以色胺酸及FlAsH組成之凋亡蛋白酶3螢光感測多肽

FlAsH Labeled Peptide with Tryptophan Residues as a Caspase-3 Sensor

鄭皓蔚(Hao-Wei Cheng)

指導教授 ： 李賢明 ； 謝發坤

國立中央大學/理學院/化學系/碩士(2015年)

摘要

本論文是以色胺酸對螢光團螢光淬滅的特性為基礎，設計並合成蛋白酶活性
感測器 FlAsH-3W2F-TCǶ利用一系列 FlAsH 螢光染料標示胜肽的螢光性質進行研究，發現帶有芳香環側鏈的胺基酸中，相較於阻胺酸與苯Ҙ胺酸，色胺酸對FlAsH 螢光染料的螢光有最好的淬滅效果；胜肽上的色胺酸數量越多對螢光的淬滅效果也越好Ƕ色胺酸對於螢光染料的淬滅是根據光誘發電子轉移，透過螢光分子與色胺酸之間碰撞所形成暫時的分子軌域，提供電子轉移的通道，使螢光團激發的能量得到鬆弛Ƕ在以 Trypsin 為模型的蛋白酶活性實驗中，感測器水解前後有將近 9 倍的螢光強度增加；在 Caspase-3 目標蛋白酶活性實驗中，也觀測到明顯的螢光增強Ƕ以色胺酸做為淬滅團在螢光感測器上的應用是可行的。

關鍵字

感測器 ； 凋亡蛋白酶3 ； 螢光

並列摘要

We synthesized a Caspase-3 sensor, FlAsH-3W2F-TC, whose fluorescence was quenched by tryptophan inside the peptidyl sensor backbone. By studying a series of FlAsH-labeled peptide sequences, tryptophan containing peptide sequence showed higher quenching ability than histidine and phenylalanine peptide sequences; the more tryptophan the peptide sequence contains, the higher quenching efficiency we get. The quenching phenomena is based on photo-induced electron transfer(PET). The excited fluorophore relax by conducting an electron through temporary collisional molecular orbital. In trypsin model experiment, our sensor showed ~9-fold fluorescence increasing after hydrolysis; in Caspase-3 experiment, our sensor also showed significant fluorescence increasing. Finally, the application of tryptophan as a natural quencher in peptidyl fluorescent sensor is feasible and potential.

參考文獻

(1) Elmore, S. Toxicologic Pathology. 2007, 35, 495.

J. Nature Protocols. 2010, 5, 294.

Biological Chemistry 1998, 273, 9357.

(8) Ashkenazi, A.; Dixit, V. M. Science 1998, 281, 1305.

(10) Fulda, S.; Debatin, K. M. Oncogene 2006, 25, 4798.