Structure-guided design and cloning of peptide inhibitors targeting CDK9/cyclin T1 protein-protein interaction

CDK9 (cyclin-dependent kinase 9) plays a significant role in numerous pathological conditions, such as HIV-1 infection and cancer. The interaction between CDK9 and cyclin T1 is crucial for maintaining the kinase’s active state. Therefore, targeting this protein-protein interaction offers a promising strategy for inhibiting CDK9. In this study, we aimed to design and characterize a library of mutant peptides based on the binding region of cyclin T1 to CDK9. Using Osprey software, a total of 7,776 mutant peptides were generated. After conducting a comprehensive analysis, three peptides, namely, mp3 (RAADVEGQRKRRE), mp20 (RAATVEGQRKRRE), and mp29 (RAADVEGQDKRRE), were identified as promising inhibitors that possess the ability to bind to CDK9 with high affinity and exhibit low free binding energy. These peptides exhibited favorable safety profiles and displayed promising dynamic behaviors. Notably, our findings revealed that the mp3 and mp29 peptides interacted with a conserved sequence in CDK9 (residues 60–66). In addition, by designing the structure of potential peptides in the plasmid vector pET28a (+), we have been able to pave the way for facilitating the process of their recombinant production in an Escherichia coli expression system in future studies. Predictions indicated good solubility upon overexpression, further supporting their potential for downstream applications. While these results demonstrate the promise of the designed peptides as blockers of CDK9 with high affinity, additional experimental studies are required to validate their biological activity and assess their selectivity. Such investigations will provide valuable insights into their therapeutic potential and pave the way for the future development of peptide-based inhibitors targeting the CDK9-cyclin T1 complex.