探討酵母菌Glutaminyl-tRNA synthetase 對於粒腺體功能之影響

Is glutaminyl-tRNA synthetase essential for the mitochondrial function of yeast?

葉曜榮(Yao-Jung Yeh)

指導教授 ： 王健家

國立中央大學/理學院/生命科學系/碩士(2008年)

摘要

Aminoacyl-tRNA (簡稱aa-tRNA) 的合成，對於蛋白質的生合成是非常重要的步驟，通常是藉由tRNA合成酶將胺基酸接到相對應的tRNA上。不過在Gln-tRNAGln的合成上卻是一個例外。在之前對於Saccharomyces cerevisiae的研究中指出，細胞質的glutaminyl-tRNA synthetase (簡稱GlnRS )是由GLN4基因所提供的，最近的研究發現細胞質的GlnRS會送到粒腺體中參與Gln-tRNAGln的合成。在本篇論文中我們利用細胞質的valyl-tRNA synthetase作為回報基因進一步找出其粒腺體標的訊號的範圍為包含第524 ~ 564號胺基酸，這段序列接近ATP的結合位 (KMSKS保留區域)，這是少數的例子，其粒腺體標的訊號位於在內部序列當中同時也與主要的催化區重疊，但是對於細胞質的GlnRS是怎樣利用一段非傳統的N端粒腺體標的訊號去作用，其機制還不清楚。在Schizosaccharomyces pombe中並沒有發現有粒腺體標的訊號存在，所以我們認為這樣的標的訊號並不是所有酵母菌都擁有的。藉由功能性互補實驗、回報基因測試，發現GlnRS對於粒腺體功能的維持並不是必要的，因此在粒腺體中可能同時存在兩條途徑去合成Gln-tRNAGln。

關鍵字

tRNA 合成酶 ； 酵母菌

並列摘要

Aminoacyl-tRNA (aa-tRNA) formation, an essential process in protein biosynthesis, is generally achieved by direct attachment of an amino acid to cognate tRNA by the aa-tRNA synthetases. An exception is Gln-tRNAGln synthesis. In Saccharomyces cerevisiae, the cytoplasmic glutaminyl-tRNA synthetase (GlnRS) activity is provided by the translational product of GLN4. Previous reports showed that this cytoplasmic GlnRS is also involved in mitochondrial Gln-tRNAGln synthesis. In this thesis, functional mapping using the cytoplasmic form of valyl-tRNA synthetase as the passenger protein identifies the peptide containing amino acids 524 ~ 564 of the enzyme as the mitochondrial targeting signal (MTS), which is close to the ATP-binding site (KMSKS conserved motif). This is one of the few examples, where MTS is embedded in the internal sequence, and is overlapped with the catalytic core domain of the enzyme. However, the detailed mechanism that enables GlnRS to be imported into mitochondria is not clear. In contrast, no MTS is found in Schizosaccharomyces pombe GlnRS, so this may be not a common feature for all yeast GlnRSs. Complementation tests further suggest that yeast GlnRS is not essential for mitochondrial function, and may serve as a redundant system for Gln-tRNAGln synthesis in mitochondria.

並列關鍵字

GlnRS ； Aminoacyl-tRNA synthetase ； AARS

參考文獻

張嘉珮 (2007)酵母菌使用罕見轉譯起始密碼的可能性探討。中央大學碩士

桂妤 (2006)一個雙重功能的酵母菌tRNA合成酶之研究。中央大學碩士

Hughes, T.R., Marton, M.J., Jones, A.R., Roberts, C.J., Stoughton, R., Armour, C.D., Bennett, H.A., Coffey, E., Dai, H., He, Y.D., et al. (2000) Functional discovery via a compendium of expression profiles. Cell 102: 109–126.

Simos, G., Segref, A., Fasiolo, F., Hellmuth, K., Shevchenko, A., Mann, M., and Hurt, E. C. (1996) The yeast protein Arc1p binds to tRNA and functions as a cofactor for the methionyl- and glutamyl-tRNA synthetases. EMBO J 15: 5437–5448

Cahuzac, B., Berthonneau, E., Birlirakis, N., Guittet, E. and Mirande, M. (2000) A recurrent RNA-binding domain is appended to eukaryotic aminoacyl-tRNA synthetases. EMBO J 19: 445-52.