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High-yield arginine deiminase bacterial strain and method for producing L-citrulline by same

A technology of citrulline and bacterial liquid, which is applied in the field of high-yield arginine deiminase strain and its production of L-citrulline, can solve the problem of low specific enzyme activity, achieve high acid production and stable properties , important economic and social benefits

Active Publication Date: 2012-10-03
山东恩贝生物工程有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to solve the defects of existing arginine deiminase production strains and solve the problem of low specific enzyme activity

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The breeding of embodiment 1 high production arginine deiminase bacterial strain BM-2

[0022] (1) Preparation of the starting strain ATCC 10180 bacterial suspension:

[0023] One ring of Streptococcus faecalis (streptococcus faecalis) ATCC10180 strain preserved on a slant was picked with a sterile inoculation needle, and streak culture was carried out on a solid medium. After activation, pick a ring of fresh bacteria and dilute it with sterile saline, and the concentration of the bacteria solution is diluted to 10 7 cells / ml to obtain a bacterial suspension.

[0024] Solid medium and seed medium (the latter without agar) consisted of the following components by weight: 10 parts of casein peptone, 20 parts of beef extract, 20 parts of yeast extract, 20 parts of glucose, 10 parts of sodium acetate, diammonium citrate 5 parts, Tween 802 parts, dipotassium hydrogen phosphate 2 parts, magnesium sulfate heptahydrate 0.1 part, manganese sulfate monohydrate 0.05 part, agar 2...

Embodiment 2

[0043] Embodiment 2L-citrulline production method

[0044] Seed medium composition (parts by weight): 7.5 parts of casein peptone, 10 parts of beef extract, 5 parts of yeast extract, 10 parts of glucose, 5 parts of sodium acetate, 2 parts of diammonium citrate, 801 parts of Tween, dipotassium hydrogen phosphate 4 parts, 0.3 parts of magnesium sulfate heptahydrate, 0.03 parts of manganese sulfate monohydrate and 1000 parts of water;

[0045] Composition of fermentation medium (parts by weight): 5 parts of glucose, 2 parts of corn steep liquor, 10 parts of peptone, 30 parts of beef extract, 20 parts of arginine, 5 parts of potassium dihydrogen phosphate, 0.5 part of magnesium sulfate heptahydrate, sodium chloride 3 parts, 0.03 parts of manganese sulfate monohydrate and 1000 parts of water.

[0046] (1) Take a glycerol tube, place it at room temperature until it melts, draw 1ml of the bacterial solution and inoculate it into a first-grade seed shaker flask, and incubate at 30°C ...

Embodiment 3

[0050] Seed medium composition (parts by weight): 20 parts of casein peptone, 5 parts of beef extract, 10 parts of yeast extract, 20 parts of glucose, 10 parts of sodium acetate, 10 parts of diammonium citrate, 10 parts of Tween 8010 parts, dipotassium hydrogen phosphate 10 parts, 0.5 parts of magnesium sulfate heptahydrate, 0.05 parts of manganese sulfate monohydrate and 1000 parts of water;

[0051] Composition of fermentation medium (parts by weight): 20 parts of glucose, 10 parts of corn steep liquor, 20 parts of peptone, 20 parts of beef extract, 30 parts of arginine, 10 parts of potassium dihydrogen phosphate, 0.8 part of magnesium sulfate heptahydrate, sodium chloride 10 parts, 0.02 parts of manganese sulfate monohydrate and 1000 parts of water.

[0052] (1) Take a glycerol tube, place it at room temperature until it melts, inoculate 0.5ml of bacterial solution into a first-grade seed shake flask, and incubate at 37°C for 24 hours;

[0053] (2) inoculate into secondary...

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Abstract

The invention discloses a high-yield arginine deiminase bacterial strain and a method for producing L-citrulline by the same. The high-yield arginine deiminase bacterial strain of the invention is streptococcus faecalis BM-2 CGMCC No.4990. After small-scale fermentation of the bacterial strain in a 30-L fermentation tank, the L-citrulline yield is up to 98 g / L, which is increased by 50% when compared with the L-citrulline yield of the original strain; after continuous fermentation for 5 batches, the L-citrulline yield of the bacterial strain is stable, and the L-citrulline yield level is obviously higher than that of reported bacterial strains; and the bacterial strain of the invention has important industrial production application value.

Description

technical field [0001] The invention relates to a high-yield arginine deiminase strain and a method for producing L-citrulline with it, belonging to the field of biotechnology. Background technique [0002] Arginine deiminase (ADI) is a protein that can decompose L-arginine. Scholars first purified ADI from the extract of Mycoplasma arginini, and found that the enzyme can inhibit the proliferation of animal vascular endothelial cells. Related studies have further confirmed that arginine deiminase can not only inhibit the proliferation of various malignant tumor cells in vitro, It can also inhibit the proliferation of malignant tumors in the body. As a new anti-tumor substance, arginine deiminase has been paid more and more attention by researchers. [0003] According to reports, arginine deiminase can catalyze the conversion of L-arginine into L-citrulline. This enzymatic reaction is only one step, which can effectively avoid the complex feedback regulation in the total sy...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P13/10C12R1/46
Inventor 郑雄敏王震刘倩
Owner 山东恩贝生物工程有限公司
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