Peptide Thioester Synthesis via Cysteine Residue Activation
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Existing methods for synthesizing peptide thioesters are limited by the requirement for non-native amino acid derivatives and specific three-dimensional structures, restricting the size and complexity of peptides that can be synthesized.
Innovation Solution
A process involving the selective activation of a cysteine residue's thiol group with a compound having a leaving group, followed by a thiol exchange reaction to convert the peptide chain into a thioester, allowing for the synthesis of longer peptides without the need for native amino acid derivatives or specific structures.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If solid-phase synthesis is used to construct peptide thioester, then peptide thioester can be obtained, but the size of synthesizable peptide is limited and non-native amino acid derivatives or linkers are required
Solution Approach 1:
The patent introduces a thionoformate group as an intermediary functional group that enables peptide bond cleavage and thioester formation. This intermediary group acts as a bridge between the peptide chain and the thioester product, allowing the transformation without requiring non-native amino acid derivatives or linkers. The thionoformate group is formed by reacting a cysteine thiol group with a thionoformate reagent, then subsequently reacts with an amide to cleave the peptide bond and form the thioester.
2Ease of manufacture
If linker-based methods are used for peptide thioester construction, then thioester can be obtained by thiolysis, but non-native amino acid derivatives must be chemically synthesized separately, complicating the procedure
Solution Approach 1:
The patent extracts the essential functional requirement (thioester formation) from the complex linker-based methods. Instead of using entire linker systems that require separate synthesis, the invention isolates and utilizes only the thionoformate functional group, which can be introduced directly to cysteine residues in native peptides. This extraction simplifies the procedure by eliminating the need for non-native amino acid derivatives while maintaining the ability to form peptide thioesters.
3Reliability
If native chemical ligation is used to ligate peptide chains, then native amide bond can be formed, but peptide thioester raw material must be prepared in advance through complex methods
Solution Approach 1:
The patent enables the peptide chain itself to serve the dual function of both substrate and reagent for thioester formation. The cysteine residue within the native peptide chain reacts with the thionoformate reagent to generate the thioester functionality in situ. This self-service approach eliminates the need for separate preparation of peptide thioester raw materials through complex solid-phase synthesis or linker-based methods, while maintaining the reliability of native amide bond formation in subsequent ligation reactions.
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 easy thioesterification of long chain polypeptides, facilitating the production of modified proteins by combining biosynthesis and solid-phase synthesis, and allows for site-specific addition of sugar chains, improving the synthesis of complex peptides.
Implementation Method 1
reacting a compound A represented by the following formula (I) with a thiol group of a cysteine residue in a peptide chain having the cysteine residue to eliminate R2
Implementation Method 2
reacting a compound B represented by the following formula (II) with the first intermediate in an organic solvent to add a -NH-C(=Y)NHR3 group to a carboxyl group forming a peptide bond with an amino acid adjacent to an N-terminal-side of the cysteine residue
Implementation Method 3
thioesterifying a C-terminal of the second intermediate by reacting the second intermediate with thiol to exchange the -NH-C(=Y)NHR3 group for the thiol group at the C-terminal
Data Source
AI summary
Provided is a novel process for chemically converting a peptide chain into a peptide thioester. The inventors paid attention to the cysteine residue of a peptide chain. As a result, the following has been found. When a -C(=X)-R1 group is introduced to the thiol group of the cysteine residue and then the resulting peptide is reacted with a compound having a leaving group represented by the formula: -NH-C(=Y)NHR3 in an organic solvent, the -NH-C(=Y)NHR3 group binds via addition reaction to the carboxyl group of the N-terminal-side peptide bond of the cysteine residue, whereby the peptide bond is cleaved and the C-terminal-side peptide fragment is cut off. Further, when the resulting peptide chain having the -NH-C(=Y)NHR3 group is reacted with a thiol in a buffer solution, a thiol exchange reaction occurs, namely, the thiol group of the thiol binds to the carbonyl carbon to which the -NH-C(=Y)NHR3 group has bound, whereby the -NH-C(=Y)NHR3 group is eliminated. Thus, the conversion into a peptide thioester is attained.