Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: May 1, 2013
Publication Date: May 24, 2013
Citation: Silva, C.J., Dynin, I.A., Erickson-Beltran, M.L., Hui, C., Carter, J.M. 2013. Oxidation of methionine in PrP is dependent upon the oxidant and the amino acid two positions removed(Abstract). Meeting Abstract. PS 01. Technical Abstract: Background/Introduction. Methionine oxidation has been shown both to be associated with prion formation and implicated in the inhibition of amyloid formation in model systems. This work is based on model systems where hydrogen peroxide was used as an oxidant. Materials and Methods. We developed a sensitive mass spectrometry-based method to study the oxidation of a methionine at position 216 (Met216) in sheep PrP. Oxidation of Met216(MetSO216)has been implicated in prion formation. In order to test the susceptibility of Met216 to oxidation by molecular oxygen, we prepared clones containing three polymorphisms of sheep PrP at position 218 (Ile, Val and Thr). The clones were grown in medium supplemented with either natural abundance NH4Cl or 15NH4Cl, in order to obtain isotopically labeled recombinant protein. The recombinant proteins were purified and then subjected to air oxidation. Results and Conclusions. Our analysis showed that the proportion of MetSO216 was highly dependent upon the amino acid residue at position 218. The Met216 in the sheep rPrP protein containing 218Ile was more susceptible than the Met216 in the sheep rPrP with Val218. The Met216 was least oxidized when the protein contained Thr218. In model studies, the Met216 is resistant to oxidation when the oxidant is hydrogen peroxide. This indicates that the presence of Ile218 in sheep and elk PrP renders the Met216 intrinsically more susceptible to air oxidation. Our results also indicate that Val218 is likely to make the analogous methionine in the human form of PrP more susceptible to air oxidation.