Proline-Rich Peptide Tags for Soluble Protein Expression
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Existing methods for producing recombinant proteins in a soluble fraction are inefficient, particularly when using peptide tags like the ENTROPIC BRISTLE DOMAIN (EBD) that contain serine between prolines, leading to low total protein production.
Innovation Solution
The use of a peptide tag with a specific amino acid sequence, such as X(PY)qPZ, where X, Y, and Z are defined by specific amino acids, improves the expression level of proteins in a soluble fraction, ensuring the tag contains multiple Q, M, L, N, or T residues, enhancing solubility and production efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If a peptide tag containing serine between prolines (e.g., EBD peptide) is used to improve protein expression in soluble fraction, then the expression of useful protein is improved, but total protein production becomes low
Solution Approach 1:
The patent modifies the amino acid composition and sequence parameters of the peptide tag. Specifically, it replaces serine residues between prolines with alternative amino acids (such as glycine, alanine, or other non-serine residues) while maintaining the proline-rich structure. This parameter change resolves the contradiction by eliminating the harmful effect of serine-proline sequences that cause low total protein production, while preserving the beneficial solubility-enhancing properties through the modified peptide tag structure
Solution Approach 2:
The patent uses a simplified peptide tag structure that is easier to produce and more stable than the original EBD peptide. By replacing complex serine-proline sequences with simpler alternative sequences, the modified peptide tag achieves better overall performance in terms of total protein production while maintaining its function as a solubility-enhancing tag
2Productivity
If peptide tags are linked to improve protein expression, then expression in soluble fraction is improved, but protein aggregation and insolubility occur
Solution Approach 1:
The patent changes the amino acid sequence parameters of the peptide tag by replacing serine residues with alternative amino acids. This modification alters the physical and chemical properties of the peptide tag, reducing its tendency to cause protein aggregation while maintaining its ability to enhance solubility. The modified sequence parameters optimize the balance between expression improvement and solubility maintenance
Solution Approach 2:
The modified peptide tag acts as an intermediary structure that mediates between the protein of interest and the cellular environment. By modifying its amino acid composition (replacing serine with alternative residues), the peptide tag becomes a more effective mediator that enhances protein solubility without causing aggregation, thus resolving the contradiction between expression improvement and solubility stability
Data Source
AI summary
By linking a peptide tag having the amino acid sequence X(PY)qPZ to a protein of interest, and then expressing the protein, the protein of interest is efficiently accumulated in a soluble fraction, where P is proline; X is an amino acid sequence composed of 0 to 5 amino acids independently selected from R, G, S, K, T, L, N, Q, and M; Y is an amino acid sequence composed of 1 to 4 amino acids independently selected from R, G, K, T, L, N, Q, and M; q is an integer 1 to 10; and Z is an amino acid sequence composed of 0 to 10 amino acids independently selected from R, G, S, K, T, L, N, Q, and M; where the amino acid sequence of the peptide tag contains three or more Qs, Ms, Ls, Ns, and/or Ts in total.