Cyclic biospecific affinity chromatographic method for the purification of the sex steroid binding protein (SBP): Application to the purification of SBP from toad

Abstract

A novel biospecific affinity chromatographic procedure was developed for the purification of the sex steroid binding protein from Bufo arenarum. A charcoal column connected in series to the affinity column allows the removal of any ligand non-covalently bound to the matrix or released during its storage, thus avoiding the need for exhaustive and prolonged washing procedures. In addition, it is not necessary to remove the endogenous ligand from the starting material and the binding to the affinity column can be monitored to determine the time required to achieve the maximum yield. The advantages are the charcoal adsorption of the ligand “washed” from the affinity column by the protein to be purified and the amplification provided by the cyclic use of the system. The procedure improves the yield from less than 1% (by conventional procedures) to more than 50%. With minor modifications this procedure can be useful for the purification of binding proteins and receptors.