Octarepeat 是 prion protein (PrP)中的一段序列，這段序列會重複的出現在prion protein 胺基酸序列 60-91這段區域中， 而且 octarepeat 已知對於 Cu2+ 有相當良好的親和力。Proline 可以形成 exo 或是 endo 兩種構形，且由於立體電子效應的關係，當 4R 位置有拉電子基取代時 proline 會偏好 Cγ-exo 構形，而拉電子取代基位於 4S 位置時 proline 則會偏好 Cγ-endo 構形，在 octarepeat 的原始結構中，proline 較偏向 endo 構形。因此在論文中我們以 4號位置具有拉電子取代基的 proline 衍生物置換octarepeat的 proline，探討 proline 的構形對於 octarepeat 與 Cu2+ 的親和力是否會有影響，而置換的 proline 衍生物包括 4R-hydroxyproline (Hyp)、4R-fluoroproline (Flp)、4R-methoxyproline (Mop), 4S-hydroxyproline (hyp)、 4S-fluoroproline (flp) 和 4S-methoxyproline (mop)。經過置換後的胜肽在加入 Cu2+ 之後的螢光焠熄比例和錯合比例都沒有改變，而從 CD 和螢光的 glycine 競爭實驗所得之解離常數 (Kd) 比較得知，除了 Hyp 和 hyp 取代的胜肽解離常數有明顯提高之外，大部分經過置換的胜肽都沒有影響。解離常數的改變應該是因為 Hyp 和 hyp 的 OH 官能基產生的氫鍵影響到了胜肽和 Cu2+ 周圍環境中的水，導致 Hyp、hyp 取代的胜肽和 Cu2+ 配位能力降低，解離常數提高。雖然 Hyp和 hyp 取代的胜肽解離常數都有改變，但是從 CD 的 Far UV、 Near UV 和Visible 的光譜測量發現，hyp 取代的胜肽和 Cu2+ 後的構形與其他胜肽類似，而 Hyp 取代卻明顯不同，這其中差異應該是在於Hyp 和 hyp 兩者的 pyrrolidine ring 有不同的構形所造成。

The octapeptide (PHGGGWGQ) is a fragment of the prion protein (PrP) and this sequence is repeated in the human PrP 60-91 region. The octarepeat is known to have great affinity to copper (II) ions. Proline can form either a Cγ-exo ring pucker or a Cγ-endo ring pucker. Due to stereoelectronic effects, an exo ring pucker is preferred if an electron-withdrawing group is on the 4R position of proline while an electron-withdrawing substituent on 4S position favors an endo ring pucker. The proline of the octarepeat peptide in the native prion protein adopts an endo pucker. In this work we have studied how the ring pucker of proline affects the binding affinity of the peptide to Cu2+ ions by using 4R-hydroxyproline (Hyp)、4R-fluoroproline (Flp)、4R-methoxyproline (Mop), 4S-hydroxyproline (hyp)、 4S-fluoroproline (flp) and 4S-methoxyproline (mop). The fluorescence quenching behavior and binding stoichiometry of proline derivatives containing PrP peptides are similar to the wild type. The dissociation constant (Kd) measured by the fluorescence and CD glycine competition experiments is similar to that of the wild type with the exception of Hyp and hyp substituted peptides. The significent change in the dissociation constants of these two peptides might be due to the hydration of the OH group, which pertubes the environments of the coordination site and affects the coordination between Cu2+ and the peptides. Especially, Hyp substituted peptide has a significently different CD spectrum from those of the other PrP peptides upon binding to Cu2+. The difference between Hyp and hyp sunstituted peptides could be due to the conformation of their pyrrolidine rings.

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