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Method for preparing medicinal recombinant human interleukin-11

A technology for interleukin and fusion protein, which is applied in the field of preparation of pharmaceutical recombinant human interleukin-11, can solve the problems of uncontrollable degree of glycosylation, poor process stability, high process difficulty, etc., and achieves safety and effectiveness Sexual guarantee, quality controllable, good repeatability

Inactive Publication Date: 2010-11-24
XIAMEN AMOYTOP BIOTECH
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Problems solved by technology

However, as far as human IL-11 has no intramolecular / intermolecular disulfide bonds and no potential glycosylation sites, glycosylation will occur when expressed in yeast, and the degree of glycosylation is not easy to control. At the same time, compared with the large intestine Bacillus expression system, yeast expression system has a long fermentation period, difficult process, high cost, and poor process stability; expressed in Escherichia coli, the expression level is high, which is conducive to purification

Method used

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  • Method for preparing medicinal recombinant human interleukin-11
  • Method for preparing medicinal recombinant human interleukin-11
  • Method for preparing medicinal recombinant human interleukin-11

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Experimental program
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Embodiment 1

[0031] 1) Preliminary purification of rhIL-11

[0032] a. Bacterial cell disruption: The bacterial cells were resuspended with 50 mM Tris+500 mM NaCl, pH 8.0 buffer at a ratio of 1:10 (w / v). Ultrasonic crushing can be used for a small amount, and high-pressure crushing can be used for a large amount. The centrifuged supernatants of the cells suspended in the disrupted solution were compared using 100-900 Psi pressure. The result shows: 200Psi has higher yield and higher purity.

[0033] b. Separation by chelating column: According to the characteristics of MB residues in the fusion protein, Chelating Sepharose FF was selected as the filler for preliminary purification. Use 100mM imidazole to wash away impurity proteins, and the elution condition of the target protein is 50mM Tris-Cl, 285-325mM imidazole, 1% Tween 80 gradient elution, the purification results show that this step of purification can make the target fusion protein The purity is as high as more than 90%, which ...

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Abstract

The invention discloses a method for preparing medicinal recombinant human interleukin-11, which adopts the following preparation steps of: 1) initial purification of a fusion protein in the interleukin-11, which comprises steps of crushing thalli and separating a chelate column; 2) digesting the fusion protein containing the interleukin-11 by enterokinase; and 3) fine purification of the interleukin-11, wherein Macro Cap SP column chromatography and Superdex-75 column chromatography are carried out to obtain the medicinal recombinant human interleukin-11. The method has the characteristics of simple process, high expression amount, high stability, uniform product, low production cost, environmental protection and high activity, improves the safety, controllability and validity of medicaments and is suitable for industrial production.

Description

technical field [0001] The invention relates to a preparation method of medicinal recombinant human interleukin-11. Belongs to the field of biotechnology. Background technique [0002] Human interleukin-11 (IL-11) is a cytokine that exists in various tissues such as human bone marrow, nerves, respiratory tract and digestive tract epithelium, and has a wide range of physiological functions, especially its effect on the hematopoietic system. have important clinical value. Currently, recombinant human IL-11 ( Genetics Institute) was officially approved by the FDA in November 1997. Its indication is chemotherapy-induced thrombocytopenia, and it has become the only drug of choice for the treatment of chemotherapy-induced thrombocytopenia. [0003] IL-11 was only discovered in the late 1980s. It was found that the culture medium derived from the bone marrow stromal cell line PU-34 could stimulate the growth of the T1165 cell line neutralized by excess IL-6 antibody. In 1990, ...

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Application Information

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IPC IPC(8): C12P21/06C07K1/16C07K1/14
Inventor 孙黎郑成已郑建华王世媛
Owner XIAMEN AMOYTOP BIOTECH
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