Conformational change in full-length mouse prion: a site-directed spin-labeling study.

Inanami, Osamu; Hashida, Shukichi; Iizuka, Daisuke; Horiuchi, Motohiro; Hiraoka, Wakako; Shimoyama, Yuhei; Nakamura, Hideo; Inagaki, Fuyuhiko; Kuwabara, Mikinori

Conformational change in full-length mouse prion: a site-directed spin-labeling study.

Inanami, Osamu; Hashida, Shukichi; Iizuka, Daisuke et al.

Biochemical and Biophysical Research Communications, 2005, 335(3), 785-792

Permalink : https://hdl.handle.net/2115/27969

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アクセス権	open access
URI	https://hdl.handle.net/2115/27969
タイトル	Conformational change in full-length mouse prion: a site-directed spin-labeling study.
著者	Inanami, Osamu ; Hashida, Shukichi ; Iizuka, Daisuke ; Horiuchi, Motohiro ; Hiraoka, Wakako ; Shimoyama, Yuhei ; Nakamura, Hideo ; Inagaki, Fuyuhiko ; Kuwabara, Mikinori
e-Rad_Researcher 10193559 著者名 Inanami, Osamu
著者名 Hashida, Shukichi
著者名 Iizuka, Daisuke
e-Rad_Researcher 30219216 著者名 Horiuchi, Motohiro
著者名 Hiraoka, Wakako
著者名 Shimoyama, Yuhei
e-Rad_Researcher 30207906 著者名 Nakamura, Hideo
e-Rad_Researcher 70011757 著者名 Inagaki, Fuyuhiko
著者名 Kuwabara, Mikinori
言語	英語
キーワード	Site-directed spin-labeling
	Electron spin resonance
	Prion
	Conformational change
	pH-sensitive region
発行日	2005-09-30
出版者	Elsevier Inc.
収録物名	Biochemical and Biophysical Research Communications
巻(号)	335(3)
ページ	785-792
抄録	The structure of the mouse prion (moPrP) was studied using site-directed spin-labeling electron spin resonance (SDSL-ESR). Since a previous NMR study by Hornemanna et al., [Hornemanna, Korthb, Oeschb, Rieka, Widera, Wüthricha, Glockshubera, Recombinant full-length murine prion protein, mPrP (23–231): purification and spectroscopic characterization, FEBS Lett. 413 (1997) 277–281] has indicated that N96, D143, and T189 in moPrP are localized in a Cu2+ binding region, Helix1 and Helix2, respectively, three recombinant moPrP mutations (N96C, D143C, and T189C) were expressed in an Escherichia coli system, and then refolded by dialysis under low pH and purified by reverse-phase HPLC. By using the preparation, we succeeded in preserving a target cystein residue without alteration of the α-helix structure of moPrP and were able to apply SDSL-ESR with a methane thiosulfonate spin label to the full-length prion protein. The rotational correlation times (τ) of 1.1, 3.3, and 4.8 ns were evaluated from the X-band ESR spectra at pH 7.4 and 20 °C for N96R1, D143R1, and T189R1, respectively. τ reflects the fact that the Cu2+ binding region is more flexible than Helix1 or Helix2. ESR spectra recorded at various temperatures revealed two phases together with a transition point at around 20 °C in D143R1 and T189R1, but not in N96R1. With the variation of pH from 4.0 to 7.8, ESR spectra of T189R1 at 20 °C showed a gradual increase of τ from 2.9 to 4.8 ns. On the other hand, the pH-dependent conformational changes in N96R1 and D143R1 were negligible. These results indicated that T189 located in Helix2 possessed a structure sensitive to physiological pH changes; simultaneously, N96 in the Cu2+ binding region and D143 in Helix1 were conserved.
資源タイプ	学術雑誌論文
出版タイプ	Accepted Manuscript
PISSN	0006-291X
EISSN	1090-2104
関連情報	PMID 16095563
関連情報	URI http://www.sciencedirect.com/science/journal/0006291X
関連情報 (isVersionOf)	出版社版DOI https://doi.org/10.1016/j.bbrc.2005.07.148