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 d enatured PA-GFP migrated at 57 kDa which corresponds to the theoretica l molecular mass, Although the reason(s) for this mobility shift betwe en fluorescent and non-fluorescent molecules remains unclear, the smal l 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.