Lysine Production Corynebacterium gapN Gene Knockout

Resolve Bottlenecks,
Find Innovative SolutionsGenerate Solutions

Solution Overview

Problem

Corynebacterium strains lack NADP-dependent glyceraldehyde-3-phosphate dehydrogenase activity, limiting L-lysine productivity, as existing attempts to introduce foreign gapN genes have not yielded satisfactory results.

Innovation Solution

Inactivate the endogenous glyceraldehyde-3-phosphate dehydrogenase gene (gapA) and introduce exogenous NADP-dependent glyceraldehyde-3-phosphate dehydrogenase gene (gapN) from Streptococcus mutans, Streptococcus agalactiae, or Bacillus cereus, using expression vectors like pECCG122-Pcj7-gapN, to enable NADP-dependent activity for enhanced L-lysine production.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If foreign gapN genes are introduced into Corynebacterium strains, then L-lysine productivity should be enhanced through increased NADPH levels, but the endogenous gapA gene continues to compete for substrate and coenzyme, limiting the effectiveness of gapN introduction

Engineering Contradiction:

ImproveL-lysine productivityVSAvoidenzymatic system complexity

Core Design Contradiction:

ProductivityVSDevice complexity

Solution Approach 1:

The patent applies the 'Taking out' principle by removing the endogenous gapA gene from the Corynebacterium strain through gene knockout technology. This eliminates the competing enzymatic pathway that was consuming glyceraldehyde-3-phosphate and NAD+, thereby allowing the introduced gapN gene to operate without competition and maximize NADPH production for L-lysine synthesis.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent applies the 'Parameter changes' principle by changing the coenzyme specificity parameter of the glyceraldehyde-3-phosphate dehydrogenase enzyme. By replacing the NAD+-dependent gapA enzyme with the NADP+-dependent gapN enzyme, the metabolic flux is redirected to produce NADPH instead of NADH, providing the reducing power needed for enhanced L-lysine productivity.

Inventive Principle:

Principle #35Parameter changes

2Adaptability or versatility

If both gapA and gapN genes are present in the strain, then metabolic flexibility is maintained, but substrate competition between NAD+-dependent and NADP+-dependent pathways reduces overall L-lysine production efficiency

Engineering Contradiction:

Improvemetabolic flexibilityVSAvoidL-lysine production efficiency

Core Design Contradiction:

Adaptability or versatilityVSProductivity

Solution Approach 1:

The patent removes the gapA gene to eliminate the competing metabolic pathway. This ensures that all glyceraldehyde-3-phosphate is directed through the gapN pathway, maximizing NADPH production and L-lysine synthesis efficiency without the dilution effect of parallel pathway competition.

Inventive Principle:

Principle #2Taking out (Extraction)

3Stability of the object's composition

If gapN gene is introduced without inactivating gapA, then the strain maintains native metabolic pathways, but the introduced gapN gene fails to achieve satisfactory L-lysine productivity enhancement

Engineering Contradiction:

Improvenative metabolic pathway stabilityVSAvoidL-lysine productivity

Core Design Contradiction:

Stability of the object's compositionVSProductivity

Solution Approach 1:

The patent applies gene knockout technology to remove the gapA gene, creating a metabolic void that is then filled by the introduced gapN gene. This ensures that the gapN pathway becomes the dominant or sole pathway for glyceraldehyde-3-phosphate metabolism, guaranteeing sufficient NADPH production for high-level L-lysine synthesis.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

Instead of adding gapN to an existing gapA-containing strain (which would create competition), the patent inverts the approach by first removing gapA and then introducing gapN. This inversion ensures that gapN operates without competition and becomes the primary metabolic pathway, achieving the desired productivity enhancement.

Inventive Principle:

Principle #13The other way round (Inversion)

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 introduction of gapN gene increases NADPH levels, providing reducing power for improved L-lysine biosynthesis, resulting in enhanced productivity in Corynebacterium strains.

Implementation Method 1

the gapN gene encoding enzyme which converts glyceraldehyde-3-phosphate into 3-phosphoglycerate using NADP as a coenzyme

Methodology Applied

Scientific EffectEnzyme catalysis: Enzyme

Implementation Method 2

produces NADPH... the resulting increase in NADPH levels is used as the source of reducing power for lysine biosynthesis

Methodology Applied

Scientific EffectRedox reaction: Redox Reactions

Data Source

PatentEP2453006B1Method for producing l-lysine using corynebacterium sp. that has obtained the activity of glyceraldehyde-3-phosphate dehydrogenase derived from an alien species

Publication Date: 2015.10.07 CJ CHEILJEDANG CORP

Figure 1~2

AI summary

The present invention relates to a Corynebacterium sp. strain having an activity of NADP-dependent glyceraldehyde-3-phosphate dehydrogenase and an improved productivity of L-lysine, and a method for producing L-lysine using the same. According to the Corynebacterium sp. strain of the present invention and the method for producing L-lysine using the same, L-lysine can be produced in a high yield.