Citrate Synthase D5V Mutation for L-Amino Acid Production
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for improving amino acid production in Corynebacterium glutamicum, such as fermentation technology and recombinant DNA techniques, face limitations in enhancing citrate synthase activity and amino acid biosynthesis efficiency.
Innovation Solution
Introduction of a polynucleotide encoding a citrate synthase polypeptide with an amino acid sequence modification at position 5, specifically replacing L-aspartic acid with L-valine, and optional conservative amino acid exchanges, integrated into bacteria like Corynebacterium glutamicum and Escherichia coli to enhance L-amino acid production during fermentation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional fermentation technology and recombinant DNA techniques are used to improve amino acid production, then production levels can be enhanced, but the efficiency of citrate synthase activity and amino acid biosynthesis remains limited
Solution Approach 1:
The invention applies parameter changes by modifying the amino acid sequence of citrate synthase at position 5 (replacing L-aspartic acid with L-valine) to optimize enzyme function. This specific parameter modification in the protein structure leads to improved citrate synthase activity and enhanced amino acid production efficiency, resolving the contradiction between productivity and enzyme efficiency
Solution Approach 2:
The invention applies local quality by making a targeted local modification at position 5 of the citrate synthase polypeptide chain. This localized amino acid substitution (D5V) specifically enhances the catalytic efficiency of citrate synthase without affecting the overall structure and function of the enzyme, thereby improving amino acid biosynthesis efficiency while maintaining productivity
2Quantity of substance
If amino acid production is improved through existing methods, then yield increases, but further enhancement of biosynthesis efficiency faces limitations
Solution Approach 1:
The invention changes the biochemical parameter of citrate synthase by substituting L-aspartic acid with L-valine at position 5, which optimizes the enzyme's catalytic properties. This parameter change in the enzyme structure accelerates the biosynthesis rate of amino acids, thereby improving both the yield and productivity simultaneously
3Productivity
If citrate synthase activity is enhanced through mutation, then amino acid production improves, but the complexity of strain development increases
Solution Approach 1:
The invention applies local quality by implementing a single, specific amino acid substitution (position 5: L-aspartic acid to L-valine) in the citrate synthase gene. This focused local modification simplifies the strain development process compared to multiple gene manipulations, while still achieving improved amino acid production, thus reducing the complexity of strain development
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The modified citrate synthase activity leads to improved L-amino acid production, particularly L-lysine, L-valine, and L-isoleucine, with strains showing increased enzyme activity and efficiency in amino acid secretion or accumulation, optimizing fermentation processes.
Implementation Method 1
the polypeptide possesses citrate synthase activity (EC No. 4.1.3.7)
Implementation Method 2
Fermentation of the bacteria of the genus Corynebacterium according to the invention in a suitable nutrient medium, and accumulation of the L-amino acid in the nutrient medium or in the cells of the bacteria
Data Source
Figure 1
AI summary
The invention relates to novel polynucleotides, coding for a polypeptide with citrate synthase activity, bacteria comprising the same, preferably of the species Corynebacterium and Escherichia, and methods for producing amino acids using said bacteria. The invention particularly relates to an isolated polynucleotide, coding for a polypeptide with the amino acid sequence SEQ ID NO:2, wherein the L-aspartic acid in position 5 of the amino acid sequence is replaced by another proteinogenic amino acid and the polypeptide has citrate synthase activity.