured value. Our computational studies suggested that binding to the hydrophobic pocket is much weaker than previously reported. These findings suggest that c-VEVFV is drawn away from intended hydrophobic pocket and instead binds to the deep central pocket within HbS, where it is ineffective at inhibiting polymerization. In the future, this cyclic peptide, which has high binding affinity for HbS, could potentially be modified to disrupt key contacts involved in polymerization. Our work highlights importance of combining experimental and computational approaches to guide rational design of more effective anti-sickling agents.
Mamadou Sangare (Sun,) studied this question.