Optically Active Amine Production via Transaminase Extraction
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Solution Overview
Problem
Current methods for producing optically-active amine compounds using transaminases face challenges in achieving high yields due to reversible reaction equilibria, often requiring expensive coenzymes like NADH and complicating the reaction system with additional enzymes, leading to inefficiencies and increased costs.
Innovation Solution
A method involving the concomitant use of a transaminase, an α-keto acid reductase, and a coenzyme regenerating enzyme, such as glucose dehydrogenase, to convert ketone compounds into optically-active amine compounds without the need for expensive coenzymes like NADH, by biasing the reaction equilibrium towards the desired product.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If a transaminase is used to produce optically-active amine compounds through amination reaction, then the desired amine compound can be produced, but the reversible reaction equilibrium limits the yield and requires additional measures to bias the equilibrium
Solution Approach 1:
The patent removes the by-product α-keto acid from the reaction system through extraction with an organic solvent. This extraction process continuously removes the by-product from the aqueous phase where the transaminase reaction occurs, shifting the reversible equilibrium toward product formation and significantly improving the yield of optically-active amine compounds without requiring additional enzymes or expensive coenzymes.
Solution Approach 2:
The patent changes the physical-chemical parameters of the reaction system by adjusting pH and using buffer solutions to optimize transaminase activity. Additionally, the patent employs coupled enzymes (amino acid dehydrogenase and racemase) that change the chemical state of the by-product α-keto acid, converting it into D-amino acids that can serve as additional amino donors, thereby further biasing the equilibrium toward product formation.
2Productivity
If by-product removal methods are used to bias reaction equilibrium, then yield improves, but the reaction system becomes more complex with additional enzymes and coenzymes
Solution Approach 1:
The patent employs a simple extraction method using organic solvents to remove the by-product α-keto acid from the reaction system. This physical separation method avoids the need for additional complex enzymatic systems while effectively biasing the reaction equilibrium. The extraction process is straightforward and does not require coupling multiple enzymes or expensive coenzymes like NADH, thus maintaining system simplicity while improving yield.
3Productivity
If expensive coenzymes like NADH are used to convert by-products, then reaction equilibrium is biased toward product formation, but production costs increase
Solution Approach 1:
The patent uses extraction with organic solvents to remove the by-product α-keto acid, eliminating the need for expensive coenzymes like NADH that would otherwise be required to convert the by-product. This approach achieves equilibrium bias through simple physical separation rather than costly biochemical conversion, significantly reducing production costs while maintaining high yield.
Solution Approach 2:
The patent replaces expensive coenzymes with inexpensive buffer solutions and simple organic solvents for by-product removal. The buffer solutions and solvents can be used in large quantities without significant cost, making the process economically viable for large-scale production while achieving the same effect of biasing reaction equilibrium.
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
This approach enables the efficient production of optically-active amine compounds in good yield by reducing the concentration of by-products and regenerating coenzymes, thereby improving productivity and reducing production costs.
Implementation Method 1
a transaminase, using an α-amino acid as an amino-group donor, which has an ability to act on the ketone compound to convert the ketone compound into the optically-active amine compound
Implementation Method 2
an α-keto acid reductase, using a reduced β-nicotinamide adenine dinucleotide (NADH) or a reduced β-nicotinamide adenine dinucleotide phosphate (NADPH) as a coenzyme, which has an ability to reduce, to an α-hydroxy acid, an α-keto acid produced from the α-amino acid through action of the transaminase (A)
Implementation Method 3
an enzyme that has an ability to convert, into NADH, an oxidized β-nicotinamide adenine dinucleotide (NAD+) produced from the NADH through action of the α-keto acid reductase (B) or that has an ability to convert, into NADPH, an oxidized β-nicotinamide adenine dinucleotide phosphate (NADP+) produced from the NADPH through action of the α-keto acid reductase (B)
Data Source
AI summary
The present invention relates to a method for producing an optically-active amine compound. The method is characterized by using a transaminase (A), an α-keto acid reductase (B), and an enzyme (C), each having specific properties, in an identical reaction system to convert a ketone compound into a corresponding optically-active amine compound in which a carbon atom with an amino group bonded thereto serves as an asymmetric point. The present invention also relates to a recombinant vector for use in the method. The present invention makes it possible to efficiently produce an optically-active amine compound.