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。 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.