Method for efficiently producing pro-TGase through fermentation

A technology for producing transglutaminase enzymes and strains is applied in the field of high-efficiency fermentation and production of transglutaminase proenzymes, which can solve problems such as low secretion of transglutaminase proenzymes, and achieve good application prospects. Effect

Inactive Publication Date: 2012-07-11
JIANGNAN UNIV
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Problems solved by technology

[0004] The technical problem to be solved in the present invention is to provide a method for efficient fermentation and production of transglutaminase zymogen to solve the technical problem of low secretion of transglutaminase zymogen

Method used

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  • Method for efficiently producing pro-TGase through fermentation
  • Method for efficiently producing pro-TGase through fermentation
  • Method for efficiently producing pro-TGase through fermentation

Examples

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Embodiment 1

[0017] The logarithmic growth phase controls the temperature at 37°C, when the bacteria grow to OD 600 At 4, 6, 8, and 10 o'clock respectively, the temperature was lowered to 20°C and IPTG with a final concentration of 0.4mM was added to promote the high-efficiency expression of pro-TGase. Until the end of fermentation, the induced OD 600 =8, the highest enzyme activity was 6.22U / mL, and the production intensity was 0.16U / mL / h ( figure 1 ).

Embodiment 2

[0019] Change the initial glycerol concentration to 4g / L, 8g / L and 12g / L respectively, at OD 600 = Induction at 8 hours. Culture condition is the same as example 1, until fermentation finishes ( figure 2 ). When the initial glycerol concentration was 8g / L, the highest enzyme activity was 9.92U / mL, and the production intensity was 021U / mL / h, which was 31.25% higher than the previous production intensity ( image 3 ).

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Abstract

The invention discloses a method for producing pro-TGase by adding isopropyl thiogalactoside (IPTG) and inducing to improve the fermentation of recombinant Escherichia Coli BL21(DE3) through a low temperature strategy. A secretary expression vector pET22b recombinant Escherichia Coli BL21(DE3) is taken as a fermentation strain, and the temperature is controlled to be 37DEG C in an initial growth stage; when thalli grow to reach a certain thallus concentration, cool induction is carried out, the IPTG at the final concentration of 0.4mM is added, the temperature is reduced to 20DEG C, and the expression of the pro-TGase is induced. The temperature of an induction stage is reduced in the fermentation process, and the aim of improving the yield of the pro-TGase is finally fulfilled. The method has the advantages that: the yield and the productivity of the pro-TGase can be effectively improved, the content of other proteins in a fermentation liquor is reduced, the efficiency during downstream extraction is improved, the product purity is improved, energy consumption is reduced, and the method is suitable for industrial large-scale production.

Description

technical field [0001] The present invention relates to a method for high-efficiency fermentative production of pro-TGase (transglutaminase) pro-TGase (transglutaminase) fermentatively produced by recombinant Escherichia Coli BL21 (DE3) using an inducer low-temperature control strategy. zymogen) method. Background technique [0002] Transglutaminase EC 2.3.2.13 full name R-glutaminyl-peptide: amine-γ-glutayle-transferase referred to as TGase), also known as transglutaminase or glutaminyl-peptide γ-glutaminyl Transferase, with a variety of catalytic functions. It can catalyze the formation of ε-(γ-glutamyl) lysine covalent bonds between or within the protein molecules, causing intramolecular and intermolecular cross-linking of proteins, connections between proteins and amino acids, and glutamine in protein molecules. Hydrolysis of the amide group. The unique catalytic function of TGase makes it play an important role in many fields. At present, the enzyme is widely used i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/10C12R1/19
Inventor 陈坚马建龙刘松陈康康张娟张东旭堵国成
Owner JIANGNAN UNIV
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