Friden P, Schimmel P
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Mol Cell Biol. 1987 Aug;7(8):2708-17. doi: 10.1128/mcb.7.8.2708-2717.1987.
Although the majority of genes for amino acid biosynthesis which have been examined are under general amino acid control, LEU1 and LEU2 of Saccharomyces cerevisiae respond specifically to leucine. We report here an analysis of LEU3, a putative leucine-specific regulatory locus. We show that LEU3 is necessary for expression of wild-type levels of LEU1- and LEU2-specific RNAs and, further, that the levels of LEU4-specific transcripts are also affected by LEU3. We cloned LEU3 and showed by DNA sequence analysis that it contained an open reading frame of 886 amino acids. A striking feature of the predicted LEU3 protein was a cluster of acidic amino acids (19 of 20) located in the C-terminal half of the coding region. The protein also had a repeated cysteine motif which was conserved in a number of other yeast proteins implicated in gene regulation. We show that whole-cell extracts contained a LEU3-dependent DNA-binding activity that interacted with the 5' region of LEU2. Subdivision of the LEU2 5' region established that the LEU3-dependent DNA-binding activity interacted with the segment which had the previously reported homology with LEU1.
尽管已检测的大多数氨基酸生物合成基因都受一般氨基酸调控，但酿酒酵母的LEU1和LEU2基因对亮氨酸有特异性反应。我们在此报告对LEU3（一个假定的亮氨酸特异性调控位点）的分析。我们发现LEU3对于野生型水平的LEU1和LEU2特异性RNA的表达是必需的，而且LEU4特异性转录本的水平也受LEU3影响。我们克隆了LEU3，并通过DNA序列分析表明它包含一个886个氨基酸的开放阅读框。预测的LEU3蛋白的一个显著特征是位于编码区C端一半的一簇酸性氨基酸（20个中有19个）。该蛋白还有一个重复的半胱氨酸基序，在许多其他与基因调控有关的酵母蛋白中是保守的。我们发现全细胞提取物中含有一种依赖LEU3的DNA结合活性，它与LEU2的5'区域相互作用。对LEU2 5'区域的细分表明，依赖LEU3的DNA结合活性与先前报道的与LEU1有同源性的片段相互作用。
