7β-HSDH Enzyme Stereospecific Reduction for UDCA Synthesis
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Solution Overview
Problem
Current methods for producing ursodeoxycholic acid (UDCA) face challenges such as incomplete conversion due to reaction equilibria and the need for multiple enzymes, increasing process costs, and the difficulty in removing cofactor coproducts, which affects the efficiency and selectivity of the synthesis process.
Innovation Solution
Isolation and characterization of a novel regio- and stereospecific 7β-hydroxysteroid dehydrogenase (7β-HSDH) from Collinsella aerofaciens, which enables the stereospecific reduction of dehydrocholic acid (DHCA) to 3,12-diketo-7β-cholanic acid, allowing for the production of UDCA in three steps with high conversion and selectivity using recombinantly produced 7β-HSDH and 3α-HSDH enzymes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If chemical oxidation method is used for UDCA production, then conversion can be achieved, but selectivity is poor requiring protection steps
Solution Approach 1:
The patent replaces chemical oxidation methods with enzymatic oxidation using 7β-HSDH from Collinsella aerofaciens. This biological catalyst provides high stereoselectivity for oxidizing the 7β-hydroxy group while leaving other functional groups untouched, eliminating the need for protection steps required in chemical methods.
2Productivity
If multiple enzymes are used for UDCA synthesis, then conversion can be achieved, but process cost increases
Solution Approach 1:
The patent discovers that 7β-HSDH from Collinsella aerofaciens can perform multiple oxidation steps in the UDCA synthesis pathway. This single enzyme can oxidize both the 7β-hydroxy group and the 12α-hydroxy group, replacing what previously required multiple different enzymes, thereby simplifying the process and reducing costs.
3Productivity
If cofactor regeneration is used, then reaction can proceed, but coproduct removal is difficult affecting equilibrium
Solution Approach 1:
The patent employs a two-phase system where the coproduct (NAD+) is extracted into an organic phase, separating it from the aqueous reaction medium. This continuous removal of coproduct shifts the reaction equilibrium forward according to Le Chatelier's principle, maintaining high conversion without coproduct accumulation interfering with the reaction.
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 novel method enables high-yield, selective production of UDCA with reduced byproduct formation, facilitating large-scale production and efficient cofactor regeneration, thereby overcoming the limitations of existing methods.
Implementation Method 1
Isolation and characterization of a novel regio- and stereosspecific 7β-hydroxysteroid dehydrogenase (7β-HSDH) from Collinsella aerofaciens, which enables the stereospecific reduction of dehydrocholic acid (DHCA) to 3,12-diketo-7β-cholanic acid
Implementation Method 2
efficient cofactor regeneration, thereby overcoming the limitations of existing methods
Data Source
AI summary
The invention relates to novel 7β-hydroxysteroid dehydrogenases which are obtainable from bacteria of the genus Collinsella, especially of the strain Collinsella aerofaciens, to the sequences encoding said enzymes, to methods for producing said enzymes and to their use in the enzymatic conversion of cholic acid compounds, and especially in the production of ursodeoxycholic acid (UDCS). The invention also relates to novel methods for the synthesis UDCS.