Ankyrin Repeat Domain N-Terminal Capping Thermostability
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Solution Overview
Problem
There is a need to further improve the thermostability of ankyrin repeat domains, which are crucial for maintaining protein stability and function, especially in applications requiring high temperatures or prolonged storage.
Innovation Solution
Specific mutations in the N-terminal capping module, particularly at position 15, are introduced to enhance the thermostability of ankyrin repeat domains, improving their melting temperature and folding properties while reducing aggregation and immunogenicity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Temperature
If standard ankyrin repeat domains are used, then basic binding function is achieved, but thermostability is insufficient for high temperature applications
Solution Approach 1:
The patent applies local quality by introducing specific mutations at position 15 of the N-terminal capping module while leaving the rest of the ankyrin repeat domain structure intact. This localized modification at a specific position (position 15) optimizes thermostability without disrupting the overall folding and binding function of the domain. The specific amino acid substitutions (e.g., to alanine, valine, leucine, isoleucine, or methionine) create local structural improvements that enhance global stability.
Solution Approach 2:
The patent employs parameter changes by systematically varying the amino acid residue at position 15 of the N-terminal capping module to optimize thermostability. Different amino acids (alanine, valine, leucine, isoleucine, methionine) are tested at this specific position to determine which substitution yields the highest melting temperature and stability. This parameter optimization approach directly addresses the need for improved thermostability while maintaining functional integrity.
2Reliability
If ankyrin repeat domains are designed for high stability, then thermostability improves, but aggregation and immunogenicity may increase
Solution Approach 1:
The patent uses local quality by making a targeted substitution at position 15 of the N-terminal capping module, which locally optimizes the protein's surface properties and folding characteristics. This localized change improves overall stability and reduces aggregation propensity without requiring extensive modifications that could create new immunogenic epitopes elsewhere in the structure.
Solution Approach 2:
The patent applies parameter changes by selecting specific amino acids (alanine, valine, leucine, isoleucine, methionine) at position 15 that optimize the balance between stability and reduced aggregation. These parameter selections are based on their side chain properties that promote proper folding and reduce exposed hydrophobic surfaces, thereby minimizing aggregation and immunogenicity while maximizing thermostability.
Data Source
AI summary
Described herein are proteins comprising an ankyrin repeat domain having an N-terminal capping module with improved properties, as well as corresponding protein libraries, pharmaceutical compositions and nucleic acids encoding such proteins. In other aspects, the disclosure relates to methods using such proteins, corresponding protein libraries or pharmaceutical compositions.