PPR First-Motif Amino Acid Changes to Reduce Protein Aggregation
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Solution Overview
Problem
PPR proteins designed with multiple motifs exhibit aggregation properties when expressed in cultured animal cells, hindering their effective use in nucleic acid manipulation techniques.
Innovation Solution
Modify the 6th and 9th amino acids of the first motif in PPR proteins to hydrophilic amino acids, such as asparagine or aspartic acid, to reduce aggregation and enhance binding specificity to nucleic acids.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If PPR proteins are expressed in cultured animal cells, then they can perform nucleic acid binding functions, but aggregation occurs reducing their effectiveness
Solution Approach 1:
The patent applies parameter changes by modifying the amino acid sequence of PPR proteins, specifically changing the 6th and 9th amino acids of the first motif to hydrophilic amino acids. This chemical parameter change reduces aggregation while preserving nucleic acid binding function, directly resolving the contradiction between functionality and aggregation.
2Measurement precision
If multiple PPR motifs are linked to create high-performance PPR proteins, then binding specificity improves, but aggregation property increases
Solution Approach 1:
The patent applies local quality by making a specific local modification to the PPR protein structure - changing only the 6th and 9th amino acids of the first motif while leaving the rest of the multiple motif structure intact. This localized change reduces aggregation without compromising the overall binding specificity provided by the multiple motifs.
Solution Approach 2:
The patent modifies the chemical parameters of specific amino acid residues (6th and 9th positions) to hydrophilic types, which changes the local physicochemical properties of the protein. This parameter change reduces aggregation tendency while maintaining the binding specificity function of the multiple PPR motifs.
Data Source
AI summary
In order to improve aggregation property of a PPR protein, the A6 amino acid of the 1st PPR motif from the N-terminus (M1) is made more hydrophilic. Further, the A9 amino acid of M1 is made to be a hydrophilic amino acid or glycine. The A6 amino acid is preferably asparagine or aspartic acid, and the A9 amino acid is preferably glutamine, glutamic acid, lysine, or glycine. Proteins containing such a PPR motif as M1 motif may have not only improved aggregation property, but also high binding power to a target nucleic acid.