Method for forming supramolecular hydrogel for biological catalysis by utilizing enzyme self-assembly
A self-assembly method and porous hydrogel technology, applied in biochemical equipment and methods, biofuels, enzymes, etc., can solve problems such as poor stability and easy falling off, large mass transfer resistance, small immobilization amount, etc., and achieve immobilization steps Simple, good organic solvent tolerance, enzyme activity and stability improvement effect
Pending Publication Date: 2022-06-03
TSINGHUA UNIV
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Problems solved by technology
Traditional immobilization methods include carrier-based physical adsorption, covalent grafting, affinity, cross-linking, etc. If the immobilization force is too strong, the activity will be greatly lost, and if the force is too weak, the stability will be poor and easy to fall off. Therefore, the activity and There is a contradiction in stability
In addition, some methods immobilize enzymes through embedding and encapsulation, such as MOF-encapsulated enzymes, although the stability is high, the immobilization amount is small and there is a large mass transfer resistance
Method used
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Embodiment 1
[0049] Example general method:
[0050] 1. Add alcohol dehydrogenase (ADH) and nitrogen-containing organic ligands to a beaker with a mass ratio of 1:50. 9 mL of phosphate buffer was added to bring the enzyme concentration to 0.6 mg / mL. The temperature is controlled at 25°C. Stir for 10 minutes.
[0051] 2. After stirring, add 1 mL of a metal ion solution (the metal ion is copper ion or cobalt ion or magnesium ion or calcium ion) into the beaker, and the concentration of the metal ion in the metal ion solution is 13 mg / mL. The temperature is controlled at 25°C. Stirring was continued for 2 h.
[0052] 3. The solution is collected, centrifuged and washed three times, and then freeze-dried to obtain a porous hydrogel product.
[0053] 4. Take the synthetic hydrogel catalyzing reaction containing 0.5mg enzyme, the reaction solution includes phosphate buffer, the reaction substrate is acetophenone, the concentration of acetophenone is 5mmol / L, and the concentration of coenzym...
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Abstract
The invention discloses a method for forming supramolecular hydrogel through enzyme self-assembly for biological catalysis, and relates to a method for immobilizing ethanol dehydrogenase through an enzyme self-assembly method, application of the ethanol dehydrogenase and a method for synthesizing R-chiral alcohol. The method for immobilizing the ethanol dehydrogenase by using the enzyme self-assembly method comprises the following steps: mixing the ethanol dehydrogenase, a nitrogen-containing organic ligand and a pH buffer solution to obtain a pre-gelatinized initial solution; mixing the initial solution and the metal ion solution, and continuously carrying out gelation reaction to obtain a gel solution; and centrifuging, washing and freeze-drying the gel solution to obtain the porous hydrogel containing the ethanol dehydrogenase. The immobilization method is simple in step, a carrier is not needed, chemical modification and additional additives are not needed, and compared with free enzyme, the hydrogel obtained through enzyme self-assembly has better organic solvent tolerance, heat stability and storage stability and can be used for preparing the hydrogel. The catalyst is used for catalyzing asymmetric reduction of aryl ketone to synthesize R-chiral alcohol, and has very high yield and chiral selectivity.
Description
technical field [0001] The invention belongs to the technical field of biocatalysis, and relates to a method for forming a supramolecular hydrogel by using enzyme self-assembly for biocatalysis, and specifically includes a method for immobilizing alcohol dehydrogenase by using the enzyme self-assembly method and its application, and the synthesis of R-hand alcohol method. Background technique [0002] Biocatalysis has the characteristics of high selectivity, high efficiency, green and mildness, and is a powerful driving force for the progress of human material production. Among them, biocatalyst is the core. Enzymes are widely used biocatalysts, and their separation, purification and reuse are the main obstacles restricting industrial applications. In order to solve this problem, the enzyme immobilization technology was born, that is, the water-soluble enzyme is treated by physical or chemical methods to make it insoluble in water, but still has enzyme activity. Tradition...
Claims
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IPC IPC(8): C12N9/04C12N11/02C12P7/02
CPCC12N9/0006C12Y101/01001C12N11/02C12P7/02Y02E50/10
Inventor 王玉军陈强骆广生
Owner TSINGHUA UNIV
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