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Method for purifying recombinant human follistatin protein

A follistatin and purification method technology is applied in the field of efficient purification of a new genetic engineering drug-recombinant human follistatin protein, and achieves the effects of low production cost, high product yield and simple production process

Inactive Publication Date: 2017-03-15
德清知诺同丰生物科技有限公司
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  • Summary
  • Abstract
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Problems solved by technology

[0004] Using genetic engineering fermentation method to express recombinant human follistatin protein, the expression level is relatively high, but due to the high hydrophobicity of the protein and rich disulfide bonds, the protein mostly exists in the form of inclusion bodies

Method used

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Embodiment Construction

[0022] The high-efficiency purification process of the recombinant human follistatin of the present invention includes the following steps: bacterial cell crushing → inclusion body washing and dissolving → rough purification of the target protein → fine purification of the target protein.

[0023] The specific process is as follows:

[0024] 1. Bacterial fragmentation:

[0025] 1.1 Take out the frozen bacteria from the low-temperature refrigerator, break them into small pieces of about 2 cubic centimeters, put them into a large beaker, add 20mM PB solution with a volume 20 times the weight of the bacteria, and add lysate with a final concentration of 25mg / L Enzymes, stir with a mechanical stirrer for 1 hour.

[0026] 1.2 Under the condition of ice bath, break up the bacteria in batches with an ultrasonic breaker to ensure that the broken liquid is no longer viscous.

[0027] 1.3 Use a high-capacity refrigerated high-speed centrifuge at 7000rpm / min, centrifuge at 4°C for 20 m...

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Abstract

The invention relates to an efficient method for purifying a new genetic engineering medicine, namely recombinant human follistatin protein. The method for purifying the recombinant human follistatin protein comprises the following steps: (1) performing ultrasonic pyrolysis on thallus, washing an inclusion body, and performing hydrotropy with 6M guanidine hydrochloride; (2) performing nickel column affinity chromatography by using a 6His label; and (3) performing DEAE Sepharose FF anion exchange chromatography, thereby obtaining the high-purity recombinant human follistatin protein. According to the method, the purified protein yield is high and the protein purity is high, and the method has the characteristics of simple steps, economy and practicability and convenience in large-scale production and purification.

Description

technical field [0001] The invention relates to a high-efficiency purification method of a genetic engineering new drug-recombinant human follistatin protein. Background technique [0002] Follistatin is a cysteine-rich glycosylated single-chain polypeptide isolated from bovine and pig follicular fluid by Robertson and Ueno in 1987, which has a strong inhibitory effect on follicle stimulating hormone. effect. Follistatin is widely distributed, and follistatin can be detected in different tissues. As the binding protein of Activin, Follistatin can regulate the reproductive activity of animals through the Activin / Follistatin system. Experiments in recent years have shown that follistatin can participate in the regulation of various physiological functions of the body through its action system, and play an important role in the regulation of ovarian granulosa cell differentiation, embryonic development, erythropoiesis and osteoblast function. biological significance. [000...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/47C07K1/22C07K1/18
CPCC07K14/4703
Inventor 雷振松王勇波
Owner 德清知诺同丰生物科技有限公司
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