Purification and partial characterization of the normal cellular homologue of the scrapie agent protein

Abstract

The scrapie agent protein (Sp33-37) is a degradation-resistant protein that aggregates into fibrils and amyloid plaques. This protein is derived from a normal cellular protein (Cp33-37). Understanding the mechanism responsible for the conversion of Cp33-37 to Sp33-37 may explain scrapie agent replication. Cp33-37 was extracted from normal hamster brain and purified 2700-fold by an immunoaffinity method. Both Cp33-37 purified from normal hamster brain and Sp33-37 purified from scrapie-affected hamster brain had apparent masses of 33-37 kilodaltons and displayed microheterogeneity characteristic of glycoproteins. Cp33-37 was completely digested by proteinase K under conditions that resulted in conversion of Sp33-37 to the protease-resistant fragment PrP27-30. Cp33-37 did not cause scrapie when inoculated intracerebrally into hamsters. Fractions containing purified Sp33-37 had average titers of greater than 10(11) LD50 of the scrapie agent/mg of protein; these titers were not diminished by proteinase K. These results indicate that altered sensitivity to proteolysis in vitro reflects an intrinsic difference between Sp33-37 and Cp33-37.