Construction of a fusion protein between protein A and green fluorescent protein and its application to Western blotting

Aequorea green fluorescent protein (GFP) and protein A were fused and expressed in Escherichia coli, The fluorescent native fusion protein (PA-GFP) migrated at 47 kDa in SDS-PAGE, However, the non-fluorescent denatured PA-GFP migrated at 57 kDa which corresponds to the theoretical molecular mass, Although the reason(s) for this mobility shift between fluorescent and non-fluorescent molecules remains unclear, the small ring structure within the native molecules may affect their mobility. The cell extract, prepared from an E. coli strain producing PA-GFP, was used in Western and dot blots. The sensitivity and specificity of the PA-GFP detection mere sufficient for rapid and easy screening.