- 學位論文
側鏈修飾對Tat衍生胜辨認核糖核酸與細胞穿透之影響
Effect of Side Chain Modifications of Tat-derived Peptides on RNA Recognition and Cellular Uptake
若您是本文的作者，可授權文章由華藝線上圖書館中協助推廣。
摘要
愛滋病是由人類免疫缺乏病毒（HIV）所導致，而關於病毒以及治療方式的研究也在近幾十年來不斷有新結果。其中一種可能的治療方式是以干擾Tat蛋白與TAR RNA之間的交互作用力，進而影響病毒的繁衍；這樣的構思已應用在先前的研究中並設計出一種以Tat蛋白為模版的胜肽—TatC4。TatC4具有和TAR RNA的高度結合力，但其專一性不足，造成TatC4抑制Tat-TAR交互作用的能力不如預期。所以本次研究是希望透過更改胺基酸側鏈的長度與電荷，可以增進Tat衍生胜肽對TAR RNA的專一性。 實驗方式是以凝膠電泳（Electrophoretic mobility shift assay）來代表胜肽與TAR RNA的結合度；根據Tat衍生胜肽與對照組的比較，新設計出來的胜肽中有部分衍生胜肽對TAR RNA具有比較高的專一性。同時也使用流式細胞儀，針對與TAR RNA有較高親和性與專一性的Tat衍生胜肽進行細胞穿膜實驗，結果顯示Tat衍生胜肽皆具有較高的穿膜能力。由此可知，側鏈長度與電荷的更動不只影響胜肽與TAR RNA的結合度，也同時影響胜肽的細胞穿膜能力。
並列摘要
HIV (Human Immunodeficiency virus) has been studied for decades, and research on developing drugs for treating HIV infection has been ongoing. Interfering with the interaction between TAR RNA and Tat-protein might be a therapeutically viable strategy, which gave rise to the development of peptide TatC4 to bind TAR RNA in the Cheng lab (Org. Biomol. Chem. 2015, 13, 11096). Unfortunately, the ability of TatC4 to inhibit Tat-TAR mediated protein production didn’t meet our expectations. The results suggested that the specificity for binding TAR remained to be improved. This lack of specificity could perhaps be remediated by simultaneously changing the side chain length and charge of the Arg residues in the Tat-derived peptide. Accordingly, a series of Tat-derived peptides with Arg side chain modifications have been synthesized by solid phase methods and purified. The binding of the peptides to TAR RNA was investigated by EMSA (electrophoretic mobility shift assay). Some of the Tat-derived peptides showed enhanced TAR RNA binding affinity and specificity. Four peptides with higher binding specificity or affinity were chosen for cellular uptake studies, which all showed higher cellular uptake compared to the native peptide. Overall, the research should contribute to developing molecules to treat HIV infections.
參考文獻
Chapter 1
1. Crick, F. Central dogma of molecular biology. Nature 1970, 227, 561-563.
2. Davey, M. J.; O'Donnell, M. Mechanisms of DNA replication. Curr. Opin. Chem. Biol. 2000, 4, 581-586.
3. Conaway, J. W.; Shilatifard, A.; Dvir, A.; Conaway, R. C. Control of elongation by RNA polymerase II. Trends Biochem. Sci. 2000, 25, 375-380.
4. Ramakrishnan, V. Ribosome structure and the mechanism of translation. Cell 2002, 108, 557-572.