Peptides have emerged as promising agents to target challenging protein-protein interactions (PPIs). One such difficult interface is the Skp1-Skp2 interaction, which has proven resistant to effective inhibition by small molecules. Here, we rationally designed stapled peptide candidates that effectively disrupt the Skp1-Skp2 interface. Two distinct stapling strategies were employed to enhance peptide stability and potency. The peptides’ efficacy was validated using surface plasmon resonance (SPR) and cancer cell line assays, demonstrating their mechanistic impact. To deepen our understanding and identify more potent binders, we examined peptide interactions with Skp1, revealing a stapling-dependent effect on binding affinity. This was further investigated through combined SPR and nuclear magnetic resonance (NMR) spectroscopy analyses. Correlations between NMR relaxation parameters, chemical shift perturbations, and binding affinity allowed us to distinguish enthalpic and entropic contributions influenced by stapling. Leveraging these insights into dynamic changes and their impact on peptide activity, we aim to develop highly effective peptide inhibitors against Skp1-Skp2 and extend this approach to other challenging PPI interfaces.
Tolani et al. (Sun,) studied this question.