A novel PRNP-P105S mutation associated with atypical prion disease and a rare PrPSc conformation.

OBJECTIVE: To define the clinicopathologic, genetic, and pathogenic prion protein (PrP(Sc)) characteristics associated with a novel mutation of the prion protein gene (PRNP). METHODS: The coding segment of PRNP from the proband and family members was sequenced and the brain of the proband was histologically studied. The Western blot profile of the proteinase K (PK) resistant fraction of PrP(Sc), an approximation of its conformation, or "PrP(Sc)-type," was determined. RESULTS: We detected a novel mutation at codon 105 of PRNP that results in a serine (S) substitution of proline (P) (P105S), in a young woman who developed progressive aphasia, behavioral changes, dementia, and parkinsonism, lasting 10 years to her death. Histopathologic findings included an intense focus of multicentric PrP-plaques within the hippocampus, punctate plaques scattered throughout the cerebellum, and intense spongiform degeneration focally within the putamen, suggesting a variant of Gerstmann-Sträussler-Scheinker syndrome (GSS). However, PrP(Sc)-typing revealed two PK-resistant PrP(Sc) fragments (approximately 21 and 26 kDa), a pattern not previously detected in GSS. CONCLUSIONS: This mutation is the third sequence variation at codon 105 of PRNP. The unusual phenotype and PrP(Sc)-type distinguishes this genetic prion disease from typical Gerstmann-Sträussler-Scheinker syndrome and other codon 105 substitutions, suggesting that, in addition to the loss of proline at this position, the PrP(Sc) conformation and phenotype is dependent on the specific amino acid substitution.

High-resolution autoreactive epitope mapping and structural modeling of the 65 kDa form of human glutamic acid decarboxylase.

The smaller isoform of the GABA-synthesizing enzyme, glutamic acid decarboxylase 65 (GAD65), is unusually susceptible to becoming a target of autoimmunity affecting its major sites of expression, GABA-ergic neurons and pancreatic beta-cells. In contrast, a highly homologous isoform, GAD67, is not an autoantigen. We used homolog-scanning mutagenesis to identify GAD65-specific amino acid residues which form autoreactive B-cell epitopes in this molecule. Detailed mapping of 13 conformational epitopes, recognized by human monoclonal antibodies derived from patients, together with two and three-dimensional structure prediction led to a model of the GAD65 dimer. GAD65 has structural similarities to ornithine decarboxylase in the pyridoxal-5'-phosphate-binding middle domain (residues 201-460) and to dialkylglycine decarboxylase in the C-terminal domain (residues 461-585). Six distinct conformational and one linear epitopes cluster on the hydrophilic face of three amphipathic alpha-helices in exons 14-16 in the C-terminal domain. Two of those epitopes also require amino acids in exon 4 in the N-terminal domain. Two distinct epitopes reside entirely in the N-terminal domain. In the middle domain, four distinct conformational epitopes cluster on a charged patch formed by amino acids from three alpha-helices away from the active site, and a fifth epitope resides at the back of the pyridoxal 5'-phosphate binding site and involves amino acid residues in exons 6 and 11-12. The epitopes localize to multiple hydrophilic patches, several of which also harbor DR*0401-restricted T-cell epitopes, and cover most of the surface of the protein. The results reveal a remarkable spectrum of human autoreactivity to GAD65, targeting almost the entire surface, and suggest that native folded GAD65 is the immunogen for autoreactive B-cells.