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Figure 2
Efficient production of a receptor–ligand complex by co-transfection in HEK293T cells. Cells were transfected with either the receptor construct (sample 1) or the ligand (sample 2, N-glycosylated) or a 1:1 mix of the two plasmids (1 + 2 complex and lane 3 in the inset). Protein levels appear to be improved by co-expression, considering that only half of the normal amount of vector DNA for each partner was used. Size-exclusion chromatography demonstrates complex formation and reveals that a significant amount of ligand (expressed normally at higher levels than the receptor) remains unbound. In the chromatogram, sample 2 appears larger than sample 1 owing to N-glycosylation. For the Western blot analysis (inset) the samples first were deglycosylated by treatment with PNGase; they therefore migrate at the predicted molecular weights. |