小分子藥物普拉辛活化EphA2受體產生的生物物理特徵探討

Biophysics Characterization of EphA2 Receptor Upon Activation by Prazosin

林裕凱(Yu-Kie Lin)

指導教授 ： 周綠蘋

國立臺灣大學/醫學院/生物化學暨分子生物學研究所/碩士(2018年)

https://doi.org/10.6342/NTU201801267

摘要

根據實驗室過去的研究發現，在具有蕾莎瓦藥物抗性的肝癌細胞株中有大量EphA2受體(Ephrin type-A receptor 2 or EphA2 receptor)的表現。近期文獻顯示，在有配體結合誘導的情況下，EphA2受體絲胺酸897位點的磷酸化下降並抑制Akt的磷酸化，使得癌症細胞的增生及轉移受阻，成為有效抑制癌症生長的途徑。然而，尚未有人發表強力有效的促效劑可以應用在癌症治療上。而且近期關於EphA2受體分子作用層面的研究也相當有限，使得找尋EphA2受體的促效劑受阻。因此，我們將在本篇研究中，透過實驗室過去研究發現的，具有活化EphA2受體潛力的促效劑普拉辛(prazosin)，作為研究的材料。我們嘗試更專注於EphA2 LBD(ligand binding domain)的作用機轉以及與普拉辛之間的作用力，並成功純化了EphA2 LBD重組蛋白，也進行了表面電漿共振的分析。更進一步利用分子動力學模擬分析EphA2受體與普拉辛結合後產生的結構變化及動性變化，並將之與EphA2受體及其天然配體Ephrin A5結合的結果作比較，藉此了解兩組模擬模型分子作用的異同處。

在我們的研究中觀察到EphA2受體中，由組胺酸54(GLN 54)到纈胺酸69(VAL 69)組成的β折板，在EphA2受體與配體結合後產生的角度偏折可能與EphA2受體活化有關。在普拉辛的分子動力學模擬中，普拉辛結合下的EphA2受體組胺酸54(GLN 54)到纈胺酸69(VAL 69)組成的β折板，呈現出與Ephrin A5相同的結構動性特徵。這個二級結構的動性變化使得未來在藥物設計上有一個可能的篩選指標，成為EphA2受體促效劑開發上的一大助力。

關鍵字

EphA2 ； 普拉辛 ； 分子動力學模擬 ； 喹唑啉

並列摘要

The EphA2 receptor is overexpressed in sorafenib-resistance hepatic cancer cells. When the ligand is absent, EphA2 has been reported to be a pro-oncogenic factor due to Akt activation. However, the effect can be reversed when ligand binds to EphA2. Currently, none of therapeutic agonist of the EphA2 receptor has been reported. Because current understanding of molecular mechanism of the EphA2 receptor activation is limited, designing an effective agonist on the EphA2 receptor is a difficult task. In this work, we successfully purified the EphA2 LBD recombinant protein and used surface plasma resonance analysis to understand the relationship between prazosin and EphA2 LBD recombinant protein. Subsequently, we have built EphA2-ligand docking models: EphA2-prazosin, in which prazosin is a potential EphA2 agonist that was previous identified in our lab. We have also conducted molecular dynamics simulations (MDS) to investigate the difference of conformational and dynamics changes upon EphA2 receptor activation.

Based on the analysis of MDS trajectories, we found that there is a significant dynamics change of the EphA2 β-sheet between Gln54 and Val69, which may be related to EphA2 receptor activation. Prazosin-EphA2 MDS showed that the dynamics changes of the EphA2 β-sheet were correlated with the activation of the EphA2 receptor-Ephrin A5. Therefore, it is anticipated that this finding could serve as a structural and dynamical fingerprint for discovering novel EphA2 agonists.

並列關鍵字

EphA2 ； Prazosin ； Molecular dynamic simulation ； Quinazoline

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