mRNA folding energy optimization for translation efficiency
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Solution Overview
Problem
Current methods fail to systematically optimize mRNA folding strength along the coding sequence, which affects translation efficiency and is not well understood across different species, leading to inefficiencies in protein expression.
Innovation Solution
Introducing synonymous mutations in specific regions upstream and downstream of the stop codon to increase folding energy, optimizing the coding sequence to enhance translation efficiency by modifying the mRNA folding strength.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If mRNA folding strength is increased in the coding region, then translation initiation may be improved, but translation elongation may be slowed down causing ribosomal traffic jams
Solution Approach 1:
The patent applies local quality by differentiating mRNA folding strength requirements across different regions of the coding sequence. Specifically, it strengthens folding in the 5' region (nucleotides 1-70) to enhance translation initiation while maintaining or reducing folding strength in downstream regions to prevent ribosomal traffic jams during elongation. This regional differentiation resolves the contradiction by optimizing each region's folding properties for its specific functional requirement.
2Productivity
If synonymous mutations are introduced to increase folding energy, then translation efficiency is improved, but coding sequence complexity increases
Solution Approach 1:
The patent employs parameter changes by systematically modifying the folding energy parameter of mRNA in specific coding regions through synonymous mutations. It uses computational algorithms to calculate and optimize folding free energy values, introducing mutations that increase folding energy in the 5' region (nucleotides 1-70) while controlling mutations in downstream regions. This approach improves translation efficiency by changing the folding energy parameter without altering the encoded protein sequence.
3Productivity
If folding energy is increased upstream of stop codon, then translation termination is optimized, but mRNA stability may be affected
Solution Approach 1:
The patent applies local quality by specifically targeting the region upstream of the stop codon (nucleotides -30 to 0) for folding energy optimization. It introduces synonymous mutations in this specific local region to enhance translation termination efficiency while being mindful of the potential impact on overall mRNA stability. The method carefully balances folding energy increases in this terminal region to achieve optimal termination without compromising the stability of the entire mRNA molecule.
Data Source
AI summary
Nucleic acid molecule comprising a coding sequence and a region of increased folding energy upstream of a stop codon are provided. Expression vectors and cells comprising the nucleic acid molecule are also provided. Methods for optimizing a coding sequence comprising increasing folding energy in a region upstream of the stop codon are also provided.