Production of ß-Galactosidase Fused Protein A and the Purification of it in an Aqueous Two-Phase System
We have studied the production of the genetically fused protein Staphylococcal protein A/Escherichia coli ß-galactosidase (SpA-ßgal). The cultivation conditions were found to have a profound effect on the overall productivity of SpA-ßgal. Biologically active SpA-ßgal was produced corresponding to 12% of the cell dry weight. The purification of SpA-ßgal was based on the partitioning properties of ß-galactosidase during extraction in an aqueous two-phase system consisting of poly(ethylene glycol) 4000 and potassium phosphate. Stability studies of SpA-ßgal revealed that the ß-galactosidase moiety remained intact both with respect to size and enzyme activity, while the SpA moiety was completely degraded by E. coli proteases.
KeywordsPotassium Phosphate Cell Debris Phase System Bottom Phase Binodial Curve
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