Fusion-tag-free rhIL-11 and soluble expression and efficient purification method of mutant of fusion-tag-free rhIL-11

Abstract

The invention relates to the technical field of gene recombination, and discloses a fusion-tag-free rhIL-11 and a soluble expression and efficient purification method of a mutant of the fusion-tag-free rhIL-11, and a gene for expressing the fusion-tag-free rhIL-11 is obtained by performing codon escherichia coli preference optimization on human IL-11. The mutant of the rhIL-11 without the fusion tag is derived by performing mutation of a specific amino acid site on the basis of the rhIL-11. The rhIL-11 or the mutant thereof without any fusion tag can be expressed in prokaryotic system escherichia coli cells in a soluble form at high yield, a fusion tag cutting step is not needed in subsequent downstream treatment, inclusion body denaturation and renaturation procedures are not involved, the culture process is simple and extremely low in cost, the expression period of the rhIL-11 or the mutant thereof is short, and the method is suitable for industrial production. The downstream purification step is simple, and the prepared rhIL-11 or the mutant protein thereof has a correct structure and good in-vitro activity.

Description

technical field [0001] The invention relates to the technical field of gene recombination, in particular to a method for soluble expression and high-efficiency purification of rhIL-11 without a fusion tag and a mutant thereof. Background technique [0002] Human interleukin-11 (hIL-11) is one of many important cytokines in the human body. Natural human interleukin-11 is composed of 178 amino acids, which contains a relatively high proportion of proline and leucine, and does not contain cysteine , without glycosylation sites, its theoretical molecular weight is about 19kDa, and its theoretical isoelectric point is 11.16. IL-11 can cooperate with various cytokines in the human body to exert physiological effects, jointly stimulate the growth of precursor cells such as human erythrocytes and megakaryocytes, induce the maturation of megakaryocytes, and promote platelet production. rhIL-11 produced by Genetic Institute (Oprelvekin, ) was approved by the US FDA for the treatmen...

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Problems solved by technology

[0006] Based on the above problems, the present invention aims at the deficiencies of the prior art and the above-mentioned difficult problems, and provides a method for soluble expression and efficient purification of rhIL-11 and its mutants without fusion tags. The present invention does not have any fusion tags. rhIL-11 or its mutants can be expressed in a soluble form in the prokaryotic Escherichia coli at high yields, and the subsequent downstream processing does not require fusion tag cutting steps, nor does it involve inclusion body denaturation and renaturation procedures, and its cultivation process is simple and convenient. The cost is extremely low, the expression cycle of rhIL-11 or its mutants is short, the downstream purification steps are simple, the prepared rhIL-11 or its mutant protein has a correct structure, and its in vitro activity remains good

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