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Method for improving activity of glutamate decarboxylase by virtue of 732 cation exchange resin

A glutamic acid decarboxylase and cation exchange technology, applied in the biological field, can solve the problem of high cost of γ-aminobutyric acid, and achieve the effects of easy separation and recovery, lower production costs, and no environmental pollution

Active Publication Date: 2018-01-12
LINGNAN NORMAL UNIV
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0029] The object of the present invention is to provide a kind of method that improves Enterococcus faecium glutamic acid decarboxylase conversion activity by 732 cation exchange resins, by using 732 cation exchange resins as auxiliary catalyst, 732 cation exchange resins, containing glutamic acid decarboxylase Enterococcus faecium cells, buffers and reaction substrates L - Glutamate or L -Glutamic acid monosodium salt constitutes a co-catalytic system, and finally achieves the purpose of promoting the conversion activity of glutamate decarboxylase in Enterococcus faecium cells, increasing the production of γ-aminobutyric acid, and solving the problem of high cost of biosynthesis of γ-aminobutyric acid

Method used

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  • Method for improving activity of glutamate decarboxylase by virtue of 732 cation exchange resin
  • Method for improving activity of glutamate decarboxylase by virtue of 732 cation exchange resin
  • Method for improving activity of glutamate decarboxylase by virtue of 732 cation exchange resin

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Embodiment 1

[0064] A method of utilizing 732 cation exchange resin to improve the activity of glutamic acid decarboxylase according to the present invention comprises the following steps:

[0065] ①. L - Monosodium glutamate (monosodium glutamate) was dissolved in 0.2 mol / L acetic acid-sodium acetate buffer, adjusted to pH 4.2 to 4.6, and prepared to 0.2 mol / L to 0.3 mol / L L - Monosodium glutamate solution as substrate solution;

[0066] ②. The 732 cation exchange resin was regenerated or pretreated with 1 mol / L NaOH and 1 mol / L HCl according to the resin instructions, and then fully equilibrated with pH 4.2~pH 4.6, 0.2 mol / L acetic acid-sodium acetate buffer, and then Equilibrate with substrate solution;

[0067] ③. Mix the three in the ratio of 732 cation exchange resin balanced in step ②: volume of substrate solution: volume of Enterococcus faecium GDMCC 60203 bacterial suspension is 1: 1: 1, the mixture is glutamate decarboxylase compound transformation reaction system;

[0068] ④...

Embodiment 2

[0071] A method of utilizing 732 cation exchange resin to improve the activity of glutamic acid decarboxylase according to the present invention comprises the following steps:

[0072] ①. L - Glutamic acid was dissolved in 0.2 mol / L acetic acid-sodium acetate buffer, adjusted to pH 4.2-4.6, and prepared to 0.2 mol / L-0.3 mol / L L - Monosodium glutamate solution as substrate solution;

[0073] ②. The 732 cation exchange resin was regenerated or pretreated with 1 mol / L NaOH and 1 mol / L HCl according to the resin instructions, and then fully equilibrated with pH 4.2~pH 4.6, 0.2 mol / L acetic acid-sodium acetate buffer, and then Equilibrate with substrate solution;

[0074] ③. Mix the three in the ratio of 732 cation exchange resin balanced in step ②: volume of substrate solution: volume of Enterococcus faecium GDMCC 60203 bacterial suspension is 1: 1: 1, the mixture is glutamate decarboxylase compound transformation reaction system;

[0075] ④. React the mixture in step ③ at 80 ...

Embodiment 3

[0078] A method of utilizing 732 cation exchange resin to improve the activity of glutamic acid decarboxylase according to the present invention comprises the following steps:

[0079] ①. L - Monosodium glutamate (monosodium glutamate) was dissolved in 0.2 mol / L acetic acid-sodium acetate buffer, adjusted to pH 4.2 to 4.6, and prepared to 0.2 mol / L to 0.3 mol / L L - Monosodium glutamate solution as substrate solution;

[0080] ②. The 732 cation exchange resin was regenerated or pretreated with 1 mol / L NaOH and 1 mol / L HCl according to the resin instructions, and then fully equilibrated with pH 4.2~pH 4.6, 0.2 mol / L acetic acid-sodium acetate buffer, and then Equilibrate with substrate solution;

[0081] ③. Mix the three 732 cation-exchange resin balance in step ②: volume of substrate solution: volume of Enterococcus faecium GDMCC 60203 glutamate decarboxylase free enzyme solution in a ratio of 1:1:1, the mixture is glutamate Acid decarboxylase complex conversion reaction sys...

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Abstract

The invention relates to a method for improving activity of enterococcus faecium glutamate decarboxylase by virtue of 732 cation exchange resin, and belongs to the field of bio-technology. According to the method provided by the invention, the 732 cation exchange resin is taken as an enzyme activity accelerant of the enterococcus faecium glutamate decarboxylase; a 732 cation exchange resin-glutamate decarboxylase composite catalysis system is constructed by mixing the 732 cation exchange resin, an L-glutamic acid solution and an enterococcus faecium suspension or glutamate decarboxylase free enzyme liquid at the ratio of 1 to 1 to 1 of the mass of the 732 cation exchange resin to the volume of the L-glutamic acid solution to the volume of the enterococcus faecium suspension or glutamate decarboxylase free enzyme liquid; and the yield of [gamma]-aminobutyric acid can be improved by 25.31-143.23% when it conducts a reaction in a water-bath oscillator at 80r / min and 37-43 DEG C or it conducts a stirring reaction at a low speed for 24-36h. According to the method provided by the invention, the 732 cation exchange resin can obviously improve the activity of the glutamate decarboxylase,and meanwhile, an exchange adsorption effect of the 732 cation exchange resin on [gamma]-aminobutyric acid is a process of purifying the [gamma]-aminobutyric acid, so that a downstream extraction andpurification process is simplified and production cost is reduced; and the method is simple and convenient and is green and environment-friendly.

Description

technical field [0001] The invention relates to a method for improving the activity of glutamic acid decarboxylase with 732 cation exchange resin, which belongs to the field of biotechnology. Background technique [0002] γ-Aminobutyric acid (γ-Aminobutyric acid, GABA) is a non-protein amino acid with 4 carbon atoms. It is the main inhibitory neurotransmitter in the mammalian central nervous system. Antioxidant, anti-inflammatory, anti-cancer, blood pressure lowering, sedative and sleep improving effects have become the ingredients of pharmaceuticals and health products that have attracted much attention. GABA is also an important raw material for the production of bioplastic polyamide 4 due to the formation of 2-pyrrolidone by lactamization. [0003] Microbes grow and multiply quickly, and their production of GABA is not limited by time and space. Therefore, the production of GABA by microbial glutamate decarboxylase (glutamate decarboxylase, GAD, EC4.1.1.15) has attracted...

Claims

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Application Information

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IPC IPC(8): C12P13/00C12R1/01
Inventor 杨胜远韦锦
Owner LINGNAN NORMAL UNIV
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