SSD Guide Protein Fusion for Peptide Production Yield
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Solution Overview
Problem
Current methods for producing peptides, such as chemical synthesis and traditional recombinant production, face challenges with yield, cost, scalability, and stability, especially for peptides between 30 and 50 amino acids, due to issues like degradation by proteases and inefficient purification processes.
Innovation Solution
The use of a guide protein derived from the SH3 domain of spectrin (SSD guide protein) in a recombinant fusion protein system, which allows for high expression yields, solubility, and simplified purification, including a cleavage site and purification tag, facilitating the production and recovery of peptides at a lower temperature range (23-26°C) for improved productivity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If chemical synthesis is used to produce short peptides, then high purity can be achieved, but the yield decreases with increasing peptide size and production cost increases
Solution Approach 1:
The patent uses a guide protein as an intermediary carrier to produce peptides. The guide protein is expressed as a fusion protein with the peptide sequence, then purified and processed to release the target peptide. This intermediary approach allows high-yield production of larger peptides while maintaining purity through the purification of the fusion protein and controlled cleavage.
2Productivity
If traditional recombinant production is used to express peptides in host cells, then scalability and cost efficiency improve, but peptide stability deteriorates due to degradation by endogenous proteases
Solution Approach 1:
The guide protein serves as a protective intermediary that shields the expressed peptide from degradation by endogenous proteases during cellular expression. The fusion protein structure and subsequent purification process ensure peptide stability while enabling scalable production.
Solution Approach 2:
The patent performs preliminary purification of the fusion protein before cleavage to release the peptide. This preliminary action removes potential proteases and other degrading factors from the system before the peptide is released, preventing subsequent degradation and ensuring long-term stability.
3Productivity
If existing guide proteins are used for peptide production, then expression yield improves, but purification difficulty increases due to additional residues and chemical modifications
Solution Approach 1:
The patent strategically removes problematic elements from the guide protein sequence. The guide protein is designed without cysteine residues that would require disulfide bond management, and without methionine at the cleavage junction that would complicate chemical cleavage. This extraction of problematic features simplifies both expression and purification while maintaining high yield.
Solution Approach 2:
The patent applies local quality optimization by specifically designing the guide protein sequence with favorable properties at critical positions: no cysteine residues throughout the sequence to avoid disulfide complications, and a specific amino acid at the cleavage junction to enable simple chemical cleavage. This localized optimization of sequence properties greatly simplifies the overall manufacturing process.
Data Source
AI summary
A peptide sequence of a guide protein for the production of a peptide of interest; a peptide sequence that has a similarity of at least 90% to SEQ. ID. NO 1; a nucleotide sequence encoding the guide protein; an expression vector comprising the nucleotide sequence; a host cell that expresses a fusion protein comprising a peptide of interest; a method of production of a peptide of interest, comprising the steps of A) constructing an expression vector; B) inserting the expression vector into a host cell; C) expressing the fusion protein, culturing the host cell in a culture medium; D) recovering the accumulated fusion protein in the host cell; E) cleaving the fusion protein; F) purifying the peptide of interest.