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Purification process of mussel-like mucin

A mussel mucin and process technology, applied in the field of protein purification, can solve the problems of low oxidation rate and slow reaction speed, achieve high purity, simplify the impurity removal method, improve the catalytic reaction speed and the effect of tyrosine oxidation rate

Pending Publication Date: 2022-05-10
CHENGDU ENZPRO BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method has the problems of slow reaction speed and low oxidation rate when modifying tyrosine residues to dopa, and the content of dopa in the recombinant Mgfp-5 protein after tyrosinase oxidation is only 9.6pmol / g

Method used

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  • Purification process of mussel-like mucin
  • Purification process of mussel-like mucin
  • Purification process of mussel-like mucin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Embodiment 1, a kind of purification process of mussel-like mucin

[0059] Purification process steps are as follows:

[0060] 1. Cell destruction: Add 0.2% mass final concentration of Triton X-100 to the fermentation broth, stir evenly, raise the temperature to 85°C under stirring conditions, and maintain it for 15 minutes; use a disc centrifuge at 16000×g to collect the precipitate; Add water of 1 times the volume of the fermentation broth to the precipitate, and add Triton X-114 with a final concentration of 0.2% mass, wash, continue to centrifuge at 16000×g with a disc centrifuge, and collect the precipitate.

[0061] 2. Acid extraction: add 0.3 times the acetic acid of the precipitated mass to the precipitate obtained in step 1, stir for 12 hours; add water to 5 times the original volume, use a disc centrifuge to centrifuge at 16000 × g, and collect the supernatant; The filter membrane (molecular weight cut-off 10KD, made of polyethersulfone) was concentrated to 1...

Embodiment 2

[0082] Embodiment 2, a kind of purification process of mussel-like mucin

[0083] Purification process steps are as follows:

[0084] 1. Cell destruction: add 0.2% mass final concentration of Triton X-100 to the fermentation broth, stir evenly, raise the temperature to 85°C under stirring conditions, and maintain it for 15 minutes; use a disc centrifuge at 16000×g to collect the precipitate ; Add 1 times the volume of fermentation broth water to the precipitate, and add 0.2% mass final concentration of Triton X-114, wash, centrifuge at 16000×g, and collect the precipitate.

[0085] 2. Acid extraction: add 2% citric acid solution to the precipitate obtained in step 1 (the mass ratio of the precipitate to 2% citric acid solution is 1:20), stir for 12 hours; use a disc centrifuge to centrifuge at 16000 × g, collect Supernatant: Concentrate the supernatant to 1 / 10 of the original volume with an ultrafiltration membrane (molecular weight cut-off 3KD, polyethersulfone material), add ...

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Abstract

The invention discloses a purification process of mussel-like mucin, and belongs to the field of protein purification. The purification process comprises the following steps: (1) breaking cells; (2) acid extraction; (3) adding salt to remove impurities, namely adding salt into the concentrated solution obtained in the step (2) until the final concentration is 0.20-0.80 M, centrifuging, collecting supernate, adding salt into the supernate until the final concentration is 1-3 M, centrifuging, and collecting precipitate; (4) enzyme catalytic oxidation; and (5) ion exchange chromatography. The mussel-like mucin with high yield, high purity and high dopa content is obtained by using the purification process provided by the invention. The purification process disclosed by the invention is short in steps and low in cost, and has a wide application prospect in large-scale production of the mussel-like mucin.

Description

technical field [0001] The invention belongs to the field of protein purification, and in particular relates to a purification process of mussel-like mucin. Background technique [0002] Mussels are a kind of crustacean marine organisms widely distributed in coastal and offshore areas. Their silk glands can secrete silk and fix mussels on the surface of various substrates through silk disks formed by silk. The main component of its silk is mussel foot silk protein (Mytilus edulis foot protein, MFP), also known as mussel adhesive protein (mussel adhesive protein, MAP), the protein has high strength, high toughness and water resistance. Mussel mucin also has the characteristics of strong corrosion resistance, good biocompatibility, and no immune reaction. It has broad application prospects in the fields of marine engineering, biomedicine, and surface chemistry. [0003] A large number of studies have shown that the key to mussel mucin's ability to adhere to almost all solid m...

Claims

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Application Information

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IPC IPC(8): C07K14/435C07K1/18C07K1/30C07K1/14
CPCC07K14/43509
Inventor 刘杨昌杨明颜饶峻彭钰洁唐玲莉袁夕茹李端华王辂赵晨
Owner CHENGDU ENZPRO BIOTECHNOLOGY CO LTD