Heterogeneity of normal prion protein in two-dimensional immunoblot: presence of various glycosylated and truncated forms

The common use of one-dimensional (1-D) immunoblot with a single monoclonal antibody (Mab) engenders the notion that the normal or cellular prion protein (PrPC ) comprises few and simple forms. In this study we used two-dimensional (2-D) immunoblot with a panel Mabs to various regions of the prion protein to demonstrate the complexity of the PrPC present in human brain. We distinguished over 50 immunoblot spots, each representing a distinct PrPC species based on combinations of different molecular weights and isoelectric points (pIs). The PrPC heterogeneity is due to the presence of a full-length and two major truncated forms as well as to the diversity of the glycans linked to most of these forms. The two major truncated forms result from distinct cleavage sites located at the N-terminus. In addition, enzymatic removal of sialic acid and lectin binding studies indicate that the glycans linked to the full-length and truncated PrPC forms differ in their structure and ratios of the glycoforms. The truncation of PrPC and the heterogeneity of the linked glycans may play a role in regulating PrPC function. Furthermore, the presence of relatively large quantities of different PrPC species may provide additional mechanisms by which the diversity of prion strains could be generated.