Title page for 952204014

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Student Number 952204014
Author Yi-Yi Kuo(郭亦亦)
Author's Email Address No Public.
Statistics This thesis had been viewed 1550 times. Download 630 times.
Department Life Science
Year 2007
Semester 2
Degree Master
Type of Document Master's Thesis
Language zh-TW.Big5 Chinese
Title Translation initiation of HTS1 in yeast
Date of Defense 2008-06-25
Page Count 78
  • Histidyl-tRNA synthetase
  • translation initiation
  • Abstract In yeast, there are two sets of aminoacyl-tRNA synthetases, one localized in the cytoplasm and the other in the mitochondria. Most of the mitochondrial tRNA synthetases are encoded by nuclear genes distinct from those encoding their cytoplasmic counterparts. However, some mitochondrial tRNA synthetases are encoded by the same genes that code for their cytoplasmic homologues. For example, the cytoplasmic and mitochondrial histidyl-tRNA synthetases of Saccharomyces cerevisiae (ScHisRS) are encoded by the same nuclear gene, HTS1, through alternative initiation of translation from two in-frame AUG codons. The gene specifies two messages, the longer one with two 5’-end in-frame AUGs and the short one with only the second AUG. In this study, we showed that the HisRS genes of Schizosaccharomyces pombe、Candida albicans and Aspergillus fumigatus have only one gene that also contain two in-frame AUG initiation codons. The mitochodrial and cytosolic forms are translated from the first and second AUG initiation codons, respectively. Besides, we found that ScHisRS had a lysine-rich N-terminal polypeptide extension of 34 residues, which was absent from its E. coli counterpart. Attachment of this appended domain to E. coli glutaminyl-tRNA synthetase did not enhance the enzyme’s tRNA-binding and aminoacylation activity toward yeast tRNAs. Deletion of the appended domain of ScHisRS had little effect on the enzyme’s complementation activity in vivo. These results suggest that the appended domain is not essential for the cytoplasmic and mitochondrial functions of ScHisRS in vivo.
    Table of Content 目錄
    第一章 緒論1
    1.1 Aminoacyl-tRNA synthetases(aaRSs)的簡介1
    1.1.1 aaRSs的生化功能1
    1.1.2 aaRSs 的分類2
    1.2 原核與真核細胞在蛋白質合成轉譯方式的差異3
    1.2.1 原核與真核細胞內蛋白質的合成3
    1.3 Histidyl-tRNA synthetase(HisRS)的簡介5
    1.3.1 HisRS的生化功能5
    1.3.2 HTS1基因的表現6
    第二章 材料與方法9
    2.5點突變(Site-directed Mutagenesis)13
    2.6 hts1剔除株的製備14
    2.10 SDS-PAGE之蛋白質分子量分析19
    2.11西方點墨法(western blotting)20
    第三章 結果24
    3.3 ScHTS1基因的表現26
    3.6 ScHisRS的附加區段是否具鍵結tRNA的能力32
    3.7 ScHisRS附加區段對其細胞質與粒腺體酵素的重要性33
    第四章 討論35
    4.2 ScHisRS附加區段的特性37
    第五章 參考文獻39

    圖十二、酵母菌及大腸桿菌 HisRS 的比較54
    圖十三、利用E. coli GlnRS的融合蛋白進行互補性試驗55
    圖十四、利用E. coli GlnRS的融合蛋白進行互補性試驗56
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  • Chien-Chia Wang(王健家)
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    Date of Submission 2008-07-02

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