臺灣博碩士論文加值系統

蛇是會讓人害怕的動物之一，蛇毒更是讓人與其他動物死亡的主要原因，因為毒液會引起溶血作用，破壞紅血球，有些也會造成血液的凝結等。在本論文裡首先概略的提到目前的實驗報告，目前將眼鏡蛇毒蛋白分成三大類：心臟毒 (Cardiotoxin)、神經毒 (Neurotoxin)、和磷脂水解酵素 (Phospholipase A2)。
台灣眼鏡蛇心臟毒素被分析後，發現有五種蛋白，命為CTX A1~A5，接著從序列中Pro30和Ser28存在與否，將這五種分成P型和S型兩類。其中比較有意思的是，發現在台灣眼鏡蛇毒蛋白中，第三型心臟蛇毒有束縛水 (bound water) 的現象，第三型心臟蛇毒是屬於P型的，由三維結構來看，束縛水存在於第二個迴圈 (loop II) 內，而束縛水的功用，目前是認為它與迴圈上的某幾個胺基酸產生氫鍵，氫鍵網可以穩定其上方β-摺板 (β-sheet) 的結構。
本論文中，我們從熱力學中Gibbs自由能的觀點，藉由分子動力模擬的計算方式，用經驗價鍵模型 (Empirical valence bond model)，以及自由能微擾法 (Free energy perturbation method) 的方法，代替實驗上真實蛋白質的突變，有效的計算出自由能差的變化量，來研究此現象並用以解釋第二迴圈上的Proline30胺基酸的重要。

Snake venoms make people die often and snakes only live by them. They consist of cardiotoxin (CTX), neurotoxin, and phospholipase A2 (PLA2). From the published reports, there are five kinds of venom proteins in the CTX molecules found in Taiwan cobra (Naja naja atra) venom, and they were named CTX A1~A5. Next, CTX proteins have been classified into two types, P-type and S-type, which are distinguished by the presence of Pro30 and Ser28, respectively, near the tip of loop II.
An interesting phenomenon called "bound water" was found in recent reports. By the NMR experiments, this water molecule exists in loop II of CTX A3 protein, which is a P-type protein. It is possible for its function to form a hydrogen-binding network in loop II, and this network may keep the above β-sheet structures stable.
In this thesis, we study this phenomenon by molecular dynamics simulations. We first mutate Pro30 to Gly by using the empirical valence bond model. And then by using the free energy perturbation method, we calculate the free energy differences. From the point of view of the thermodynamic cycle, we can get the values of ΔΔG, the changes of the Gibbs free energy differences. The meaning of ΔΔG is to discuss the effects of mutation of side chains with bound water on protein. Finally, after comparing the results of P-type proteins with S-type ones, we suggest that the amino acid, Pro30, may be a "key" for the bound water. Namely, although Pro30 is not directly bound with the bound water by hydrogen bonds, this residue still affects this bound water molecule.

CONTENTS
I. Introduction 1
I-1.Snake venom, 2
I-2.Overall structure of CTX, 3
I-3.Function of CTX, 4
I-4.The aim of this thesis, 6
II. Principle and methods 15
II-1.Thermodynamic cycle, 16
II-2.Empirical Valence Bond model, 17
II-3.Free Energy Perturbation method, 19
III. Results 21
IV. Conclusion 24
References 26

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