For the preparation of the complex of IGF-II and IGFBP-6, a co-expression vector containing two copies of human IGF-II and IGFBP-6 expression cassette was constructed with alcohol oxidase (AOX1) promoter and secretion signal sequence of %-factor, and transformed to Pichia pastoris yeast. Through a purification procedure involving anionexchange chromatography and gel filtration, a complex of IGF-II with IGFBP-6 was obtained. An additional C-terminal sequence of IGFBP-6 (CS-BP6) was found to be bound to this complex. Dynamic light scattering showed that this complex was very stable and homogenous in solution. Western blotting based on non-reducing Tricine-SDS-PAGE indicated that IGF-II expression coupled with IGFBP-6 might significantly avoid the mispairing of disulfide bonds compared with the IGF-II expressed alone.
Keywords: Pichia pastoris, insulin-like growth factor II, insulin-like growth factor binding protein-6, co-expression, complex
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