Novel Differences between Two Human Prion Strains Revealed by Two-dimensional Gel Electrophoresis

Tao Pan,Monica Colucci,Boon-Seng Wong,Ruliang Li,Tong Liu,Robert B Petersen,Shu Chen,Pierluigi Gambetti,Man-Sun Sy

doi:10.1074/jbc.m107358200

Abstract

The phenotype of human sporadic prion diseases is affected by patient genotype at codon 129 of the prion protein (PrP) gene, the site of a common methionine/valine polymorphism, and by the type of the scrapie PrP (PrP(Sc)), which likely reflects the prion strain. However, two distinct disease phenotypes, identified as sporadic Creutzfeldt-Jakob disease (M/M2 sCJD) and sporadic fatal insomnia (sFI), share methionine homozygosity at codon 129 and PrP(Sc) type 2. One-dimensional gel electrophoresis and immunoblotting reveal no difference between the M/M2 sCJD and sFI species of PrP(Sc) in gel mobility and glycoform ratio. In contrast, the two-dimensional immunoblot demonstrates that in M/M2 sCJD the full-length PrP(Sc) form is overrepresented and carries glycans that are different from those present in the PrP(Sc) of sFI. Because the altered glycans are detectable only in the PrP(Sc) and not in the normal or cellular PrP (PrP(C)), they are likely to result from preferential conversion to PrP(Sc) of rare PrP(C) glycoforms. This is the first evidence that a qualitative difference in glycans contributes to prion diversity.

Highlights

The normal or cellular prion protein (PrPC)1 in humans is a cell-surface, glycosylphosphatidylinositol-anchored glycoprotein that has 209 residues and two potential N-glycosylation sites [1,2,3]
On one-dimensional immunoblots, all untreated PrP preparations from control, M/M2 subtypes of Creutzfeldt-Jakob disease (sCJD), and sporadic fatal insomnia (sFI) brains display the typical pattern of human brain PrP, control brains contain only PrPC whereas PrP from M/M2 sCJD and sFI comprises both PrPC and PrPSc, referred to as total PrP
Analysis of PrPSc by one-dimensional gel electrophoresis and immunoblotting has shown that PrPSc associated with distinct animal and human prion diseases may differ in gel mobility and in the ratio of the glycoforms, providing the first insight into the molecular basis of prion diversity (4 – 6, 12, 13)

Summary

The abbreviations used are

PrPC, cellular (normal) prion protein; PAGE, polyacrylamide gel electrophoresis; PK, proteinase K; PrPSc, scrapie PrP; sCJD, sporadic Creutzfeldt-Jakob disease; sFI, sporadic form of fatal insomnia; M/M, Met homozygous at codon 129; M/M2, Met homozygous at codon 129 and having PrPSc type 2; PNGase F, peptideN-glycosidase F; mAb, monoclonal antibody. Prion strains are thought to reflect different conformations that PrPSc may adopt [5, 12, 13] Both the genotype of the host and the prion strains determine the disease phenotype [2, 3]. The mechanism of the phenotypic diversity in sporadic prion diseases is unclear but is believed to be determined by the genotype at codon 129, the site of a common Met/Val polymorphism [17, 18] and the PrPSc type [18]. We demonstrate for the first time major differences between the PrPSc associated with sFI and that present in M/M2 sCJD.

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION