Abstract
Bovine spongiform encephalopathy (BSE) is a prion disease of cattle that is caused by the misfolding of the cellular prion protein (PrPC) into an infectious conformation (PrPSc). PrPC is a predominantly α-helical membrane protein that misfolds into a β-sheet rich, infectious state, which has a high propensity to self-assemble into amyloid fibrils. Three strains of BSE prions can cause prion disease in cattle, including classical BSE (C-type) and two atypical strains, named L-type and H-type BSE. To date, there is no detailed information available about the structure of any of the infectious BSE prion strains. In this study, we purified L-type BSE prions from transgenic mouse brains and investigated their biochemical and ultrastructural characteristics using electron microscopy, image processing, and immunogold labeling techniques. By using phosphotungstate anions (PTA) to precipitate PrPSc combined with sucrose gradient centrifugation, a high yield of proteinase K-resistant BSE amyloid fibrils was obtained. A morphological examination using electron microscopy, two-dimensional class averages, and three-dimensional reconstructions revealed two structural classes of L-type BSE amyloid fibrils; fibrils that consisted of two protofilaments with a central gap and an average width of 22.5 nm and one-protofilament fibrils that were 10.6 nm wide. The one-protofilament fibrils were found to be more abundant compared to the thicker two-protofilament fibrils. Both fibrillar assemblies were successfully decorated with monoclonal antibodies against N- and C-terminal epitopes of PrP using immunogold-labeling techniques, confirming the presence of polypeptides that span residues 100-110 to 227-237. The fact that the one-protofilament fibrils contain both N- and C-terminal PrP epitopes constrains molecular models for the structure of the infectious conformer in favour of a compact four-rung β-solenoid fold.
Highlights
Prion diseases, called transmissible spongiform encephalopathies (TSE), belong to a group of zoonotic, fatal neurodegenerative diseases that cause spongiform changes in the brain [1]
Bovine spongiform encephalopathy (BSE) prions exist in three variants or strains: C-type BSE prions, which caused the epizootic “mad cow disease” outbreak, and two atypical forms L-type and Htype BSE prions, named according to their migration patterns during gel electrophoresis
Our study revealed that L-type BSE prions assemble into oneand two-protofilament containing amyloid fibrils and that the width of the two-protofilament fibrils is approximately twice that of one-protofilament fibrils
Summary
Called transmissible spongiform encephalopathies (TSE), belong to a group of zoonotic, fatal neurodegenerative diseases that cause spongiform changes in the brain [1]. Bovine spongiform encephalopathy (BSE), known as ‘mad cow disease’, was first detected in 1986 in the U.K. BSE is the only zoonotic prion disease with the confirmed capability of transmission to humans, resulting in the occurrence of variant Creutzfeldt-Jakob disease, an acquired form of prion disease in humans. Three BSE strains have been reported to cause prion disease in cattle, including classical BSE (C-type) and two atypical forms termed L-type and H-type BSE, which differ in their neuropathological and molecular phenotypes [7,8,9]. The disease caused by L-type BSE is called bovine amyloidotic spongiform encephalopathy (BASE), due to the atypical deposition of amyloid plaques in the brain [8,10]
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