聚脯胺酸 (polyproline) 可以形成 PPI 或是 PPII 結構，而胜肽鏈上的
n→π* 作用力被視為是穩定 PPII 結構的重要因素。本實驗室在先前的研究中利
用脯胺酸 (proline) 衍生物，合成一系列聚脯胺酸胜肽，探討立體電子效應對於
聚脯胺酸結構在 PPI 與 PPII 結構間轉換的活化能之影響，並在 HP7 胜肽中置
換脯胺酸衍生物來探討立體電子效應對於 β-hairpin 的結構與穩定度之影響。
本次研究中，除了將 (2S)-4,4-difluoroproline (Dfp) 與 (2S)-4-ketoproline
(Kep) 兩種脯胺酸衍生物置換到聚脯胺酸上，且利用圓二色光譜探討其對於 PPI
與 PPII 結構的影響外，並藉由理論計算研究得到一系列聚脯胺酸胜肽的結構資
訊，包含 φ 、 ψ 以及 χ 二面角都是在聚脯胺酸中重要的結構資訊。理論計算
部分我們使用 Gaussian 軟體， DFT 方法進行結構最佳化，實驗所合成之 HN-
(Pro)5X(Pro)5-OH 胜肽在計算上都以Ac-(Pro)2X(Pro)2-OMe 分子模型來取代。計
算結果顯示，當 χ 為正值時， Dfp 、 Kep 與 thiaproline (Thp) 的 pyrrolidine
ring 不會形成典型的 endo pucker ，而是形成類似扭曲平面的特殊構形，其被置
換到聚脯胺酸胜肽上，對於 PPI結構會有明顯的影響。
另外在 HP7 胜肽的部分，先前研究顯示當 Pro 被置換成 Thp ，原本 wild-
type 的 β-hairpin 結構會崩解，形成 random coil 。因此我們截取中間形成 turn
的序列 NPATGK 以及 turn 兩端的 Trp ，建立 WNPATGKW 作為分子計算模
型，利用 ONIOM 計算方法進行一系列的計算。計算結果顯示， Thp 的置換使
得 Asn 的 C=O 與 Lys 的 N-H 之間的距離加大，氫鍵作用力消失，並且造成
Pro5 與 Trp3 之間的距離縮短，使得兩者之間的空間推擠作用變大，結構變得
不穩定，因此無法形成穩定的 β-hairpin 結構。

Polyproline can form either type I (PPI) or type II (PPII) helices. The backbone
n→π* interactions have been suggested to play a critical role in stabilizing a PPII helix.
In our previous study, we prepared a series of polyproline peptides with proline
derivatives incorporated to evaluate stereoelectronic effects on the transition energy
barrier of the conversion between PPI and PPII conformation. We also incorporated
proline derivatives into the HP7 peptide to investigate stereoelectronic effects on β-
hairpin.
In this work we prepared two polyproline peptides in which (2S)-4-ketoproline
(Kep) and (2S)-4,4-difluoroproline (Dfp) were incorporated. Circular dichroism (CD)
spectroscopy was used to characterize their conformation. We also used a
computational approach to study the effects of Kep, Dfp and thiaproline (Thp) on
polyproline conformation. For each model compound and system, hybrid density
function theory (DFT) calculations, as implement in Gaussian 09, were carried out on
the conformation, and the dihedral angels φ, ψ and χ were measured for polyproline.
We used Ac-(Pro)2X(Pro)2-OMe as a model system for the computation. Typical 4-
substituted Pro derivatives display either a Cγ
-endo or a Cγ
-exo ring pucker. Such
conformations are not readily accessible to Ac-Kep-OMe, Ac-Dfp-OMe and Ac-Thp-
OMe. When Ac-Kep-OMe, Ac-Dfp-OMe and Ac-Thp-OMe adopt a Cγ
-endo pucker,
their pyrrolidine ring is significantly distorted and distinctly different from other 4-
substituted proline derivative. This may explain why Kep, Dfp, and Thp destabilize PPI
conformation as observed in experiments.
Moreover, the HP7 peptide could not form β-hairpin upon incorporating Thp into
the peptide. We chose WN-X-ATGKW sequence as a model system for computation as
it can form H-bonds required for a β-turn in HP7. ONIOM calculation was carried out on the structure. The calculated result reveals that Thp substitution on HP7 makes the
H-bond between C=O of Asn4 and N-H of Lys9 broken as the distance between the
residues increases. Thp substitution also makes the distance between Pro5 and Trp3
decrease, which may increase the steric repulsion between the residues. The
computational results may in part explain why Thp dramatically destabilize HP7.

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