Codon 3-5 Sequence Tuning for Precise Protein Translation Control
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Solution Overview
Problem
Existing methods for manipulating gene expression to alter protein levels are time-consuming, organism-specific, costly, and lack the ability to tightly control the extent of protein reduction or increase, often being inconsistent.
Innovation Solution
Modifying the nucleic acid sequence at codons 3, 4, and 5 by inserting or changing nucleotides to create a modified nucleic acid sequence that encodes a second protein, which can be expressed in a cell or cell-free system, allowing for precise control over protein expression levels.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If existing methods for manipulating gene expression are used, then protein levels can be altered, but the methods are time-consuming, costly, and lack tight control over the extent of protein reduction or increase
Solution Approach 1:
The invention changes the nucleotide parameters at specific codon positions (3, 4, and 5) to modulate translation efficiency. By substituting nucleotides at these specific positions while maintaining the same amino acid sequence, the method achieves precise control over protein expression levels without altering the protein's functional sequence, thereby resolving the contradiction between control precision and time efficiency.
Solution Approach 2:
The invention applies local quality changes by modifying only specific positions (codons 3, 4, and 5) within the coding sequence rather than making global changes. This localized modification approach allows for precise control of translation efficiency at specific regions of the mRNA, enabling tight control over protein expression levels while minimizing the overall changes to the gene sequence, thus reducing time and resource requirements.
2Reliability
If existing methods for manipulating gene expression are used, then protein levels can be altered, but the methods are inconsistent and organism-specific
Solution Approach 1:
The invention establishes a universal parameter change strategy by modifying codons at positions 3, 4, and 5, which are universally recognized by the genetic code across different organisms. This approach creates a consistent and reliable method for controlling translation efficiency that can be applied across various species without requiring organism-specific optimizations, thereby improving reliability while maintaining method simplicity.
Solution Approach 2:
The invention achieves universality by designing a codon modification approach that functions across different organisms. The specific nucleotide substitutions at codons 3, 4, and 5 create a universal mechanism for controlling translation efficiency that is not limited to a single organism type, enabling consistent application in bacteria, yeast, mammalian cells, and other systems without requiring method redesign.
3Manufacturing precision
If nucleic acid sequence is modified to control translation efficiency, then protein expression levels can be precisely modulated, but the amino acid sequence may change
Solution Approach 1:
The invention applies parameter changes at the nucleotide level rather than the amino acid level. By specifically modifying nucleotides at positions 3, 4, and 5 of codons while maintaining the same amino acid encoding, the method achieves precise control over translation efficiency without altering the protein's amino acid sequence, thus resolving the contradiction between expression control precision and sequence fidelity.
Solution Approach 2:
The invention creates a copy of the original coding sequence with modified nucleotides at specific positions. The modified sequence serves as a functional copy that maintains the same amino acid sequence (and thus the same protein function) while having altered translation efficiency properties. This copying approach allows precise modulation of expression levels without compromising the integrity of the amino acid sequence.
Data Source
AI summary
The present disclosure relates to compositions and methods to modulate the level of expression of a protein in a deliberate manner (i.e., tunable regulation of expression) with only a minimal change to the genetic sequence of the gene of interest. The present disclosure therefore also provides compositions and methods to predictably alter protein abundance.