Prion Protein-Detergent Micelle Interactions Studied by NMR in Solution

Abstract

Transmissible spongiform encephalopathies (TSEs), 2 such as Creutzfeldt-Jakob disease and the Gerstmann-Sträussler-Scheinker syndrome in humans, are accompanied by the appearance in the brain of an aggregated "scrapie" isoform of the host-encoded prion protein, PrP Sc (1-3). The cellular form, PrP C , consists of an unstructured N-terminal "tail" of residues 23-125 and a globular domain of residues 126 -231, and is attached by a C-terminal glycosylphosphatidylinositol (GPI) anchor to the outer plasma membrane. This structure ensures a role of membrane interactions in the physiological function of PrP C and probably also in the disease-related events leading to TSEs. For example, transgenic mice expressing a prion protein variant lacking the GPI membrane anchor did not develop the typical clinical signs of TSE after inoculation with infectious brain homogenate, although significant amounts of PrP Sc accumulated in the brain (4). This finding led to the conclusion that membrane-association of PrP C is necessary for the development of a TSE. Independent evidence for the importance of membrane interactions for the onset of prion diseases was derived from cell-free conversion assays and cell culture experiments (5, 6).Data have also been presented that indicate that in addition to the normal form with the C terminus linked to a GPI anchor and the C-terminal domain located on the cell surface, PrP C can adopt two different transmembrane topologies, Ctm PrP and Ntm PrP, which have the C-terminal polypeptide segment located in the lumen of the endoplasmic reticulum ( Ctm PrP) or in the cytoplasm ( Ntm PrP) (7-9). The population of the Ctm PrP variant is Ͻ10% of the total wild-type prion protein present during cellular biosynthesis but is increased to 20 -30% for the pathogenic mutations P102L, P105L, and A117V of human PrP and the designed variant mouse PrPs obtained with the amino acid exchanges A113V/A115V/A118V and K110I/H111I (10 -13). The population of Ctm PrP was further increased when an additional mutation, L9R, was present in the N-terminal signal sequence (14), so that ϳ50% of the PrP was synthesized as the Ctm PrP variant in granule neurons obtained from transgenic mice expressing a prion protein construct carrying the four amino acid replacements L9R, A113V, A115V, and A118V (15). Quite generally, an increase in the population of Ctm PrP was also shown to be associated with severe neurodegeneration in transgenic mice, and it has been suggested that Ctm PrP may be the proximate cause of neuronal death in certain prion disorders (10,11,15).In vitro studies on interactions of full-length and N-terminally truncated forms of recombinant PrP showed that acidic membranes caused the N-terminal part of the protein to become more structured, whereas the C-terminal domain was destabilized (16 -19). Furthermore, zwitterionic gel-phase dipalmitoylphosphatidylcholine or raft-like membranes were shown to induce increased ␣-helical structure in recombinant Syrian hamster PrP-(90 -231) at pH 7.0 (18, 19). Membrane interactio...

Copper Alters Aggregation Behavior of Prion Protein and Induces Novel Interactions between Its N- and C-terminal Regions

“…Our NMR studies indicate previously unknown interactions of the N-terminal region with the C-terminal region in the Cu 2ϩ -bound PrP . The NMR of full-length PrP has been reported earlier; there are also reports of NMR studies on Cu 2ϩ -binding to fragments of PrP (28,32,59). To the best of our knowledge, this is the first investigation of NMR of Cu 2ϩ -bound full-length PrP, which shows novel long range inter-domain interactions.…”

“…Conformational studies on binding of Cu 2ϩ to PrP in vitro have been performed using peptides (26 -28) or truncated variants of PrP (29 -31) and at nonphysiological temperatures (2,10,(13)(14)(15)(16)(17) and in water or buffers of low ionic strength (3,9,13,(32)(33)(34). We have used full-length PrP to investigate conformational changes and aggregation upon Cu 2ϩ -binding at physiological temperature (37°C).…”

Interactions between the Conserved Hydrophobic Region of the Prion Protein and Dodecylphosphocholine Micelles

“…Several studies using circular dichroism and NMR (15,16,17,18) have taken a close look at these interactions in structural terms, but an atomic level (high resolution) description is not yet available. From these reports, Hornemann et al (18) have studied the interactions between dodecylphosphocholine (DPC) and the mouse prion protein (mPrP(90 -231)) and disease-related mutants. The data showed little or no interaction between the wild type and the detergent whereas the mutants showed weak interactions.…”

Residues within a lipid-associated segment of the PECAM-1 cytoplasmic domain are susceptible to inducible, sequential phosphorylation

“…Upfield 15 N chemical shift changes previously have been correlated with a micelle-induced conformational change from random coil to ␣-helix on interaction. 25,26 To identify the residues involved in DPC binding, we assigned backbone 15 …”