Recombinant Resilin Secretion and Purification
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Solution Overview
Problem
Current methods for producing recombinant elastomeric proteins like resilin suffer from low purity due to cellular contamination and intracellular accumulation, leading to reduced efficiency and mechanical property defects, necessitating improved expression and purification techniques that recover proteins from extracellular portions.
Innovation Solution
A method involving culturing recombinant host cells expressing a secreted resilin coding sequence, primarily using yeast cells, to secrete recombinant resilin proteins, which are then purified from fermentation broths using centrifugation and chaotrope treatments, achieving high yields and purity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If recombinant host cells are lysed to extract recombinant resilin protein, then protein production is achieved, but protein purity is reduced due to cellular contamination
Solution Approach 1:
The patent extracts the target recombinant resilin protein from the cellular environment by selectively removing cellular contaminants through purification steps, separating the protein of interest from cellular debris and other proteins to achieve high purity
Solution Approach 2:
The patent employs an intermediary purification process that facilitates the separation of recombinant resilin from cellular components, using selective binding or precipitation mechanisms to isolate the protein while leaving contaminants behind
2Quantity of substance
If recombinant resilin is accumulated intracellularly, then protein production increases, but production efficiency decreases due to toxicity
Solution Approach 1:
The patent removes the harmful accumulation of recombinant resilin from the intracellular environment by implementing secretion strategies and purification steps that extract the protein from cells, preventing toxic buildup while maintaining production
Solution Approach 2:
The patent incorporates feedback mechanisms that monitor and regulate recombinant resilin production and secretion, adjusting expression levels to prevent intracellular accumulation and toxicity while optimizing overall productivity
3Device complexity
If simple precipitation-based purification is used, then purification process is simplified, but resulting protein has low purity due to cellular protein contamination
Solution Approach 1:
The patent introduces an intermediary purification mechanism that enhances the simple precipitation process, adding selective binding or chromatography steps that specifically target and remove cellular proteins while maintaining the simplicity of the overall process
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 approach results in high-yield, high-purity recombinant resilin production with enhanced mechanical properties, overcoming the limitations of traditional methods by minimizing cellular contamination and toxicity, and allowing for scalable and efficient protein production.
Implementation Method 1
recombinant host cells secrete a recombinant resilin protein encoded by said secreted resilin coding sequence
Implementation Method 2
purifying said recombinant resilin protein from said fermentation
Implementation Method 3
purifying said recombinant resilin protein from said fermentation
Data Source
AI summary
This invention relates to elastomeric protein and elastomeric protein production. In particular, the invention is directed to elastomeric protein sequences, including methods and compositions for production of elastomeric protein sequences, such as expression constructs, and host cells, and including compositions generated from the elastomeric protein sequences.