Characterization of recombinant glycosylated human interleukin 2 produced by a recombinant plasmid transformed CHO cell line.
A recombinant plasmid containing expression units for human pre-interleukin 2 (pre-IL-2) and the selectable marker mouse DHFR, was constructed and used to transform DHFR- CHO cells to the DHFR+ phenotype. Selected colonies were isolated and tested for IL-2 production. Twelve highly IL-2-producing clones were amplified in stepwise increasing concentrations of methotrexate. The IL-2 secreted into the culture medium by one of these clones was purified to homogeneity and partially characterized. N-terminal sequence analysis showed that pre-IL-2 was correctly processed during secretion. SDS gel electrophoresis and chromatofocusing experiments in conjunction with neuraminidase treatment indicated a posttranslational glycosylation of the secreted mature protein similar to that described for the tetrasaccharide structure of the N2 form of natural IL-2. This recombinant IL-2 has a specific activity of 2.5 x 10(7) U/mg.
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D005784:Gene Amplification; D005796:Genes; D006031:Glycosylation; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics* / Q000302:isolation & purification; D010957:Plasmids; D011994:Recombinant Proteins / Q000302:isolation & purification*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014170:Transformation, Genetic
D007376:Interleukin-2; D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase