Strain capable of producing D-mannose isomerase and method for producing D-mannose isomerase by using same

A technology of mannose isomerase and mannose, applied in the field of food biology, can solve the problems of severe reaction conditions, high cost of plant enrichment method, poor safety and the like, and achieve the effect of efficient production

Inactive Publication Date: 2015-07-15
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In contrast, the chemical synthesis method has a long history and rapid reaction, but the raw materials used are expensive, highly toxic, the reaction conditions are severe, and the safety is poor. The solvent used is highly corrosive and has potential safety hazards, so it is not suitable for the food industry.
The plant enrichment method has high cost, low yield and great limitations

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 Isolation of producing mannose isomerase bacteria

[0031] Use naturally rotten peaches or apples, take a little pulp and add to the separation medium (in g / L): ammonium chloride 1~12 g / L, disodium hydrogen phosphate 1~6 g / L, potassium dihydrogen phosphate 0.1~1 g / L, magnesium chloride 0.05~0.1 g / L, sodium sulfate 0.1~1 g / L, D-fructose 1~20 g / L, prepared with deionized water; adjust pH to 7.0; cultivate at 30°C, 200 rpm After 14-24 h, after dilution and coating, the pure culture was obtained, cultured at 30°C for 30 h, and 15 strains with high enzyme activity were obtained, and a strain with high enzyme activity was selected to isolate the D-mannose The constitutive enzyme-producing bacteria were sent to Sangon Bioengineering (Shanghai) Co., Ltd. to determine the morphological characteristics, physiological and biochemical characteristics of the bacteria, and 16s RNA gene sequence analysis. The results were identified as Pseudomonas ( Pseudomonas sp. ) SK 27....

Embodiment 2

[0032] The preparation of embodiment 2 crude enzyme powder

[0033] The resulting Pseudomonas SK27.016, in the fermentation medium (in g / L): ammonium chloride 2 g / L, disodium hydrogen phosphate 6 g / L, potassium dihydrogen phosphate 0.5 g / L, Magnesium chloride 0.05 g / L, sodium sulfate 0.1 g / L, D-fructose 10 g / L, prepared with deionized water; adjust pH to 7.0; culture at 30°C, 200 rpm for 24 h, refrigerated and centrifuged to obtain fermented bacteria The body is the crude enzyme solution. After concentrating the crude enzyme solution, add ammonium sulfate to the concentrated enzyme solution to a saturation of 40% to precipitate to remove most of the impurity proteins, then increase the saturation of ammonium sulfate to 80%, and collect the precipitate The protein is freeze-dried to obtain the crude enzyme powder.

Embodiment 3

[0034] Embodiment 3 enzyme reaction

[0035] Use the crude enzyme powder of embodiment 2, add with 0.5 U / mL to the 50 mM phosphate buffer saline (Na 2 HPO 4 -NaH 2 PO 4 ), the enzymatic reaction was carried out at 40°C for 5 h, and 150 g / L D-mannose was converted into D-mannose.

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Abstract

The invention provides a strain capable of producing D-mannose isomerase and a method for producing D-mannose isomerase by using the same, belonging to the food biotechnical field. The invention provides a pseudomonas sp. SK27.016 screened from rotted juicy peach and collected at China Center for Type Culture Collection with a collection number of CCTCC NO.M2014411 and further provides a method for producing D-mannose isomerase through fermentation by using the strain. The method comprises the following steps: (a) fermenting the strain in a fermentation culture medium to obtain the thalli of pseudomonas sp. SK27.016; (b) converting D-fructose by using intracellular D-mannose isomerase produced by pseudomonas sp. SK27.016 to produce D-mannose; and (c) separating and purifying D-mannose. According to the method, D-mannose isomerase produced by the strain is used for efficiently producing D-mannose from D-fructose; therefore, the method is suitable for mass production and a new method is provided for enzyme-method industrial preparation of D-mannose.

Description

technical field [0001] The present invention relates to a kind of Pseudomonas ( Pseudomonas sp. The invention relates to the screening of SK 27.016 and the method for converting D-fructose to produce D-mannose by using the strain to ferment cells, belonging to the field of food biotechnology. Background technique [0002] D-Mannose, the only glyconutrient currently available clinically, is widely distributed in body fluids and tissues, especially in nerves, skin, testes, retina, liver and intestines. It is directly used to synthesize glycoproteins and participate in immune regulation. In addition, in recent years, it has been found that D-mannose can inhibit tumor growth and metastasis, increase the survival rate of cancer, and it has incomparable advantages in the treatment of urinary tract infection. D-Mannose, may be recommended as a supplemental treatment for infants and children with a rare disorder called phosphorylated mannose isomerase deficiency, which is known as...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P19/24C12R1/38
CPCC12P19/02C12P19/24C12N1/205C12R2001/38
Inventor 江波张涛胡兴沐万孟缪铭
Owner JIANGNAN UNIV
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