Abstract
Protein-protein interactions (PPIs) play many important roles in physiology and disease, yet are often difficult to inhibit by synthetic molecules. Peptides are well-suited for targeting the typically large PPI interfaces because their intermediate size allows them to bind to broad protein surfaces with high specificity. Multicyclic peptide-based inhibitors are particularly appealing for their greater structural rigidity and increased resistance to proteolytic degradation. In this study, we utilized our recently described cysteine-directed proximity-driven bicyclization cross-linker, bischlorooxime N-hydroxysuccinimide (BC-OSu), to construct N-terminus-capped bicyclic phage libraries. These bicyclic phage libraries were screened against two protein targets, Keap1 and the SARS-CoV-2 Spike protein. For both protein targets, multiple bicyclic ligands were readily identified with superior potency over those of their disulfide precursors. Importantly, these N-terminus-capped peptide bicycles exhibited remarkable stability in human serum, while their disulfide precursors degraded rapidly and primarily through N-terminal truncation. These results collectively showcase the potential and stability advantages of the N-terminus-capped bicyclic phage libraries.