Vector system pSA for high-level expression of recombinant glycoprotein in eukaryotic cells

Abstract

The invention provides a vector system pSA for high-level expression of a recombinant glycoprotein in eukaryotic cells, in particular an expression vector for expressing a foreign protein in animal cells. The vector system pSA comprises an expression cassette for expressing the foreign protein. The expression cassette is provided with the following elements sequentially from 5' to 3': (a) a first promoter, (b) a multiple cloning site and an optional coding sequence of the foreign protein, (c) a first polyA sequence; (d) a second promoter, (e) a selectable marker gene and (f) a second polyA signal sequence. The expression vector of the invention is particularly suitable to efficiently express a human activated protein C.

Description

technical field [0001] The invention relates to the field of genetic engineering. More specifically, the present invention relates to a vector system pSA for high-level expression of recombinant glycoproteins in eukaryotic cells. The present invention also provides an expression vector particularly suitable for highly expressing human activated protein C in animal cells. Background technique [0002] At present, a variety of protein expression systems have been developed to produce exogenous proteins of organisms, such as Escherichia coli expression systems, yeast expression systems, higher eukaryotic cell expression systems, etc., but these systems still have some defects that need to be improved . How to apply these new expression systems to efficiently express the desired target gene is also a problem that needs to be explored and studied urgently in this field. [0003] For example, Escherichia coli can express a variety of proteins as a host, but its existence (1) la...

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Problems solved by technology

[0003] For example, Escherichia coli can express a variety of proteins as a host, but its existence (1) lacks post-translational modification and processing of eukaryotic proteins, such as cleavage, glycosylation, formation of disulfide bonds, etc.; (2) the expressed protein Insoluble inclusion bodies are often formed, and complex renaturation is required to restore conformation and activity; (3) Bacterial proteins are not conducive to purification and other disadvantages

[0004] The Saccharomyces cerevisiae system also has limitations: (1) the yield is usually low; (2) the expression plasmid is easy to lose; (3) lacks a strong and tightly regulated promoter; (4) excessive glycosylation modification of foreign proteins It will seriously change the immunogenicity of the protein, reduce the activity, and shorten the residence time; (5) The secretion efficiency is poor and the purification is difficult

[0005] In summary, the current expression of human glycosylated proteins in animal cells has technical difficulties such as low expression, high production costs, or low activity when expressed in bacteria

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