Michalodimitrakis Konstantinos M, Sourjik Victor, Serrano Luis
EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany.
Mol Microbiol. 2005 Oct;58(1):257-66. doi: 10.1111/j.1365-2958.2005.04821.x.
Taz is a chimeric receptor consisting of the periplasmic, transmembrane and most of the HAMP linker domains of the Escherichia coli aspartate receptor (Tar(Ec)) and the cytoplasmic signalling domain of the E. coli osmosensor EnvZ. Aspartate is one of several attractant ligands normally sensed by Tar and it interacts with Taz to induce OmpR-dependent transcription from the ompC promoter--albeit with reduced sensitivity relative to the chemotactic response it evokes via Tar. By combining Taz with a reporter system that expresses green fluorescent protein (GFP) from the ompC promoter, we were able to examine the interaction of Taz with all 20 natural amino acids. Some amino acids (Leu, Met, Val and Ser) reduced GFP expression, which in the case of leucine is likely attributed to a direct effect on the receptor, rather than an indirect effect through the leucine responsive protein (Lrp). Surprisingly, amino acids like Met and Ser--which are also attractants for Tar--'inhibited' Taz. Moreover, Taz exhibits a higher sensitivity to Leu compared with Asp, which is the inverse of Tar. Our results show the exquisite sensitivity of chemotactic receptors. Small conformational changes induced by making the chimera may have changed the way it responds to different amino acids.
Taz是一种嵌合受体，由大肠杆菌天冬氨酸受体（Tar(Ec)）的周质、跨膜和大部分HAMP连接域以及大肠杆菌渗透压感受器EnvZ的细胞质信号域组成。天冬氨酸是Tar通常感知的几种吸引配体之一，它与Taz相互作用，诱导来自ompC启动子的OmpR依赖性转录——尽管相对于它通过Tar引发的趋化反应，其敏感性有所降低。通过将Taz与一个从ompC启动子表达绿色荧光蛋白（GFP）的报告系统相结合，我们能够研究Taz与所有20种天然氨基酸的相互作用。一些氨基酸（亮氨酸、甲硫氨酸、缬氨酸和丝氨酸）降低了GFP的表达，就亮氨酸而言，这可能归因于对受体的直接作用，而不是通过亮氨酸反应蛋白（Lrp）的间接作用。令人惊讶的是，像甲硫氨酸和丝氨酸这样的氨基酸——它们也是Tar的吸引剂——“抑制”了Taz。此外，与天冬氨酸相比，Taz对亮氨酸表现出更高的敏感性，这与Tar相反。我们的结果显示了趋化受体的极高敏感性。构建嵌合体所引起的微小构象变化可能改变了它对不同氨基酸的反应方式。
