Recombinant Collagen Production via Segmented Gene Design
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Solution Overview
Problem
Current methods for producing recombinant human collagen face challenges such as low yield, purity, and stability, along with high costs and long production cycles, which hinder its application in biomedical fields due to the loss of biological activity and immune rejection issues with animal-derived collagens.
Innovation Solution
A polypeptide comprising 16 repeats of a specific sequence (GERGAPGFRGPAGPNGIPGEKGPAGERGAP) linked directly, expressed in E. coli, purified using a Ni column and anion exchange chromatography, and optionally digested with TEV protease, resulting in a collagen with enhanced cell adhesion and stability, reducing the need for additional stabilizing agents and minimizing endotoxin levels.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If animal-derived tissues are used to extract collagen through acid, alkali, and enzymolysis methods, then collagen can be obtained, but the original biological activity is lost and immune rejection occurs
Solution Approach 1:
The patent uses recombinant DNA technology to copy the human collagen gene sequence and express it in a microbial system (E. coli), creating a synthetic version of human collagen that retains the original biological activity without requiring animal tissue extraction. This copying approach produces collagen with the exact amino acid sequence as native human collagen.
2Reliability
If transgenic techniques are used to prepare recombinant human collagen in animal expression systems, then biological activity is retained, but production cost increases and production cycle lengthens
Solution Approach 1:
The patent introduces a microbial expression system (E. coli) as an intermediary platform to produce human collagen. This intermediary system combines the advantages of both animal systems (retaining biological activity through correct protein folding and post-translational modifications) and microbial systems (high growth rate, low cost, short generation time), achieving efficient production of bioactive collagen.
3Reliability
If the natural collagen gene sequence is used for expression, then the correct spatial structure may be formed, but the complex structure with supercoiling and multiple chains makes mass production difficult
Solution Approach 1:
The patent segments the complex natural collagen gene sequence into a simplified repeating unit (Gly-X-Y triplet pattern) that can be easily cloned and expressed. The segmented design uses 16 repeats of a specific sequence that self-assembles into the correct triple-helical structure, making mass production feasible while maintaining spatial structure integrity.
4Reliability
If collagen is designed to form triple helix structure with specific amino acid residues, then biological function is achieved, but the requirement for precise structural organization reduces production yield
Solution Approach 1:
The patent optimizes parameters including the specific amino acid sequence (Gly-Ala-Gly-Pro-Hyp pattern), the number of repeats (16 copies), and the pH conditions during expression to maximize both triple helix formation efficiency and production yield. These parameter changes enable high-yield production of functional collagen.
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
The approach yields recombinant collagen with improved stability in aqueous solutions, higher purity, and reduced endotoxin levels, enabling long-term use and increased cell adhesion activity compared to existing recombinant type III collagens, facilitating its application in medical devices, tissue engineering, and cosmetics.
Implementation Method 1
purifying the polypeptide by a Ni column and/or an anion exchange chromatography
Implementation Method 2
purifying the polypeptide by a Ni column and/or an anion exchange chromatography
Implementation Method 3
optionally digesting the polypeptide, preferably digesting the polypeptide with TEV protease
Data Source
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AI summary
The present invention relates to a polypeptide, a process for the production thereof and a use thereof. The polypeptide of the present invention has excellent adhesion effect and is highly stable in an aqueous solution. Endotoxins can be more easily removed from the product of the polypeptide.