Fish-source anti-freezing polypeptide and preparation method thereof

A technology of antifreeze peptide and fish source, applied in the direction of peptide, peptide source, animal/human peptide, etc., can solve the problems of losing quality, destroying cell and tissue structure, etc.

Active Publication Date: 2014-01-01
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 recry

Method used

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  • Fish-source anti-freezing polypeptide and preparation method thereof
  • Fish-source anti-freezing polypeptide and preparation method thereof
  • Fish-source anti-freezing polypeptide and preparation method thereof

Examples

Experimental program
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Effect test

preparation example Construction

[0030] 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

[0036] 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 the corresponding amount of enzyme 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.

[0037] 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.

[0038] 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 so...

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PUM

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Abstract

The invention provides a fish-source anti-freezing polypeptide and a preparation method thereof. Fish skin collagen is used as a raw material which is subject to enzymolysis of papain and then separated and purified to obtain the purified specific anti-freezing polypeptide, and the amino acid complete sequence is Lys-Gly-Glu-Ala-Gly-Asp-Asn-Gly-Ala-Lys-Gly-Asp-Ala-Gly-Ser-Pro-Gly. The invention breaks through the extant research concepts and methods of the anti-freezing protein at home and abroad, avoids limitation on quantity of anti-freezing protein purified from natural living bodies and safety worries about applying the transgenic anti-freezing protein to food from the international FDA (Food and Drug Administration), and acquires the food-source-based efficient anti-freezing polypeptide, thereby laying a theoretical basis for developing the food-source-based anti-freezing polypeptide and exploring the wide application of the food-source-based anti-freezing polypeptide to food and medicines.

Description

technical field [0001] The invention relates to an antifreeze polypeptide, more specifically to a fish-derived antifreeze polypeptide and a preparation method thereof, belonging 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 p...

Claims

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Application Information

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IPC IPC(8): C07K14/46C12P21/06
CPCC07K7/08
Inventor 汪少芸蔡茜茜王文龙饶平凡
Owner FUZHOU UNIV
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