title: Oxidation of methionine 216 in sheep and elk prion protein is highly dependent upon the amino acid at position 218 but is not important for prion propagation.
abstract: We employed a sensitive mass spectrometry-based method to deconstruct, confirm, and quantitate the prions present in elk naturally infected with chronic wasting disease and sheep naturally infected with scrapie. We used this approach to study the oxidation of a methionine at position 216 (Met216), because this oxidation (MetSO216) has been implicated in prion formation. Three polymorphisms (Ile218, Val218, and Thr218) of sheep recombinant prion protein were prepared. Our analysis showed the novel result that the proportion of MetSO216 was highly dependent upon the amino acid residue at position 218 (I > V > T), indicating that Ile218 in sheep and elk prion protein (PrP) renders the Met216 intrinsically more susceptible to oxidation than the Val218 or Thr218 analogue. We were able to quantitate the prions in the attomole range. The presence of prions was verified by the detection of two confirmatory peptides: GENFTETDIK (sheep and elk) and ESQAYYQR (sheep) or ESEAYYQR (elk). This approach required much smaller amounts of tissue (600 μg) than traditional methods of detection (enzyme-linked immunosorbent assay, Western blot, and immunohistochemical analysis) (60 mg). In sheep and elk, a normal cellular prion protein containing MetSO216 is not actively recruited and converted to prions, although we observed that this Met216 is intrinsically more susceptible to oxidation.
authors: Western Regional Research Center, United States Department of Agriculture , Albany, California 94710, United States.
authors: Christopher J
authors: CJ
authors: Silva
authors: Irina
authors: I
authors: Dynin
authors: Melissa L
authors: ML
authors: Erickson
authors: Jesús R
authors: JR
authors: Requena
authors: Aru
authors: A
authors: Balachandran
authors: Colleen
authors: C
authors: Hui
authors: Bruce C
authors: BC
authors: Onisko
authors: John Mark
authors: JM
authors: Carter
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003670:Deer / Q000235:genetics / Q000378:metabolism; D008715:Methionine / Q000737:chemistry; D010084:Oxidation-Reduction; D011110:Polymorphism, Genetic; D018621:PrPC Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D018620:PrPSc Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D011328:Prions / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D012608:Scrapie / Q000235:genetics / Q000378:metabolism; D012756:Sheep / Q000235:genetics / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D034081:Wasting Disease, Chronic / Q000235:genetics / Q000378:metabolism
chemical_list: D018621:PrPC Proteins; D018620:PrPSc Proteins; D011328:Prions; D008715:Methionine