Objective: To express recombinant human interleukin-11 (rhIL-11) in methylotropic yeast Pichia pastoris.
Methods: By designing and synthesizing an artificial gene for IL-11, the expression vector pPICZ alpha-A-IL-11 was constructed and introduced into Pichia pastoris by linearized electroporation. The rhIL-11 protein was identified by ELISA and SDS-PAGE analysis. The bioactivity was analyzed by B9-11 cell line. A combination of liquid chromatography was developed to purify the rhIL-11 from ferment supernatant.
Results: The nucleotide sequence analysis indicated that the sequence of cloned artificial IL-11 gene accorded with that of designed; the secreted yield of rhIL-11 by yeast Pichia pastoris KM71-2424 in flask reached 60 mg/L. The biological activity of IL-11 in yeast supernatant and E. coli standard determined by B9-11 was 5.5 x 10(7) U/mg and 2.2 x 10(7) U/mg respectively. The rhIL-11 was purified to electrophoretic purity by a combination of liquid chromatography.
Conclusion: The human IL-11 artificial gene was obtained and successfully expressed in the Pichia pastoris(KM71-2424). The biological activity of IL-11 in yeast supernatant was significantly higher than that of E. coli standard. The rhIL-11 was purified to electrophoretic purity.