本文旨在探討陰道滴蟲裡 Myb2 蛋白(tvMyb2)以及豹蛙的 onconase 蛋白的結構與動態。在生物界中 Myb 蛋白為一個龐大的家族有著有著各式各樣的功能。多數的 Myb 蛋白都是屬於可以與去氧核醣核酸結合的轉譯因子。本篇研究了陰道滴蟲內的類Myb蛋白的結構與動態，tvMyb2 蛋白參與了鐵誘導的 ap65-1 基因的轉譯機制。Myb2 中具有最小結構的單位被鑑定為40-156 的殘基，並保有與全長蛋白相近的啟動子結合能力。類 Myb 蛋白的轉譯因子 tvMyb2 的結構與動態將被深入探討，tvMyb2 蛋白參與了鐵誘導的 ap65-1 基因的轉譯機制。tvMyb2 裡最小具結構性的單位被鑑定為 40-156 的殘基，這片段保有跟全長蛋白相近的啟動子結合能力。未結合的 tvMyb2 40-156 具有較鬆散的構型。一旦與啟動子結合後，其構型將會重新排列並形成較穩定的結構。tvMyb2 與啟動子的複合物結晶結構顯示出 tvMyb2 辨識啟動子中 5’-a/gACGAT-3’這段特異性的序列。此外，位於 R2 模段上游的 Lys49 殘基亦參與了這段特異性序列的辨識。特別的是，與另一個 tvMyb1 相較，tvMyb2 40-156 採用跟 tvMyb1 35-141 /MRE-1 複合物的 HADDOCK模型結構相反的方位與啟動子結合。Onconase 蛋白同時亦被稱為 ranpirnase 或 P-30 蛋白，過去研究中，此蛋白主要由豹蛙蛙卵的萃取物中純化而得，並在各項實驗中發現其具有抗癌的特性，另在治療腫瘤方面已經進展到第三期的臨床試驗。醣基化作用對 onconase 蛋白的結構與動態上的影響將在此被探討。結果顯示，醣基對於 onconase 蛋白的結構與動態的影響甚微。再者，使用 CPMG-based 弛豫色散的實驗方法可觀察到當 onconase 蛋白跟核酸結合後，會產生新的微秒尺度的運動。更深入的檢視此運動與配體結合動力學的關係，將有助於了解蛋白質動態在催化機制中扮演的角色以及藥物設計上的應用。

In this thesis we investigated the structure and dynamics of Myb2 protein from Trichomonas vaginalis (tvMyb2) and onconase from Rana pipiens. The Myb proteins consist of a large family of proteins with diverse functions across the biological kingdom. Most Myb proteins function as transcriptional factors by binding to DNA. Here we investigated the structure and dynamics of Myb-like transcriptional factor, tvMyb2, from Trichomonas vaginalis involved in iron-inducible transcription of ap65-1 gene. We first defined a fragment of tvMyb2 comprising residues 40–156 (tvMyb2 40–156 ) as the minimum structural unit that retains near full binding affinity with the promoter DNAs. The DNA-free tvMyb2 40–156 has a flexible and open conformation. Upon binding to the promoter DNA elements, tvMyb2 40–156 undergoes significant conformational re-arrangement and structure stabilization. Crystal structures of tvMyb2 40–156 in complex with promoter element-containing DNA oligomers showed that 5’-a/gACGAT-3’ is the specific base sequence recognized by tvMyb2 40–156 , which does not fully conform to that
of the Myb binding site sequence. Furthermore, Lys49, upstream of the R2 motif (52–102), also participates in specific DNA sequence recognition. Intriguingly, tvMyb2 40–156 binds to the promoter elements in an orientation opposite to that proposed in the HADDOCK model of the tvMyb1 35–141 /MRE-1-MRE-2r complex. Onconase (rONC), also known as ranpirnase or P-30 protein, initially purified from extracts of Rana pipiens oocytes and early embryos, exhibits anticancer activity both in vitro and in vivo and is in phase III clinical trials for tumor therapy. We examined the effect of glycosylation on the structural and dynamic of onconase. The results showed that the glycan has little effect on the overall structure and dynamics of onconase. Moreover, we employed CPMG-based relaxation dispersion method and observed a novel ms time-scale protein motion elicited by nucleic acid binding. Further examination of the correlation between this collective
motion and ligand binding kinetics may provide better understanding the role of dynamics in the catalytic mechanism and drug design of onconase.

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