Peptide Thioester Synthesis via Mild Acid Cleavage
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Solution Overview
Problem
Current methods for producing peptide thioester compounds face challenges such as poor condensation efficiency, amino acid racemization, and disruption of oligosaccharide chain structures during peptide synthesis, particularly when dealing with glycopeptides having unprotected hydroxyl groups.
Innovation Solution
A process involving solid-phase synthesis using a resin modified with a specific linker, followed by cleavage with dilute acids and reaction with a thiol compound at low temperatures in the presence of a condensing agent, to produce peptides with a carboxyl group at the C-terminus and minimize racemization.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If a safety catch linker is used on solid-phase resin for thioesterification, then peptide excision is achieved, but condensation efficiency deteriorates and amino acid racemization occurs
Solution Approach 1:
The invention changes the chemical parameters of the linker system by using a novel linker structure with specific functional groups (trityl group and acid-labile group) that enable mild excision conditions. This allows the reaction to proceed with high condensation efficiency while preventing amino acid racemization, resolving the contradiction between ease of manufacture and productivity.
2Ease of manufacture
If a safety catch linker is used on solid-phase resin for thioesterification, then peptide excision is achieved, but thiol compound reactivity deteriorates
Solution Approach 1:
The invention modifies the reaction conditions by controlling temperature (−78°C to 0°C) and using specific condensing agents that enhance thiol compound reactivity. The novel linker structure also facilitates mild excision conditions that preserve thiol compound integrity, resolving the contradiction between ease of manufacture and reliability.
3Stability of the object's composition
If hydroxyl group of oligosaccharide chain is left unprotected, then glycopeptide structure is preserved, but alkylation of sugar hydroxyl group occurs during safety catch linker activation
Solution Approach 1:
The invention applies preliminary protective measures by using a novel linker structure that prevents alkylation of sugar hydroxyl groups during activation. The linker's specific chemical structure and mild activation conditions prevent harmful alkylation reactions while preserving the native oligosaccharide chain structure, resolving the contradiction between stability and harmful factors.
4Ease of manufacture
If strong acid is used for peptide excision from solid-phase resin, then excision is achieved, but deprotection of peptide side chains and cleavage of oligosaccharide chain linkage occurs
Solution Approach 1:
The invention fundamentally changes the excision conditions by using mild acids (acetic acid, formic acid, or dilute hydrochloric acid) instead of strong acids. The novel linker structure is designed to be cleavable under these mild conditions, enabling peptide excision without deprotecting side chains or cleaving oligosaccharide linkages, thus resolving the contradiction between ease of manufacture and compositional stability.
5Stability of the object's composition
If trityl resin is used as solid phase with acetic acid for excision, then mild excision is achieved, but glycopeptides with unprotected hydroxyl group cannot be produced
Solution Approach 1:
The invention creates a universal linker system that works for both glycosylated and non-glycosylated peptides. The novel linker structure combines the mild excision properties of trityl resin with the ability to produce glycopeptides having unprotected hydroxyl groups, making the method universally applicable and resolving the contradiction between stability and adaptability.
6Stability of the object's composition
If HMPB resin is used as solid phase with 1% TFA for excision, then mild excision is achieved, but glycopeptide excision is incomplete and side chain deprotection occurs
Solution Approach 1:
The invention optimizes the excision parameters by using acetic acid or formic acid instead of TFA, and by adjusting the linker structure to be cleavable under these milder conditions. This achieves complete excision of glycopeptides while maintaining side chain protection and preventing deprotection, resolving the contradiction between stability and productivity.
7Reliability
If C-terminal amino acid is replaced by glycine to prevent racemization, then racemization is reduced, but peptide type diversity is limited
Solution Approach 1:
The invention changes the reaction parameters by controlling temperature (−78°C to 0°C) and using specific condensing agents that prevent racemization without requiring glycine substitution. This allows the use of any C-terminal amino acid, maintaining peptide diversity while preventing racemization, thus resolving the contradiction between reliability and adaptability.
8Object-generated harmful factors
If protecting groups are added in advance to prevent alkylation, then alkylation is prevented, but additional protection and deprotection steps are required
Solution Approach 1:
The invention applies preliminary anti-action by designing a novel linker structure that inherently prevents alkylation of sugar hydroxyl groups during activation. This eliminates the need for additional protecting groups and deprotection steps, resolving the contradiction between harmful factor prevention and process complexity.
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 effectively maintains side chain protecting groups and preserves oligosaccharide chain structures, achieving efficient production of peptide thioester compounds with reduced racemization, enabling the synthesis of glycoproteins with uniform oligosaccharide chains.
Implementation Method 1
forming a peptide by a solid-phase synthesis method using a resin modified with a linker represented by formula (1) as a solid phase
Implementation Method 2
cleaving a bond between the solid phase and the peptide with at least one acid selected from dilute hydrochloric acid, dilute sulfuric acid, formic acid, and acetic acid to produce a peptide having a carboxyl group at the C-terminus
Implementation Method 3
reacting a thiol compound with the peptide at −100 to 0° C. in the presence of a condensing agent in a solvent
Data Source
AI summary
The present invention provides a process for producing a peptide thioester compound. The process involves: (A) forming a peptide by a solid-phase synthesis method using a resin modified with a linker represented by the formula (1) as a solid phase:wherein R1 represents C1-4 alkyl group, R2 represents hydrogen atom or C1-4 alkoxy group, and n represents an integer of 1 to 4; (B) cleaving a bond between the solid phase and the peptide with at least one acid selected from dilute hydrochloric acid, dilute sulfuric acid, formic acid, and acetic acid to produce a peptide having a carboxyl group at the C-terminus; and (C) reacting a thiol compound with the peptide at −100 to 0° C. in the presence of a condensing agent in a solvent.