J Domain Polypeptides Enhance Recombinant Protein Folding
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Solution Overview
Problem
Current methods for producing recombinant proteins face challenges in achieving proper folding, leading to inefficient expression and accumulation of improperly folded proteins, which can result in inactive aggregates and loss of functional protein, contributing to diseases such as prion-associated diseases, Alzheimer's, and cystic fibrosis.
Innovation Solution
Incorporation of protein expression enhancing polypeptides, specifically J domain analogs, into fusion proteins to enhance the expression and proper folding of target proteins by facilitating interaction with the Hsp70 chaperone machinery, thereby increasing the yield of properly conformed functional proteins.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If recombinant protein expression is increased in host cells, then production quantity improves, but proper folding becomes more difficult leading to aggregation and loss of function
Solution Approach 1:
The patent introduces J domain polypeptides as intermediary components that mediate between the Hsp70 chaperone system and client proteins requiring folding. These J domain polypeptides act as molecular mediators that stimulate ATP hydrolysis by Hsp70, thereby enhancing the chaperone's ability to facilitate proper folding of recombinant proteins at high expression levels, preventing aggregation while maintaining high productivity
2Reliability
If Hsp70 chaperone machinery is used to facilitate protein folding, then proper conformation is achieved, but the process requires multiple components including J proteins and NEFs increasing system complexity
Solution Approach 1:
The patent extracts and utilizes specifically the J domain polypeptide component from the complex Hsp70 chaperone machinery. By isolating and applying only the J domain portion, the invention simplifies the system while retaining the essential function of stimulating Hsp70 ATP hydrolysis to promote protein folding, thereby reducing complexity while maintaining effectiveness
3Productivity
If improperly folded proteins accumulate, then functional protein yield decreases, but disease states such as prion-associated diseases, Alzheimer's, and cystic fibrosis may be contributed to
Solution Approach 1:
The patent converts the potentially harmful accumulation of misfolded proteins into a beneficial process by enhancing the Hsp70-J domain chaperone system. This converted system actively promotes proper folding of proteins that would otherwise misfold and aggregate, transforming a harmful situation into an opportunity to produce functional proteins while preventing disease-contributing aggregates
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 use of J domain analogs in fusion proteins significantly increases the expression levels of target proteins, ensuring they are properly folded and functional, effectively addressing the challenges of protein misfolding and aggregation, and improving protein production yields.
Implementation Method 1
Binding by the J protein-client complex to Hsp70-ATP stimulates ATP hydrolysis, which causes a conformational change in the Hsp70 protein
Data Source
Figure 1A~1C
Figure 2A~2B
Figure 3A~3B
AI summary
Fusion proteins comprising a protein expression enhancing polypeptide linked to a target protein binding domain and nucleic acid molecules encoding such fusion proteins are described for use in enhancing expression and/or location of a targeted protein of interest, for restoring lost functions in cells, and for treating disease. Additional fusion proteins comprising a target protein of interest modified with a fusion partner comprising a protein expression enhancing polypeptide are also disclosed.