Design of anti- and pro-aggregation variants to assess the effects of methionine oxidation in human prion protein

Abstract

Prion disease is characterized by the ␣3␤ structural conversion of the cellular prion protein (PrP C ) into the misfolded and aggregated ''scrapie'' (PrP Sc ) isoform. It has been speculated that methionine (Met) oxidation in PrP C may have a special role in this process, but has not been detailed and assigned individually to the 9 Met residues of full-length, recombinant human PrP C [rhPrP C (23-231)]. To better understand this oxidative event in PrP aggregation, the extent of periodate-induced Met oxidation was monitored by electrospray ionization-MS and correlated with aggregation propensity. Also, the Met residues were replaced with isosteric and chemically stable, nonoxidizable analogs, i.e., with the more hydrophobic norleucine (Nle) and the highly hydrophilic methoxinine (Mox). The Nle-rhPrP C variant is an ␣-helix rich protein (like MetrhPrP C ) resistant to oxidation that lacks the in vitro aggregation properties of the parent protein. Conversely, the Mox-rhPrP C variant is a ␤-sheet rich protein that features strong proaggregation behavior. In contrast to the parent Met-rhPrP C , the Nle/Moxcontaining variants are not sensitive to periodate-induced in vitro aggregation. The experimental results fully support a direct correlation of the ␣3␤ secondary structure conversion in rhPrP C with the conformational preferences of Met/Nle/Mox residues. Accordingly, sporadic prion and other neurodegenerative diseases, as well as various aging processes, might also be caused by oxidative stress leading to Met oxidation.conformational switch ͉ expanded genetic code ͉ methionine sulfoxide reductase ͉ noncanonical analogs ͉ unnatural amino acid sequence

Molecular Dynamics as an Approach to Study Prion Protein Misfolding and the Effect of Pathogenic Mutations

“…It was suggested that methionine oxidation could trigger misfolding. This hypothesis was later corroborated by experimental studies of recPrP that used norleucine and methoxinine as analogues of the hydrophobic, non-oxidized form or the hydrophilic oxidized form of methionine, respectively [165]. The norleucine variant exhibited a stabilized α-helical structure and low aggregation propensity, whereas the methoxinine variant largely consisted of β-structure and had a high tendency to aggregate.…”

Post-aggregation Oxidation of Mutant Huntingtin Controls the Interactions between Aggregates

“…Indeed, for example, oxidation of a Met residue has been reported in an A␤ peptide forming senile plaques of Alzheimer's diseases (31) and also in infectious forms of PrP proteins in prion diseases (32). Furthermore, the effects of Met oxidation on the kinetics of fibrillar aggregation in vitro have been examined in several disease-related proteins such as ␣-synuclein (33), PrP (34), and ApoA-I (35). In these previous studies, oxidative modifications have been assumed to precede the aggregation of protein molecules and thereby affect its formation kinetics; however, it remains obscure when proteins are oxidized during the formation of pathological inclusions in vivo.…”

Convergent signaling pathways—interaction between methionine oxidation and serine/threonine/tyrosine O-phosphorylation

“…However, there is also precedent for Met oxidation causing large-scale changes in protein structure (Griffiths and Cooney 2002;Wolschner et al 2009;Pan et al 2010;Marondedze et al 2013). If this were the case, it might easily lead to exposure of a previously cryptic phosphorylation site or to hide a previously exposed site.…”