Copper Has Differential Effect on Prion Protein with Polymorphism of Position 129 Boon-Seng Wong,* ,1 Christine Clive,† Stephen J. Haswell,† Ian M. Jones,* ,2 and David R. Brown‡ ,2 *NERC Institute of Virology and Environmental Microbiology, Mansfield Road, Oxford, OX1 3SR, United Kingdom; †Department of Chemistry, Hull University, Hull HU6 7RX, United Kingdom; and ‡Department of Biochemistry, Cambridge University, Tennis Court Road, Cambridge CB2 1QW, United Kingdom Received February 3, 2000 The pathology of human prion diseases is affected by polymorphism at amino acid residue 129 of the prion protein gene. Recombinant mouse prion pro- teins mimicking either form of the polymorphism were prepared to examine their effect on the confor- mation and the level of superoxide dismutase (SOD) activity of the prion protein. Following the binding of copper atoms to prion protein, antibody mapping and CD analysis detected conformational differences be- tween the two forms of protein. However, neither the level of copper binding nor the level of SOD activity associated with this form of prion protein altered with the identity of codon 129. These results suggest that in the holo-metal binding form of the protein, prion structure but not its SOD activity is affected by poly- morphism at codon 129. © 2000 Academic Press Key Words: prion; copper refolding; methionine; va- line; antibody binding; codon 129; superoxide dismutase. Prions are hypothesised to cause a group of human and animal neurodegenerative diseases, now classified together because of their aetiology and pathogenesis (for reviews see (1, 2)). The human diseases include Fatal Familial insomnia (FFI), Gerstmann-Straussler- Scheinker (GSS), and Creutzfeldt-Jakob disease (CJD). Prion proteins (PrPs) are believed to become infectious following a change in the conformation of the normal cell-surface glycoprotein, prion protein (PrP C ), to a pro- tease resistant form (PrP Sc ). Human prion diseases are unique because they can arise by inheritance as well as by infection (3) and, about 10% of the cases of CJD and all cases of GSS and FFI are linked to germ line mu- tations in the PrP gene. The mutations have been postulated to favour the conversion of PrP C into the infectious conformer (4) and the effects of various PrP mutations have been studied in transgenic mice, cell cultures, and protein models (5–7). In humans there is a polymorphism at codon 129 which, may be methionine or valine, can profoundly influences the phenotypic characteristics of the dis- eases caused by pathogenic mutations at other posi- tions (3). Residue 129 lies on one of the two short -sheet region located on the C-terminal domain of the mouse PrP (8). All FFI subjects had methionine at position 129 while all CJD subjects had valine at that position, in association with Asp178Asn mutation (9). Similarly, valine-129 in association with Phe198Ser mutation predisposed humans to the Indiana kindred variant of GSS (10). Thus, the interaction between specific mutations and codon 129 may control the phe- notypic presentation of familial human prion diseases. Recently, homozygousity for methionine at codon 129 has been suggested to predispose individual to a non- neuronal disease, sporadic inclusion-body myositis (sIBM) (11, 12). This disease presents a high level of prion mRNA expression in muscle tissues and the pathogenic effects could be the results of oxidative stress (13). Indeed, transgenic mice overexpressing wild-type PrP has shown a profound necrotizing myop- athy involving skeletal muscle (14). Recent evidence has shown that PrP C is a copper- binding protein (15–17), and defects in copper ho- meostasis are known to cause several diseases of the central nervous system (18). We recently demonstrated that when full-length prion protein binds copper it acquired superoxide dismutase (SOD) activity as a con- sequence (17). These observations are consistent with previous studies demonstrating reduced superoxide dismutase activity in neurones from Prnp 0/0 mice (15). PrP expression protects neurones from cell death due to oxidative stress (19). When recombinant PrP binds copper the acquisition of SOD activity was also accom- panied by a selective oxidation of methionine residues 1 Present address: Biomedical Research Building (9 th Floor), De- partment of Pathology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4943. 2 To whom correspondence may be addressed. Ian M. Jones at Fax: 44-1865-281635; or David R. Brown at Fax: 44-1223-333345, E-mail: drb33@cam.ac.uk. Biochemical and Biophysical Research Communications 269, 726 –731 (2000) doi:10.1006/bbrc.2000.2355, available online at http://www.idealibrary.com on 726 0006-291X/00 $35.00 Copyright © 2000 by Academic Press All rights of reproduction in any form reserved.