Mapping the prion protein using recombinant antibodies.
The fundamental event in prion disease is thought to be the posttranslational conversion of the cellular prion protein (PrPC) into a pathogenic isoform (PrPSc). The occurrence of PrPC on the cell surface and PrPSc in amyloid plaques in situ or in aggregates following purification complicates the study of the molecular events that underlie the disease process. Monoclonal antibodies are highly sensitive probes of protein conformation which can be used under these conditions. Here, we report the rescue of a diverse panel of 19 PrP-specific recombinant monoclonal antibodies from phage display libraries prepared from PrP deficient (Prnp0/0) mice immunized with infectious prions either in the form of rods or PrP 27-30 dispersed into liposomes. The antibodies recognize a number of distinct linear and discontinuous epitopes that are presented to a varying degree on different PrP preparations. The epitope reactivity of the recombinant PrP(90-231) molecule was almost indistinguishable from that of PrPC on the cell surface, validating the importance of detailed structural studies on the recombinant molecule. Only one epitope region at the C terminus of PrP was well presented on both PrPC and PrPSc, while epitopes associated with most of the antibodies in the panel were present on PrPC but absent from PrPSc.
Departments of Immunology, The Scripps Research Institute, La Jolla, California 92037, USA.
R A
RA
Williamson
D
D
Peretz
C
C
Pinilla
H
H
Ball
R B
RB
Bastidas
R
R
Rozenshteyn
R A
RA
Houghten
S B
SB
Prusiner
D R
DR
Burton
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D018604:Epitope Mapping / Q000379:methods*; D000939:Epitopes / Q000737:chemistry / Q000235:genetics; D007114:Immunization; D051379:Mice; D008969:Molecular Sequence Data; D018621:PrPC Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D018620:PrPSc Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D011328:Prions / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D018621:PrPC Proteins; D018620:PrPSc Proteins; D011328:Prions; D011994:Recombinant Proteins