分子動力模擬允許對原子層級的結構動力現象進行細節分析，例如生物領域中十分重要的分子間結合與辨識。參與(生物)分子間的結合作用力則可藉由基於熱力學定律的結合自由能計算做為評估。對於分子動態特性要點的構形柔軟度則可藉由計算構形熵，例如主成份分析的方法來估計。綜合這些方法，可對於僅止於靜態結構模型上難以說明的原子層級現象，提供合理的解釋。在這一篇我們呈現了一系列的典型分子動力模擬的結果，用於探討介於 (I) mithramycin二聚物與一段雙股DNA, (II) 數個胺基糖苷型抗生素與對應核糖體RNA A-site之寡核酸之間的結合辨識與作用力。這兩種抗生素都是由一個主要結構做為核心，並且於此處不同的碳原子位置衍生出數個糖環取代基所形成的。在第一部份(I)的研究，我們成功地建立了與實驗(核磁共振)結構及實驗結合親和力一致的動力模型，對結合作用力進行討論，並且發現此一對於DNA GC鹼基有專一結合性的抗生素對於具有兩個GC鹼基結合部位的雙股DNA(由十對核苷酸組成)之間是反協同性的結合。採用此一分子動力模擬的方法以及對這樣糖基連結的抗生素所修改的分子力場參數，在第二部分(II)的研究裡，我們對於數種胺基糖苷型抗生素以及一段能代表A-site的雙股RNA (A-site位於30S次單元上的16S核糖體RNA，30S對於細菌進行RNA轉譯形成蛋白質是相當重要的場所)之間的結合辨識及形成的水分子佔據型態進行比較。我們建立了數個具有合理的結合自由能的動態分子模型，並且與實驗數據間呈現良好的線性相關。分析在A-site上的U1406·U1495鹼基對周圍的水分子佔據部位，並區別出快速交換或者是與結合部位緊密結合的水分子。這樣的水分子佔據時間的分析在先前的研究中被提出對於循理性藥物設計是有幫助的。我們發現了在4,6-雙取代型抗生素 (tobramycin與kanamycin A) 的第三個糖環與G1405/U1406鹼基的磷酸基團上的氧原子間具有長時間的水分子佔據部位，這可能值得進一步探討作為這一類抗生素的循理性設計。

Molecular dynamics (MD) simulations allow detail analysis of structural dynamics of atomic–level phenomena such as binding recognition fundamental in Biology field. Binding interaction involved between (bio) –molecules can be evaluated by binding free energy calculation base on the law of thermodynamics. Conformational flexibility essential for investigating dynamic property can be estimated by calculating conformational entropy such as principal components analysis. Combination with these techniques can provide reasonable explanations for atomic–level phenomena that are difficult to explain on the basis of static models alone. Here we present the results of a series of conventional MD simulations on recognition and interaction between (I) a mithramycin dimer and a DNA duplex, (II) several aminoglycoside antibiotics and an oligonucleotide corresponding to rRNA A–site. Both kinds of antibiotics consist of a core structure where several sugar ring substitutions at different carbon positions. In part I of the study, we successfully built the dynamics model corresponding to the experimental structure and binding affinity, discussed the binding interaction, and found the cooperativity between this GC–specific DNA binding antibiotic and a decanucleotide duplex of two GC binding sites to be in an anticooperative manner. Following the MD protocol and modification of the force field parameters for this sugar–linked antibiotic, in part II of the study, we compared the binding recognition and hydration patterns between several aminoglycoside antibiotics and a RNA duplex corresponding to the aminoacyl–tRNA decoding site (A–site) of the 16S rRNA on the 30S subunit which is a crucial component of the bacterial translational machinery. We have built several dynamic models with reasonable binding free energies showing good linear correlation with the experimental data. The hydration sites around the U1406·U1495 pair in the A–site were analyzed to distinguish tightly bound water molecules from fast–exchanging ones which has been suggested to be useful for rational drug design. We found that the hydration sites with long residence time identified between ring III of two 4,6–disubstituted antibiotics (tobramycin and kanamycin A) and phosphate oxygen atoms of G1405/U1406 may be worthy of further exploration for rational design of this kind.

Chapter 1 Introduction -------------------------------------------------------- 6
1.1 General information --------------------------------------------------------------------------------- 7
1.2 Research background of (I) mithramycin dimer and DNA duplex ---------------------------- 10
1.3 Research background of (II) aminoglycoside antibiotics and rRNA A–site ----------------- 13
1.4 Incentive and aims ----------------------------------------------------------------------------------- 17
1.5 Figures and tables ----------------------------------------------------------------------------------- 20

Chapter 2 Materials and Methods ---------------------------------------------------------------------- 24
2.1 General introduction of conventional MD simulations ----------------------------------------- 25
2.2 Materials & modeling ------------------------------------------------------------------------------- 28
2.3 Methods: MD simulations -------------------------------------------------------------------------- 31
2.4 Methods: Binding free energy & conformational entropy calculation ----------------------- 32
2.5 Methods: Essential dynamics & PCA (principal component analysis) ----------------------- 35
2.6 Methods: Hydration analysis ----------------------------------------------------------------------- 37
2.7 Figures and tables ----------------------------------------------------------------------------------- 39

Chapter 3 Results and Discussion: (I) Mithramycin dimer and DNA duplex ----------------- 40
3.1 General inspection of MD simulations ------------------------------------------------------------ 41
3.2 Thermodynamic characterization of the binding event ----------------------------------------- 44
3.3 Cooperativity characterization: Entropic dissection and PCA analysis ----------------------- 46
3.4 Conclusion -------------------------------------------------------------------------------------------- 48
3.5 Figures and tables ----------------------------------------------------------------------------------- 49

Chapter 4 Results and Discussion: (II) Aminoglycoside antibiotics and rRNA A–site ------- 60
4.1 General inspection of MD simulations ------------------------------------------------------------ 61
4.2 Thermodynamic and essential dynamics characterization of binding affinities and
_____antibacterial activities ------------------------------------------------------------------------------- 63
4.3 Hydration sites and binding recognition ---------------------------------------------------------- 67
4.4 Conclusion -------------------------------------------------------------------------------------------- 74
4.5 Figures and tables ----------------------------------------------------------------------------------- 75

Chapter 5 Conclusion ------------------------------------------------------------------------------------- 90

Reference ---------------------------------------------------------------------------------------------------- 92

Appendix I. Publication list ------------------------------------------------------------------------------- 98
Appendix II. Abbreviation list ---------------------------------------------------------------------------- 99

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