PURIFICATION OF IMMUNOGENIC HEAT-SHOCK-PROTEIN 70-PEPTIDE COMPLEXES BY ADP-AFFINITY CHROMATOGRAPHY

Adenosine triphosphate (ATP)-affinity chromatography has been widely u sed to purify molecules of the Hsp70 family, This procedure leads to d issociation of peptides from Hsp70 molecules, resulting in Hsp70 prepa rations devoid of immunological activity. We report that substitution of ATP-affinity chromatography by ADP-affinity chromatography results in isolation of Hsp70 molecules which are still associated with peptid es and are immunogenic. The Hsp70 preparations thus obtained contain t he constitutive Hsp73 and the inducible Hsp72 molecules. These observa tions furnish a basis for an analysis of the structural heterogeneity among the members of the Hsp70 family and of the antigenic peptides as sociated with individual members of this family. They also provide a p ractical method for the isolation of large quantities of immunological ly active Hsp70-peptide preparations.