Copper(II)-induced conformational changes and protease resistance in recombinant and cellular PrP. Effect of protein age and deamidation.
While PrP(C) rearranges in the area of codons 104-113 to form PrP(Sc) during prion infections, the events that initiate sporadic Creutzfeldt-Jakob disease are undefined. As Cu(II) is a putative ligand for PrP(C) and has been implicated in the pathogenesis of Creutzfeldt-Jakob disease and other neurodegenerative diseases, we investigated the structural effects of binding. Incubation of brain microsomes with Cu(II) generated approximately 30-kDa proteinase K-resistant PrP. Cu(II) had little effect on fresh recombinant PrP23-231, but aged protein characterized by conversion of Asn-107 to Asp decreased alpha-helical content by approximately 30%, increased beta-sheet content 100%, formed aggregates, and acquired proteinase K resistance in the presence of Cu(II). These transitions took place without need for acid pH, organic solvents, denaturants, or reducing agents. Since conversion of Asn to Asp proceeds by a spontaneous pathway involving deamidation, our data suggest that covalent variants of PrP(C) arising in this manner may, in concert with Cu(II), generate PrP(Sc)-like species capable of initiating sporadic prion disease.
Qin
K
K
Centre for Research in Neurodegenerative Diseases, Department of Medical Biophysics, Mass Spectrometry Laboratory, Modern Medicine Research Centre, Toronto, Ontario M 5S 3HS, Canada.
Yang
D S
DS
Yang
Y
Y
Chishti
M A
MA
Meng
L J
LJ
Kretzschmar
H A
HA
Yip
C M
CM
Fraser
P E
PE
Westaway
D
D
D000577:Amides / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D003300:Copper / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry; D019286:Endopeptidase K / Q000378:metabolism*; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D011328:Prions / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000577:Amides; D004220:Disulfides; D011328:Prions; D011994:Recombinant Proteins; D003300:Copper; D019286:Endopeptidase K