Polypeptide DNA-Binding Specificity via Periodic Residue Variation
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Solution Overview
Problem
Conventional polypeptides with high DNA-binding domain activity often exhibit non-specific function in regions outside the target nucleotide sequence, posing safety concerns and requiring complex procedures for preparation, making it difficult to achieve compatibility between high function and specificity.
Innovation Solution
A polypeptide with a DNA-binding domain linked to a function domain via a polypeptide of 35 to 55 amino acid residues, where specific amino acid residue combinations every four DNA-binding modules ensure high function and specificity, allowing for simpler vector preparation using a vector set and library.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional polypeptides with high DNA-binding domain activity are used, then high function is achieved, but non-specific function occurs in regions outside the target nucleotide sequence
Solution Approach 1:
The patent applies local quality by introducing periodic variations in amino acid residues at specific positions (positions x and y) within every four DNA-binding modules. This creates localized differences in binding characteristics across otherwise identical modules, enabling the polypeptide to distinguish target sequences more precisely while maintaining high binding activity, thus reducing non-specific function.
2Reliability
If conventional polypeptides with high DNA-binding domain activity are used, then high function is achieved, but complex procedures are required for preparation
Solution Approach 1:
The patent employs periodic action by systematically varying amino acid residues at positions x and y every four DNA-binding modules. This periodic variation pattern can be easily encoded in the nucleotide sequence and propagated through standard molecular cloning procedures, simplifying the preparation process while achieving high function and specificity simultaneously.
3Object-affected harmful factors
If compatibility between high function and specificity is pursued, then safe genetic modification is achieved, but procedural complexity increases
Solution Approach 1:
The patent changes the parameters of the DNA-binding modules by specifying periodic variations at two particular sites (positions x and y) within every four modules. This parameter change approach maintains the overall modular structure and standard cloning procedures, avoiding increased procedural complexity while achieving enhanced safety through improved specificity.
Data Source
AI summary
The present invention provides an artificial nuclease comprising a DNA-binding domain and a function domain linked to each other via a polypeptide consisting of 35 to 55 amino acid residues wherein amino acid residues at two sites in a DNA-binding module contained in a DNA-binding domain exhibit a mode of repetition that is different for every four DNA-binding modules; a vector for expressing said artificial nuclease; a vector library for preparing said vector; and a vector set for preparing said vector library.