ATG Initiation Codon Substitution in Corynebacterium Glutamicum

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Solution Overview

Problem

Current methods for producing L-lysine in Corynebacterium glutamicum strains are limited by the suboptimal activity of enzymes such as aspartate kinase, transketolase, and pyruvate carboxylase due to their native initiation codons, which can hinder lysine biosynthesis efficiency.

Innovation Solution

The initiation codons of the lysC, tkt, and pyc genes are substituted with ATG in a modified polynucleotide, leading to enhanced enzyme activities and improved lysine productivity when introduced into Corynebacterium glutamicum strains.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If the native initiation codons (GTG or TTG) are used in lysC, tkt, and pyc genes, then the genes can be naturally expressed, but the enzyme activities are suboptimal which limits L-lysine production efficiency

Engineering Contradiction:

ImproveL-lysine production efficiencyVSAvoidenzyme activity stability

Core Design Contradiction:

ProductivityVSReliability

Solution Approach 1:

The initiation codons of lysC, tkt, and pyc genes are mutated from GTG or TTG to ATG. This parameter change in the genetic sequence improves ribosome recognition and translation initiation efficiency, thereby enhancing enzyme activities and L-lysine production while maintaining genetic stability through conservative codon substitution

Inventive Principle:

Principle #35Parameter changes

2Productivity

If the initiation codon is mutated from GTG or TTG to ATG, then translation efficiency and enzyme activity are enhanced, but the genetic sequence is modified which may affect gene stability

Engineering Contradiction:

Improveenzyme activityVSAvoidgenetic sequence stability

Core Design Contradiction:

ProductivityVSStability of the object's composition

Solution Approach 1:

The initiation codons are precisely mutated from GTG or TTG to ATG in the lysC, tkt, and pyc genes. This targeted parameter change improves translation initiation without causing frameshift or large-scale genetic instability, as ATG is a standard start codon that maintains reading frame integrity

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent uses transient expression systems and selectable markers (such as antibiotic resistance genes) to maintain and select for the mutated genes during cultivation. These auxiliary genetic elements can be discarded after serving their selection purpose, while the core initiation codon mutations remain stably integrated

Inventive Principle:

Principle #27Cheap short-living objects (Disposable)

Data Source

PatentEP3404101B1Method for producing l-lysine using microorganisms having ability to produce l-lysine

Publication Date: 2025.06.11 CJ CHEILJEDANG CORP

AI summary

The present invention relates to a modified polynucleotide encoding aspartate kinase (EC:2.7.2.4; hereinafter, referred to as LysC), transketolase (EC:2.2.1.1; hereinafter, referred to as Tkt) or pyruvate carboxylase (EC:6.4.1.1; hereinafter, referred to as Pyc), in which the initiation codon is substituted with ATG, a vector including the same, a microorganism transformed with the vector, and a method for producing L-lysine using the same.