Methionine oxidation within the prion protein

Abstract

Prion diseases are characterized by the self-templated misfolding of the cellular prion protein (PrP^C) into infectious aggregates (PrP^Sc). The detailed molecular basis of the misfolding and aggregation of PrP^C remains incompletely understood. It is believed that the transient misfolding of PrP^C into partially structured intermediates precedes the formation of insoluble protein aggregates and is a critical component of the prion misfolding pathway. A number of environmental factors have been shown to induce the destabilization of PrP^C and promote its initial misfolding. Recently, oxidative stress and reactive oxygen species (ROS) have emerged as one possible mechanism by which the destabilization of PrP^C can be induced under physiological conditions. Methionine residues are uniquely vulnerable to oxidation by ROS and the formation of methionine sulfoxides leads to the misfolding and subsequent aggregation of PrP^C. Here, we provide a review of the evidence for the oxidation of methionine residues in PrP^C and its potential role in the formation of pathogenic prion aggregates.

Keywords: Prions; methionine; oxidation; protein aggregation; protein misfolding.

Figure 1.

The location and structure of methionine residues in the structured C-terminal domain (125–228) of the human prion protein (huPrP). (a) A linear representation of the secondary structure content of huPrP generated by POLYVIEW [110]. The primary sequence of huPrP(125–228) is shown at the top and methionine residues are colored in magenta. Alpha helices are represented as red cylinders and beta strands are represented as green arrows. (b) The tertiary structure of huPrP(125–228) (PDB:1QLX) [44]. Methionine residues are labelled by their residue number in the human prion protein. Solvent exposed methionines are shown in blue and buried/partially buried methionines are shown in red. Solvent accessible surface areas (SASAs) were calculated in pyMol with sampling density set to 3. SASA values are represented as a mean and standard deviation derived from six unique solution NMR structures (PDB: 1QLX,1QLZ,1QM0,1QM1,1QM2,1QM3) [44].

Figure 2.

A schematic overview of a novel method for the accurate quantification of methionine oxidation [36]. As shown in the left panel, methionine oxidation can typically occur in vitro during sample preparation, and the measured levels of oxidation may not be reflective of levels of methionine oxidation in vivo. This problem can be solved by blocking unoxidized methionines with heavy-labelled ¹⁸O (right panel). At the time of extraction, unoxidized methionine residues are converted to methionine sulfoxide residues with a heavy oxygen atom label (red). The heavy oxygen label serves as a blocking agent and prevents the in vitro accumulation of methionine sulfoxides, labelled with a naturally occurring light oxygen atom (green). The differences in mass between heavy and light oxygen can be used to distinguish in vivo methionine oxidation from in vitro blocking by mass spectrometry. The resulting isotope clusters can then be deconvoluted and quantified by custom algorithms, resulting in a measurement of in vivo fractional oxidation.

Figure 3.

The methionine redox cycle. Methionine can be oxidized by reactive oxygen species (ROS) resulting in the formation of methionine sulfoxide. Methionine sulfoxide can be repaired by the action of methionine sulfoxide reductase (Msr) enzymes, resulting in a disulfide cysteine intermediate of Msr. Catalytic activity of Msr is restored by the action of thioredoxin (Trx) and thioredoxin reductases, using dihydronicotinamide-adenine dinucleotide phosphate (NADPH) as a cofactor, producing one molecule of nicotinamide adenine dinucleotide phosphate (NADP⁺).

Figure 4.

A proposed model for the role of methionine oxidation in prion protein misfolding and aggregation. Unoxidized methionines are represented as green circles and oxidized methionines are represented as red circles. Surface methionines in PrP can become oxidized resulting in the formation of a molten globule state and increasing the solvent exposure of buried methionines. In the molten globule state, buried methionines can become oxidized, resulting in the total destabilization and conformational rearrangement of the prion protein. It is unknown whether or not the β-structured intermediate induced by methionine oxidation is on pathway to the formation of PrP^Sc. Schematic representations of PrP^C and the molten globule state were derived from PDB:1QLX [44]. Schematic representations of PrP^Sc were derived from PDB:2RNM [111].