A Conformational Transition at the N Terminus of the Prion Protein Features in Formation of the Scrapie Isoform David Peretz 1 , R. Anthony Williamson 7 , Yoichi Matsunaga 1 Hana Serban 1 , Clemencia Pinilla 8 , Raiza B. Bastidas 7 Roman Rozenshteyn 7 , Thomas L. James 3,4 , Richard A. Houghten 8 Fred E. Cohen 2,5,6 , Stanley B. Prusiner 1,5 * and Dennis R. Burton 7 * 1 Department of Neurology 2 Cellular and Molecular Pharmacology, 3 Pharmaceutical Chemistry, 4 School of Pharmacy 5 Biochemistry and Biophysics and 6 Medicine, University of California, San Francisco CA 94143, USA 7 Departments of Immunology and Molecular Biology The Scripps Research Institute La Jolla, CA 92037, USA 8 Torrey Pines Institute for Molecular Studies, San Diego CA 92121, USA The scrapie prion protein (PrP Sc ) is formed from the cellular isoform (PrP C ) by a post-translational process that involves a profound confor- mational change. Linear epitopes for recombinant antibody Fab frag- ments (Fabs) on PrP C and on the protease-resistant core of PrP Sc , designated PrP 27-30, were identi®ed using ELISA and immunoprecipita- tion. An epitope region at the C terminus was accessible in both PrP C and PrP 27-30; in contrast, epitopes towards the N-terminal region (resi- dues 90 to 120) were accessible in PrP C but largely cryptic in PrP 27-30. Denaturation of PrP 27-30 exposed the epitopes of the N-terminal domain. We argue from our ®ndings that the major conformational change underlying PrP Sc formation occurs within the N-terminal segment of PrP 27-30. # 1997 Academic Press Limited Keywords: prion disease; scrapie; liposome; recombinant antibody; conformational rearrangement *Corresponding authors Introduction Prions cause a group of human and animal neu- rodegenerative diseases that involve profound modi®cation of the cellular prion protein (PrP C ). Spectroscopic measurements demonstrate that the conversion of PrP C into the scrapie isoform (PrP Sc) involves a major conformational transition. PrP C has a high a-helical content, some of which is con- verted into b-sheet when PrP Sc is formed (Pan et al., 1993; Pergami et al., 1996; Safar et al., 1993). The protease-resistant core of PrP Sc , denoted PrP 27-30, also has a high b-sheet content (Caughey et al., 1991; Gasset et al., 1993) but, in contrast to PrP Sc , PrP 27-30 assembles into amyloid polymers (McKinley et al., 1991; Prusiner et al., 1983). Struc- tural studies of the PrP isoforms expressed in mammalian cells and of PrP Sc have proved dif®cult because the level of PrP C expression is low and PrP Sc forms insoluble amorphous aggregates upon puri®cation (McKinley et al., 1991; Scott et al., 1988). Large synthetic PrP peptides and Escherichia coli-derived PrP as well as molecular modeling have provided some information that may prove applicable to the structure of PrP C (James et al., 1997; Mehlhorn et al., 1996; Riek et al., 1996; Zhang et al., 1995). The data reported here provide the ®rst glimpse of the localized conformational changes that PrP C undergoes as it is converted into PrP Sc . One approach to probing conformational rearrangements in prion proteins is to raise anti- bodies to diverse epitopes of PrP C and PrP Sc . There is, however, a lack of any detectable immune response, either humoral or cellular, to infectious Abbreviations used: PrP C , cellular prion protein; PrP Sc , scrapie isoform of prion protein; Fabs, recombinant antibody fragments; PrP 27-30, protease- resistant core of PrP Sc ; SHa, Syrian hamster; mAb, monoclonal antibodies; Mo, mouse; Prnp 0/0 , PrP- de®cient mice; GdnSCN, guanidinium thiocyanate; rPrP, recombinant SHaPrP(90-231); CHO, Chinese hamster ovary. J. Mol. Biol. (1997) 273, 614±622 0022 ± 2836/97/430614±9 $25.00/0/mb971328 # 1997 Academic Press Limited