開發利用聚集誘導發光染料修飾之胜肽偵測蛋白激酶活性快篩方法

Develop Aggregation-induced Emission Dyes Coupling with Peptide Substrate as Rapid Protein Kinase Activity Detection Kit.

倪永婷(Ni Yung-Ting)

指導教授 ： 李賢明 ； 陳錦地

國立臺灣師範大學/理學院/化學系/碩士(2018年)

https://doi.org/10.6345/THE.NTNU.DC.071.2018.B05

摘要

陳錦地老師實驗室設計出具有推、拉電子結構的螢光團，且不共平面的共軛結構使螢光團具有誘導堆疊放光的特性，並以帶正電荷或負電荷離子修飾增加親水性，開發此系列螢光團對生物相關及醫學研究應用性。
將此系列螢光團修飾在胜肽上，作為偵測蛋白激酶A活性的生物探針，蛋白激酶A在細胞內能催化蛋白質磷酸化，參與細胞內代謝、凋亡、分化…等重要生理機能，蛋白激酶A功能異常與癌症或神經衰退重要疾病有高度相關性。
本實驗利用鑭系元素氧化物吸附磷酸化胜肽，且氧化物不溶於水可誘導螢光團堆疊放光，搭配濾紙過濾固體並利用以螢光強度作為磷酸化胜肽半定量，胜肽磷酸化程度取決於蛋白激酶A活性強弱，成功開發具有專一性蛋白激酶活性螢光快篩方法。

關鍵字

聚集誘導發光 ； 蛋白激酶 ； 活性測試

並列摘要

We synthesized a series of aggregation-induced emission (AIE) dyes which were designed by Dr. Chin-Ti Chen’s group. The AIE dyes have nonplanar conjugat-ed structure which is crucial to provide the unique fluorescent turn-on property upon aggregation. In order to increase water solubility of our probes, we modified AIE dyes with hydrophilic ligands such as positively charge ammonium group or nega-tively charged sulfite group.
Kinase family is responsible for catalyzing intracellular protein phosphoryla-tion. It is one of the most important enzymes target in human diseases including cancer and neurodegenerative disorders. To apply AIE dyes in detection of biological events, we coupled AIE dye with peptide substrate as bioluminescent probe for Pro-tein Kinase A (PKA) detection.
We used lanthanum(III) oxide that can interact with phosphorylated AIE-peptide, and its poor solubility in water enhanced the AIE fluorescence. The solid mixture was then immobilized on a filter paper for quantitative analysis of PKA activity. The approach is potentially useful as a rapid kinase activity detection kit based on AIE dyes.

並列關鍵字

Aggregation-induced Emission ； Protein Kinase ； Activity Detection Kit

參考文獻

4. Hong, Y. N.; Lam, J. W. Y.; Tang, B. Z., Aggregation-induced emission. Chemi-cal Society Reviews 2011, 40 (11), 5361-5388.

5. Wu, J. S.; Liu, W. M.; Ge, J. C.; Zhang, H. Y.; Wang, P. F., New sensing mecha-nisms for design of fluorescent chemosensors emerging in recent years. Chemical Society Reviews 2011, 40 (7), 3483-3495.

6. Hong, Y. N.; Lam, J. W. Y.; Tang, B. Z., Aggregation-induced emission: phe-nomenon, mechanism and applications. Chemical Communications 2009, (29), 4332-4353.

7. Kwok, R. T. K.; Leung, C. W. T.; Lam, J. W. Y.; Tang, B. Z., Biosensing by lu-minogens with aggregation-induced emission characteristics. Chemical Society Re-views 2015, 44 (13), 4228-4238.

8. Liu, H. L.; Song, P. S.; Wei, R. R.; Li, K.; Tong, A. J., A facile, sensitive and se-lective fluorescent probe for heparin based on aggregation-induced emission. Talanta 2014, 118, 348-352.