Peptide Stapling via Phase-Transfer Tetrazine Chemistry
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Solution Overview
Problem
Current methods for incorporating and removing staples from peptides and proteins are inefficient, often causing decomposition or disrupting the native structure, especially when dealing with reactive side-chains like arginine and methionine.
Innovation Solution
A phase-transfer reaction system is used to incorporate and remove S,S-tetrazine moieties into peptides and proteins, allowing for the stabilization and unstabilization of these molecules by forming and breaking S,S-tetrazine bonds between cysteine or homocysteine residues, without decomposing peptides with reactive side-chains.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If conventional stapling methods are used to incorporate staples into peptides and proteins, then staple incorporation can be achieved, but the peptides decompose or the native structure is disrupted
Solution Approach 1:
The patent employs a phase-transfer catalyst as an intermediary substance that facilitates the reaction between the staple (e.g., S,S-tetrazine moiety) and the peptide/protein without directly interacting with or decomposing the peptide. This mediator enables efficient staple incorporation while maintaining peptide stability and native structure, resolving the contradiction between manufacturing efficiency and reliability.
2Adaptability or versatility
If reactive side-chains like arginine and methionine are present in the peptide, then the peptide has functional properties, but the stapling process causes decomposition
Solution Approach 1:
The patent applies local quality by creating a specific microenvironment at the reaction interface through phase-transfer catalysis. The reactive side-chains (arginine and methionine) remain in their native functional state within the peptide, while the staple incorporation occurs locally at the cysteine residue sites without affecting other parts of the peptide. This localized approach preserves functional properties while preventing decomposition.
3Ease of operation
If staple removal methods are used to unstabilize peptides, then the peptide can be returned to native conformation, but the process is inefficient or disruptive
Solution Approach 1:
The patent implements dynamics by using a reversible, controllable stapling mechanism. The S,S-tetrazine staple can be dynamically introduced and removed based on experimental needs. The phase-transfer catalysis system enables efficient and controllable unstapling, transforming a static, irreversible process into a dynamic, reversible one that can be efficiently controlled and optimized.
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 method enables the rapid and efficient incorporation of tetrazine staples into peptides and proteins, maintaining structural integrity and allowing for reversible unstapling, thereby enhancing stability and bioavailability.
Implementation Method 1
A phase-transfer reaction system is used to incorporate and remove S,S-tetrazine moieties into peptides and proteins
Implementation Method 2
irradiating the S,S-tetrazine moiety with light to form an amino acid sequence comprising two proximal thiocyanate moieties
Data Source
AI summary
The present disclosure pertains to the field peptide stapling and/or macrocyclization, where a structural motif is used to improve the properties of amino acid sequences (e.g. protease resistance, cellular penetration, biological activity). Also within the scope of the disclosure are methods for unstapling the S,S-tetrazine-containing amino acid sequence. The disclosure is also directed to methods for the reductive removal of thiocyanates from an amino acid sequence with cysteine to recycle back to the native amino acid sequence.