Shibagaki Nakako, Grossman Arthur R
Department of Plant Biology, The Carnegie Institution, Stanford, California 94305, USA.
J Biol Chem. 2006 Aug 11;281(32):22964-73. doi: 10.1074/jbc.M603462200. Epub 2006 Jun 5.
Sulfate transporters in plants represent a family of proteins containing transmembrane domains that constitute the catalytic part of the protein and a short linking region that joins this catalytic moiety with a C-terminal STAS domain. The STAS domain resembles an anti-sigma factor antagonist of Bacillus subtilis, which is one distinguishing feature of the SLC26 transporter family; this family includes transporters for sulfate and other anions such as iodide and carbonate. Recent work has demonstrated that this domain is critical for the activity of Arabidopsis thaliana sulfate transporters, and specific lesions in this domain, or the exchange of STAS domains between different sulfate transporters, can severely impair transport activity. In this work we generated a Saccharomyces cerevisiae expression library of the A. thaliana Sultr1;2 gene with random mutations in the linking region-STAS domain and identified STAS domain lesions that altered Sultr1;2 biogenesis and/or function. A number of mutations in the beta-sheet that forms the core of the STAS domain prevented intracellular accumulation of Sultr1;2. In contrast, the linking region and one surface of the STAS domain containing N termini of the first and second alpha-helices have a number of amino acids critical for the function of the protein; mutations in these regions still allow protein accumulation in the plasma membrane, but the protein is no longer capable of efficiently transporting sulfate into cells. These results suggest that the STAS domain is critical for both the activity and biosynthesis/stability of the transporter, and that STAS sub-domains correlate with these specific functions.
植物中的硫酸盐转运蛋白代表了一类蛋白质家族，它们含有构成蛋白质催化部分的跨膜结构域以及一个短连接区域，该区域将这个催化部分与C端的STAS结构域相连。STAS结构域类似于枯草芽孢杆菌的抗σ因子拮抗剂，这是SLC26转运蛋白家族的一个显著特征；该家族包括硫酸盐和其他阴离子（如碘化物和碳酸盐）的转运蛋白。最近的研究表明，这个结构域对于拟南芥硫酸盐转运蛋白的活性至关重要，该结构域中的特定损伤，或者不同硫酸盐转运蛋白之间STAS结构域的交换，会严重损害转运活性。在这项工作中，我们构建了一个拟南芥Sultr1;2基因在连接区域-STAS结构域带有随机突变的酿酒酵母表达文库，并鉴定出改变Sultr1;2生物合成和/或功能的STAS结构域损伤。形成STAS结构域核心的β折叠中的一些突变阻止了Sultr1;2在细胞内的积累。相反，连接区域以及包含第一和第二α螺旋N端的STAS结构域的一个表面有许多对该蛋白质功能至关重要的氨基酸；这些区域的突变仍然允许蛋白质在质膜中积累，但该蛋白质不再能够有效地将硫酸盐转运到细胞中。这些结果表明，STAS结构域对于转运蛋白的活性以及生物合成/稳定性都至关重要，并且STAS亚结构域与这些特定功能相关。
