Disulfide-Directed Multicyclic Peptide Libraries for the Discovery of Peptide Ligands and Drugs

Abstract

Multicyclic peptides with stable 3D structures are a kind of novel and promising peptide formats for drug design and discovery as they have the potential to combine the best characteristics of small molecules and proteins. However, the development of multicyclic peptides is largely limited to naturally occurring products. It remains a big challenge to develop multicyclic peptides with new structures and functions without recourse to the existing natural scaffolds. Here, we report a general and robust method relying on the utility of new disulfide-directing motifs for designing and discovering diverse multicyclic peptides with potent proteinbinding capability. These peptides, referred to as disulfide-directed multicyclic peptides (DDMPs), are tolerant to extensive sequence manipulations and variations of disulfide-pairing frameworks, enabling the development of de novo DDMP libraries useful for ligand and drug discovery. This study opens a new avenue for creating a new generation of multicyclic peptides in sequence and structure space inaccessible by natural scaffolds, thus would greatly benefit the field of peptide drug discovery.

De Novo Discovery of Cysteine Frameworks for Developing Multicyclic Peptide Libraries for Ligand Discovery

“…This selection can be influenced by the vast number of possible cysteine frameworks that can be configured through the adjustment of both the number and positioning of cysteine residues as well as the length of sequences. Given that our previous works have demonstrated the important role of some unique biscysteine motifs including CXC, CPPC, and CPXXC (C: cysteine; P: proline; X: any amino acid) in directing the disulfide pairing and folding of peptides, − we thus set the primary aim of this work to identify peptide sequences with a pair of CXC and CPPC motifs and two additional cysteine residues. As a proof of concept, we designed a 25-mer random sequence template, into which CPPC and CXC motifs and an isolated cysteine residue were incorporated (Figure b).…”

“…We believe that our “touchstone”-based strategy should have wide applicability to exploring the foldability of sequences with diverse cysteine frameworks, opening a new avenue for discovering many different kinds of DRPs. Particularly, it should be interesting to explore the foldability of our previously designed DRPs bearing two identical biscysteine motifs with intrinsic propensity of dimeric disulfide pairing − using the present strategy, which would enable the identification of specific cysteine frameworks with more robust foldability. Given the huge number of random sequences, selection-enriched cysteine frameworks are particularly useful for the development of peptide libraries because they can tolerate more extensive sequence variations, thus enabling access to broader regions of sequence space to discover new peptide ligands.…”

“…By following the same procedures as those before, we designed and prepared a library bearing a pair of CPPC and CPXXC motifs and an additional cysteine (Figure a), which was applied to select against MDM2. CPXXC is a new motif with superior capability of directing oxidative folding of peptides that has been identified more recently . We anticipated that the combination of CPPC and CPXXC motifs would lead to the de novo discovery of many unique DRP scaffolds.…”

“…We noticed that natural evolution leverages conserved cysteine frameworks to select sequences that are foldable in structure to diversify DRPs. − This strategy has recently been utilized artificially to design new DRPs and the corresponding peptide libraries . In addition, our works in exploiting orthogonal disulfide-directing motifs for designing DRPs with high tolerance to sequence manipulations further underline the usefulness of well-designed cysteine frameworks for creating diverse DRPs with new structures and functions. − However, the rational design of new cysteine frameworks amenable to peptide library generation is subject to the preknowledge of certain disulfide patterns directed by several precious biscysteine motifs. − We argued that the diversity of DRPs would be greatly expanded if rational design or selection strategies were available to generate a variety of new cysteine frameworks with unpredicted disulfide patterns. Such cysteine frameworks would serve as useful templates for the generation of multicyclic DRP libraries and the subsequent selection of functional peptide ligands with therapeutic potentials.…”

Nanoscale 0D/1D Heterojunction of MAPbBr3/COF Toward Efficient LED-Driven S–S Coupling Reactions

“…Disulfide bonds , hold multiples essential roles in life science, protein folding, , pharmacy, − material science, and advanced chemical application , due to their superior reversibility, flexibility, and stability. Specifically, the stable covalent bond of S–S and flexible ligands enable three-dimensional folding and cross-linking, which contribute to the assembly of proteins from polypeptides and the vulcanization of polymer rubber .…”