METHODS TO OBTAIN RECOMBINANT PROTEINS WITH INCREASED SIALYLATION FROM CELLS THAT EXPRESS ADENOVIRUS E1A PROTEIN, AND PROTEINS OBTAINED THEREBY

摘要

It includes that glycan (EPO) is connected to this that the present invention, which is provided comprising one or more isotype erythropoietin(EPO), it is characterised in that the glycan includes LewisX structures and each EPO molecules of an average of at least 6 sialic acid moiety. Present invention provides obtain to include the structure of each EPO molecules of an average of at least 6 sialic acids and Lewis x from 0 to 2 comprising glycan (EPO) wherein described glycan linked with it comprising one or more isotype erythropoietin(EPO), the described method includes: providing a kind of expressible format containing nucleic acid sequence encoding eukaryocyte adenovirus E 1 A albumen and further containing the expressible formats of EPO in core, wherein the unit further includes nucleic acid sequence encoding sialyltransferase, preferably α 2, a kind of 6-sialyltransferase or α -2, under the control of 3-sialyltransferase, in allogeneic promoter; The culture cell in serum-free culture medium and allows the EPO expressed in the cell in step b); The EPO of step c) harvest expression is from the cell and/or from culture medium; D) fraction EPO is purified and is fractionated to obtain these increased average sialic acid content N connection each EPO molecules of glycan, a kind of common mprise linked comprising the EPO of glycan with it comprising one or more equal formation are obtained, wherein the structure of each EPO molecules of an average of at least 6 sialic acids of glycan and Lewis x from 0 to 2.