Anti-freezing polypeptide and preparation method thereof

A technology of antifreeze peptide and fish source, applied in the preparation method of peptide, chemical equipment and method, peptide, etc., can solve the problems of destroying cell and tissue structure, losing quality, etc.

Active Publication Date: 2015-03-04
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Repeated rise and fall of temperature causes ice crystals to grow continuously, freeze-thaw and recrystallize, which seriously damages the cell and tissue structure and loses the quality of the product.

Method used

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  • Anti-freezing polypeptide and preparation method thereof
  • Anti-freezing polypeptide and preparation method thereof
  • Anti-freezing polypeptide and preparation method thereof

Examples

Experimental program
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Effect test

preparation example Construction

[0027] The antifreeze activity detection system for preparing antifreeze polypeptides of the present invention uses low-temperature freeze-thawed bacteria to detect the growth protection effect of adding antifreeze polypeptides on bacteria after low-temperature freeze-thaw. Inoculate the activated Lactobacillus bulgaricus into the liquid medium at 37°C, and cultivate overnight on a shaker at 130r / min as the seed liquid, inoculate the seed liquid into a new liquid medium at a ratio of 1:100, at 37°C, 130r / min Shaker culture to OD 600 =1.0 or so. Take a number of 1.5mL sterilized centrifuge tubes, add 900μL of the sample to be tested at a concentration of 250μg / mL that has been sterilized by filtration, and dilute the bacterial solution by 10 4 times, pipette 100 μL and add it to the sample to be tested, mix evenly, spread, incubate upside down in an incubator at 37°C for 18 hours, and count the number of colonies. Put the remaining sample-bacteria solution mixture at -20°C fo...

Embodiment 1

[0033] Weigh 1.65 g of fish skin collagen and dissolve it in 6 ml of Mili-Q water, then adjust its pH to 7.0 with 2 mol / L NaOH. First heat the solution to 37°C in a water bath, then add a corresponding amount of papain according to the enzyme-substrate ratio of 1:10, and the enzymatic hydrolysis time is 30 minutes. Then inactivate the enzyme in a boiling water bath for 10 minutes, then centrifuge at 14000 rpm for 10 minutes after cooling, and collect the supernatant for later use.

[0034] The supernatant was separated by Sephadex G-50 gel chromatography (length 100cm, diameter 2.6cm), the eluent was deionized water, the flow rate was 2mL / min, and the elution peak was measured at 225nm. Elution peak of antifreeze activity.

[0035] The elution peak with the best antifreeze activity separated by Sephadex G-50 gel chromatography is separated in the next step, separated by Sulfopropyl-Sepadex C-25 cation exchange chromatography (length 55cm, diameter 2.0cm), and eluted The solu...

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Abstract

The invention provides an antifreeze polypeptide and a preparation method thereof. Fish skin collagen is used as a raw material, and a purified specific antifreeze polypeptide is obtained through papain enzymolysis, separation and purification, and the complete sequence of amino acids is: Pro-Gly- Lys-Asn-Gly-Glu-Asp-Gly-Asn-Asn-Gly-Arg-Pro-Gly-Lys-Pro-Gly-Asp-Arg-Gly-Ala-Pro. The present invention breaks through the existing research ideas and methods of antifreeze protein at home and abroad, overcomes the limitation of the quantity of antifreeze protein purified from natural organisms and the concern of the international FDA organization on the safety of transgenic antifreeze protein in food application, and obtains High-efficiency antifreeze peptides based on food sources lay a theoretical foundation for the development of antifreeze peptides based on food sources and their wide application in food and medicine.

Description

technical field [0001] The invention relates to an antifreeze polypeptide, more specifically relates to an antifreeze polypeptide and a preparation method thereof, and belongs to the field of biotechnology. Background technique [0002] The problem that food and pharmaceutical products repeatedly suffer from ice crystal growth and recrystallization due to fluctuations in ambient temperature during cryogenic storage and transportation has attracted increasing attention. The repeated rise and fall of temperature causes ice crystals to grow continuously, freeze-thaw and recrystallize, which seriously damages the cell and tissue structure and loses the quality of the product. Scientists around the world are facing serious challenges: how to control the growth and recrystallization of ice crystals, and realize the control of ice crystal growth on the low-temperature cold chain is the key to restricting the quality of many food and pharmaceutical products. [0003] Antifreeze pro...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/435C12P21/06C07K1/20C07K1/18C07K1/16
CPCC07K14/461
Inventor 汪少芸邵彪王文龙饶平凡
Owner FUZHOU UNIV
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