Assessment of the different conformational states of the abnormal
prion protein (PrP(Sc)) in the CNS provides an established basis for distinguishing
transmissible spongiform encephalopathy (TSE) strains. PrP(Sc) conformers are variably resistant to N-terminal
proteinase K (PK) digestion, and analysis of the consensus products (
PrP(res)) by immunoassay enables effective, but relatively low-resolution differentiation. Determination of the precise N-terminal
amino acid profile (N-TAAP) of
PrP(res) presents a potential high-resolution means of TSE-strain typing, and thus of differential disease diagnosis. This approach was evaluated using individual mice affected by model
scrapie (22A, ME7, 87V and 79A) and
bovine spongiform encephalopathy (BSE) (301V) strains. Nano liquid chromatography-mass spectrometry (LC-MS) was used to determine
PrP(res) N-terminal tryptic digestion products. Four major N-terminal tryptic
peptides were generated from all mouse TSE strains investigated, corresponding with predominant N-termination of
PrP(res) at G(81), G(85), G(89) and G(91). Both the mass spectrometric abundance of the individual
peptides and the ratios of pairs of these
peptides were evaluated as markers of conformation in relation to their potential for strain discrimination. The yield of
peptides was significantly greater for BSE than
scrapie strains and the relative quantities of particular
peptide pairs differed between strains. Thus, whereas
peptide G(91)-K(105) was a dominant
peptide from 301V, this was not the case for other strains and, significantly, the ratio of
peptides G(91)-
K(105):G(89)-
K(105) was substantially higher for BSE-infected compared with
scrapie-infected mice. These data support the potential of the N-TAAP approach for high-resolution TSE strain typing and differential diagnosis.