Purification of the 90 kDa heat shock protein (hsp90) and simultaneous purification of hsp70/hsc70, hsp90 and hsp96 from mammalian tissues and cells using thiophilic interaction chromatography.

Heat shock proteins (HSPs) hsp70/hsc70, hsp90 and hsp96 were separated from mammalian cells and tissues on a gel obtained by the reaction of beta-mercaptoethanol with divinyl sulfone-activated Sepharose CL-6B (thiophilic gel or T-gel). Hsp90 revealed a much higher affinity towards the T-gel than the other HSPs. One-step thiophilic interaction chromatography of proteins resulted in a more than 80% purity and 85% yield of hsp90. Based on this observation, a simple and efficient method for the purification of hsp90 and a procedure for the simultaneous purification of several HSPs (hsp70/hsc70, hsp90 and hsp96) using thiophilic interaction chromatography was developed. All the HSPs were recovered with a high yield and purity (90-99%). The results indicated that the thiophilic gel is a highly efficient affinity matrix for the purification of hsp90 and can be used in the protocols of purification of different HSPs from cells and tissues of various animal species.