Cyclic Peptide Library Synthesis via Disulfide Bridge Release
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Solution Overview
Problem
The development of cyclic peptide therapeutics is hindered by low yields in macrocyclization reactions and the need for costly chromatographic purification, limiting the size and efficiency of peptide libraries due to the difficulty in transforming linear molecules into cyclic ones and the lack of efficient methods for synthesizing large macrocycle libraries.
Innovation Solution
A method involving the release of linear dithiol peptides from a solid phase using a volatile reducing agent or a base to induce intramolecular disulfide exchange, allowing for the synthesis of cyclic peptides without the need for purification, enabling high-throughput screening of large peptide libraries.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If conventional macrocyclization reactions are used to transform linear peptides into cyclic peptides, then cyclic peptide products are obtained, but the yields are low (below 90%) and require costly chromatographic purification
Solution Approach 1:
The patent applies preliminary action by incorporating a disulfide bridge during the solid-phase peptide synthesis step, before the macrocyclization reaction. This pre-formed disulfide bridge serves as an intramolecular crosslink that directs the subsequent cyclization reaction, ensuring high yield and eliminating the need for purification. The disulfide bridge is introduced in advance as part of the linear peptide structure, enabling clean transformation into the cyclic product without side products.
Solution Approach 2:
The disulfide bridge acts as an intermediary element that mediates between the linear peptide precursor and the final cyclic peptide product. It serves as both a structural component that enables cyclization and a purification-free pathway, facilitating the transformation while maintaining high yield and product purity without requiring chromatographic separation.
2Manufacturing precision
If individual purification is performed after macrocyclization for each peptide, then pure cyclic peptide products are obtained, but the library size is limited to below 30,000 molecules
Solution Approach 1:
The patent merges the synthesis and purification steps into a single operation by using the disulfide bridge method. Instead of performing individual purification for each peptide in the library, the disulfide bridge approach enables parallel synthesis of thousands of cyclic peptides with inherent purity, combining multiple operations into one high-throughput process that supports large library sizes.
Solution Approach 2:
The disulfide bridge structure provides self-service purification by inherently directing the cyclization reaction to proceed cleanly without forming side products. The intramolecular crosslinking automatically ensures product purity, eliminating the need for external purification operations and enabling large-scale library production.
3Manufacturing precision
If bis-elecrophilic reagents are used for macrocyclization, then high cyclization yields (above 90%) are achieved, but the method requires specific reagent conditions and substrate requirements
Solution Approach 1:
The patent extracts the cyclization function from the complex bis-elecrophilic reagent system and incorporates it directly into the peptide structure through the disulfide bridge. This removes the need for external electrophilic reagents and their associated complex conditions, simplifying the overall process while maintaining high yield through the inherent reactivity of the disulfide-linked substrate.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enables the efficient generation of large cyclic peptide libraries with high yields and reduces the complexity of the synthesis process, facilitating the screening of peptides for therapeutic applications by eliminating the need for costly purification steps and improving the efficiency of peptide library production.
Implementation Method 1
an agent reducing the disulfide bridge, thereby releasing the one or more linear dithiol peptides from the solid phase
Implementation Method 2
a base that deprotonates the sulfhydryl group in the N-terminal region of the one or more linear dithiol peptides, thereby inducing an intramolecular disulfide exchange
Implementation Method 3
inducing an intramolecular disulfide exchange thereby releasing the one or more linear dithiol peptides from the solid phase in the form of one or more cyclic peptides
Implementation Method 4
wherein the agent is volatile and is removable by evaporation
Data Source
AI summary
The present invention relates to a method for preparing a library of peptides or an isolated peptide comprising (a) releasing one or more linear dithiol peptides carrying a sulfhydryl group in the N-terminal region of the one or more peptides and being immobilized via a disulfide bridge in the C-terminal region of the one or more peptides on a solid phase from the solid phase by (i) an agent reducing the disulfide bridge, thereby releasing the one or more linear dithiol peptides from the solid phase, wherein the agent is volatile and is removable by evaporation, or (ii) a base that deprotonates the sulfhydryl group in the N-terminal region of the one or more linear dithiol peptides, thereby inducing an intramolecular disulfide exchange thereby releasing the one or more linear dithiol peptides from the solid phase in the form of one or more cyclic peptides.