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Strain capable of highly producing aspartate decarboxylase and method for producing alanine

An aspartic acid and decarboxylase technology, applied in the biological field, can solve problems such as low enzyme activity, achieve high enzyme activity, be beneficial to extraction and separation operations, and broaden the channels of strains.

Active Publication Date: 2016-03-02
山东中酶生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The above reports are limited by the low enzyme activity, and there is still a long way to go before industrial application

Method used

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  • Strain capable of highly producing aspartate decarboxylase and method for producing alanine
  • Strain capable of highly producing aspartate decarboxylase and method for producing alanine
  • Strain capable of highly producing aspartate decarboxylase and method for producing alanine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1: Classification of Bacillus tequila strains

[0019] 1) Characteristics of strains

[0020] exist preservation The colonies on the culture medium are milky white, nearly round, dry on the surface, opaque, wrinkled, dull, with irregular lobes at the edges, easy to pick; observed through a microscope, the bacteria are rod-shaped, single and not in chains, Gram Positive, after 48 hours of culture, it was observed that the spores were attached to the side of the bacterium or existed freely, without swelling, oval or columnar.

[0021] 2) Strain identification

[0022] The length of the 16SrDNA gene sequence obtained by DNA extraction, PCR amplification and sequencing is 1459bp, and the BLAST comparison on GenBank shows that the strain has a base similarity of 99% to Bacillus tequilensis (JX412373), and the sequence information as follows:

[0023]tggctttgcgcttgctatacatgcaagtcgagcggacagatgggagcttgctccctgatgttagcggcggacgggtgagtaacacgtgggtaacctgcctgtaagactgggataa...

Embodiment 2

[0027] 1) Strain activation: pick the strain to solid preservation Medium: yeast extract powder 5g / L, peptone 10g / L, NaCl 5g / L, agar 20g / L, pH 7.0, inoculate in a constant temperature incubator at 30°C for 20h;

[0028] 2) Preparation of liquid seeds: Pick 1 ring of activated strains and inoculate them in a 500ml Erlenmeyer flask containing 50ml of seed medium. The components of liquid seed medium: yeast extract powder 5g / L, peptone 10g / L, NaCl 5g / L, pH7 .0, sealed with eight layers of gauze, and cultivated at 30°C and 200r / min shaking table for 18h to obtain liquid seeds;

[0029] 3) liquid fermentation: according to 5% inoculum size, the components are glucose 20g / L, peptone 20g / L, aspartic acid 2g / L, MgSO 4 0.2g / L, KH 2 PO 4 0.5g / L, ammonium sulfate 3g / L fermentation medium was inserted into liquid seeds, the rotation speed was 200-700r / min, the dissolved oxygen was controlled at 20-30%, pH 7.0, temperature 30°C, aerated for 24 hours, and L-days were obtained. Fermentat...

Embodiment 3

[0034] 1) Strain activation: pick the strain to solid preservation Medium: yeast extract powder 5g / L, peptone 10g / L, NaCl 5g / L, agar 20g / L, pH 7.0, inoculate in a constant temperature incubator at 30°C for 20h;

[0035] 2) Preparation of liquid seeds: Pick 1 ring of activated strains and inoculate them in a 500ml Erlenmeyer flask containing 50ml of seed medium. The components of liquid seed medium: yeast extract powder 5g / L, peptone 10g / L, NaCl 5g / L, pH7 .0. Eight layers of gauze were sealed, and cultured at 30°C and shaken at 200r / min for 18 hours to obtain liquid seeds;

[0036] 3) Liquid fermentation: according to 7% inoculum size, the components are glucose 20g / L, peptone 20g / L, aspartic acid 2g / L, MgSO 4 0.2g / L, KH 2 PO 4 0.5g / L, ammonium sulfate 3g / L fermentation medium was inserted into liquid seeds, the rotation speed was 200-700r / min, dissolved oxygen was controlled at 25-30%, pH 7.0, temperature 30°C, aerated for 28 hours, and L-days were obtained. Fermentation ...

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Abstract

The invention relates to a strain capable of highly producing aspartate decarboxylase and a method for producing alanine and belongs to the technical field of biology. The class name of the strain is bacillus tequilensis PanD37 and the strain is preserved in the China General Microbiological Culture Collection Center with the preservation number of CGMCC No.10506 on February 2, 2015. The bacillus tequilensis PanD37 capable of producing L-aspartate-alpha-decarboxylase, provided by the invention, is reported for the first time, the strain PanD37 is good in genetic stability, and low in culture cost, the produced L-aspartate-alpha-decarboxylase is high in enzyme activity, the strain fermentation liquor can be directly converted into beta-alanine, and a surfactant, an organic solvent and other membrane permeable agents are not required to be added during catalytic synthesis, so that exogenous impurities are prevented from being introduced, the extraction separation operation is facilitated and an advantage in industrial application is realized.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a bacterial strain producing L-aspartic acid α-decarboxylase, and using the fermentation liquid of L-aspartic acid α-decarboxylase produced by the bacterial strain to catalyze the preparation of β-alanine Methods. Background technique [0002] β-alanine is the only β-type non-protein amino acid that exists in nature. It is used in the synthesis of pharmaceutical products such as carnosine, calcium pantothenate, balsalazide and sodium pamidronate, and has a wide range of uses. At present, β-alanine mainly adopts chemical synthesis method, which has disadvantages such as high cost, unfriendly conditions and production environment. The development of green and safe biological production process will have very obvious economic and social benefits. L-aspartate α-decarboxylase (L-aspartate α-decarboxylase, EC4.1.1.11, PanD) is a key enzyme in the biosynthetic pathway of pantothenic acid, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/88C12P13/06C12R1/07
CPCC12N9/88C12P13/06C12Y401/01011C12N1/205C12R2001/07
Inventor 冯志彬陈国忠张娟
Owner 山东中酶生物科技有限公司
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