NR AIBP
AU Matsunaga,Y.; Peretz,D.; Williamson,A.; Burton,D.R.; Mehlhorn,I.; Groth,D.; Cohen,F.E.; Prusiner,S.B.; Baldwin,M.A.
TI Cryptic epitopes in N-terminally truncated prion protein are exposed in the full-length molecule: dependence of conformation on pH.
QU Proteins 2001 Aug 1; 44(2): 110-8
PT journal article
AB Prion diseases are diseases of protein conformation. Structure-dependent antibodies have been sought to probe conformations of the prion protein (PrP) resulting from environmental changes, such as differences in pH. Despite the absence of such antibodies for full-length PrP, a recombinant Fab (D13) and a Fab derived from mAb 3F4 showed pH-dependent reactivity toward epitopes within the N-terminus of N-terminally truncated PrP(90-231). Refolding and maintaining this protein at pH > or =5.2 before immobilization on an ELISA plate inhibited reactivity relative to protein exposed to pH < or =4.7. The reactivity was not affected by pH changes after immobilization, showing retention of conformation after binding to the plate surface, although guanidine hydrochloride at 1.5-2 M was able to expose the cryptic epitopes after immobilization at pH > or =5.2. The alpha-helical CD spectrum of PrP(90-231) refolded at pH 5.5 was reduced somewhat by these pH changes, with a minor shift toward beta-sheet at pH 4 and then toward coil at pH 2. No covalent changes were caused by the pH differences. This pH dependence suggests titration of an acidic region that might inhibit the N-terminal epitopes. A similar pH dependence for a monoclonal antibody reactive to the central region identified an acidic region incorporating Glu152 as a significant participant.
MH Amino Acid Sequence; Animal; Antigen-Antibody Reactions; Circular Dichroism; Enzyme-Linked Immunosorbent Assay; Epitopes/*chemistry/*genetics/immunology; Guanidine/chemistry; Hamsters; Hydrogen-Ion Concentration; Immunoglobulins, Fab/chemistry/metabolism; Mesocricetus; Mice; Mice, Knockout; Molecular Sequence Data; Peptide Fragments/*chemistry/*genetics/immunology; Prions/*chemistry/*genetics/immunology; Protein Conformation; Protein Denaturation; Recombinant Proteins/chemistry/metabolism; Spectrometry, Mass, Electrospray Ionization; Support, U.S. Gov't, P.H.S.
AD Institute for Neurodegenerative Diseases, University of California, San Francisco, California 94143-0446, USA
SP englisch
PO USA