以聚脯胺酸調控色胺酸與金奈米粒子之距離探討螢光淬熄效應

Distance-dependent Fluorescence Quenching of Tryptophan by Gold Nanoparticles within 5 nm Spaced by Polyprolines

賴盈禎(Ying-Chen Lai)

指導教授 ： 朱立岡

國立清華大學/理學院/化學系所/碩士(2017年)

摘要

當金奈米粒子靠近螢光分子時，能誘發其螢光淬熄效應，且此效應與兩者的距離有關，但目前尚無對短距離(< 5 nm)進行系統性的研究。在此，吾人選用色胺酸作為螢光分子，以及利用聚脯胺酸(polyproline，PP)於水溶液中形成堅固的PPII結構作為分子尺(molecular ruler)，並藉由調整脯胺酸的數量，以控制金奈米粒子與色胺酸距離在5 nm內，探討金奈米粒子對螢光的淬熄效應。

吾人合成四種不同長度的胜肽，序列為CAPnWN，其中n=3、6、10、13，並成功地將其透過半胱胺酸的硫醇基，以金硫共價鍵鍵結於金奈米粒子表面。以靜態紫外/可見光吸收光譜、螢光光譜、衰減全反射式紅外吸收光譜以及穿透式電子顯微鏡，觀察其合成前後形貌的變化。除了靜態光譜資訊，吾人亦利用時間相關單光子技術偵測其螢光生命期，以266 nm 的雷射激發，收集355–365 nm的螢光訊號。吾人發現色胺酸在CAPnWN中的螢光生命期較其單體短，而當CAPnWN被修飾於金奈米粒子表面後，其色胺酸的螢光生命期隨著與金奈米粒子的距離越近而越短。若以奈米表面能量轉移機制分析此距離相關的螢光衰減速率，得到能量轉移速率與距離倒數呈現1.3次方關係，此結果與理論值4差異過大，因此吾人認為奈米表面能量轉移非主因。若以電子轉移機制分析，並考慮色胺酸彼此靠近可能有π-π 堆疊的影響，可以衰減常數0.1 Å-1擬合吾人數據，與以往文獻一致。故吾人認為色胺酸螢光淬熄的主因為電子轉移，以聚脯胺酸的胜肽骨架作為跳板，金奈米粒子作為電子受體。故由本實驗得知，聚脯胺酸不僅可作為分子尺，亦可作為電子轉移的模板。

關鍵字

聚脯胺酸 ； 色胺酸 ； 金奈米粒子 ； 螢光淬熄

並列摘要

Gold nanoparticles (AuNPs) often induce the quenching of excited states of fluorophores when they are close. The quenching efficiency has been observed as a function of distance between the AuNP and the fluorophore. However, there has not been discussed about AuNP-induced quenching within 5 nm. In this work, tryptophan is served as the fluorophore to study the AuNP-induced quenching and polyprolines (PP), which form the rigid PPII structure in solution, are used as molecular rulers to separate the AuNP and Trp by tuning the numbers of the proline units within 5 nm.
The surfaces of the AuNP were successfully anchored with peptides, CAPnWN which n=3, 6, 10, 13, through the terminal cysteine to form the Au-S bonding. The optical properties of AuNP-CAPnWN were characterized by UV/Visible absorption spectroscopy, fluorescence spectroscopy, ATR-FTIR and TEM. Besides the steady-state optical characterization, the temporal profiles of the tryptophan fluorescence at 355-365 nm upon 266 nm excitation were collected with the time-correlated single-photon counting. We found that the fluorescence kinetics of tryptophan in CAPnWN is slightly accelerated in comparison with the pure tryptophan. As the CAPnWN are attached onto the gold nanoparticles, the fluorescence kinetics becomes more accelerated as the separation becomes shorter. Both energy-transfer and electron-transfer processes are considered to be deactivation pathways. However, the energy transfer rate depends on the 1/d1.3 separation distance which is far from the nanosurface energy transfer mechanism. Therefore, the energy transfer from Trp to AuNP is not the major reason. Then, considering the electron transfer rate (k_ET) as an exponential function of the peptide chain length (k_ET∝e^(-βd)), and the π-stacking interaction between adjacent CAPnWN, the β could be fitted as 0.1 Å-1 same as previous studies. As a result, the fluorescence quenching of Trp by AuNP depends on their distance is attributed to the electron transfer mechanism. The polyprolines are served as stepping stones and the AuNPs are the electron acceptors. It is noteworthy that the polyproline can not only be the molecular ruler but also be the relay for electron transfer.

並列關鍵字

polyproline ； tryptophan ； nanoparticle ； fluorescence ； quenching

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