Identification of an epitope in the C terminus of normal prion protein whose expression is modulated by binding events in the N terminus

We have characterized the epitopes of a panel of 12 monoclonal antibodies ( Mabs) directed to normal human cellular prion protein (PrPC) using ELISA an d Western blotting of recombinant PrP or synthetic peptide fragments of PrP . The first group of antibodies, which is represented by Mabs 5B2 and 8B4, reacts with PrP23-145, indicating that the epitopes for these Mabs are loca ted in the 23 to 145 N-terminal region of human PrP. The second group inclu des Mabs 1A1, 6H3, 7A9, 8C6, 8H4, 9H7 and 2G8. These antibodies bind to epi topes localized within N-terminally truncated recombinant PrP90-231. Finall y, Mabs 5C3, 2C9 and 7A12 recognize both PrP23-145 and PrP90-231, suggestin g that the epitopes for this group are located in the region encompassing r esidues 90 to 145. By Western blotting with PepSpot(TM) only three of Mabs studied (5B2, 8B4 and 2G8) bind to linear epitopes that are present in 13-r esidue long synthetic peptides corresponding to human PrP fragments. The re maining nine Mabs appear to recognize conformational epitopes. Two N termin us-specific Mabs were found to prevent the binding of the C terminus-specif ic Mab 6H3. This observation suggests that the unstructured N-terminal regi on may influence the local conformation within the folded C-terminal domain of prion protein. (C) 2000 Academic Press.