YEAST AMINO ACID PERMEASES

酿酒酵母（Saccharomycescerevisiae）具有多层质膜， 将氨基酸运输到细胞中的渗透酶。这些氨基酸可以 或者被细胞代谢并用作主要氮源 用于生长或直接用于蛋白质合成。的控制 氨基酸的摄取是复杂的，并且还没有被很好地理解。特别研究 运输系统由于过程的冗余而变得复杂， 大多数渗透酶可以以不同的效率输送几种底物 并且大多数氨基酸可以通过一种以上的通透酶输入。失去一 单个通透酶对实验室中生长的细胞几乎没有影响 条件下，因此在一般实验室使用的“野生型”菌株是高度 渗透酶含量可变。这也使运输研究复杂化 在不同的菌株背景下。 已克隆和测序的已知通透酶基因显示， 与某些绝对保守的特征具有相当大的同源性。因此如 酵母基因组计划继续鉴定新的基因， 正在鉴定氨基酸渗透酶。在这个项目中， 这些未知的渗透酶的范围和特异性将通过以下方式阐明： 系统性缺失通透酶基因，然后仔细分析 氨基酸摄取的变化增加了对氨基酸类似物的敏感性， 产生的缺失菌株。当所有的渗透酶 特定氨基酸的转运已经被详细描述， 可以完成运输的动力学分析。此外，菌株中， 渗透酶的含量是有保证的，可以建造，这将是 提供给酵母界。在另一组实验中， 渗透酶分子的高度保守区域的功能将是 研究了这些地区很可能在以下方面发挥重要作用： 氨基酸转运和调节或渗透酶的定位 通过分泌途径到达质膜。这些可能性 将通过这些区域的定向诱变进行系统性测试， 两种代表性的透水性。

The yeast, Saccharomyces cerevisiae, has multiple plasma membrane permeases that transport amino acids into the cell. These amino acids can either be metabolized and used by the cell as a principal nitrogen source for growth or be utilized directly in protein synthesis. The control of amino acid uptake is complex and not well understood. Study of particular transport systems has been complicated by the redundancy of the process, most permeases can transport several substrates with varying efficiency and most amino acids can be imported by more than one permease. Loss of an individual permease has little, or no, effect on cells growing in lab conditions and thus the 'wild-type' strains in general lab use are highly variable in permease content. This also complicates studies of transport in different strain backgrounds. The known permease genes which have been cloned and sequenced show considerable homology with certain absolutely conserved features. Thus, as the Yeast Genome Project proceeds to identify new genes, more 'putative' amino acid permeases are being identified. In this project, the substrate range and specificity of these unknown permeases will be elucidated by systematic deletion of the permease genes followed by careful analysis of changes in the amino acid uptake add sensitivity to amino acid analogs of the resulting deletion strains. When all the permeases responsible for transport of a particular amino acid have been characterized a detailed kinetic analysis of transport can be completed. Also, strains in which the permease content is assured can be constructed which will be made available to the yeast community. In a separate set of experiments, the function of the highly conserved regions of the permease molecules will be investigated. It is likely that these regions will either be important in amino acid transport and regulation or in localization of the permease through the secretory pathway to the plasma membrane. These possibilities will be systematically tested by directed mutagenesis of these regions in two representative permeases.