Chen Yanling, Gollnick Paul
Department of Biological Sciences, State University of New York, Buffalo, New York 14260-1300, USA.
J Mol Biol. 2008 Apr 11;377(5):1529-43. doi: 10.1016/j.jmb.2008.02.015. Epub 2008 Feb 15.
The trp RNA-binding attenuation protein (TRAP) regulates expression of the tryptophan biosynthetic (trp) genes in response to changes in intracellular levels of free l-tryptophan in many Gram-positive bacteria. When activated by binding tryptophan, TRAP binds to the mRNAs of several genes involved in tryptophan metabolism, and down-regulates transcription or translation of these genes. Anti-TRAP (AT) is an antagonist of TRAP that binds to tryptophan-activated TRAP and prevents it from binding to its RNA targets, and thereby up-regulates trp gene expression. The crystal structure shows that AT is a cone-shaped trimer (AT(3)) with the N-terminal residues of the three subunits assembled at the apex of the cone and that these trimers can further assemble into a dodecameric (AT(12)) structure. Using alanine-scanning mutagenesis we found four residues, all located on the "top" region of AT(3), that are essential for binding to TRAP. Fluorescent labeling experiments further suggest that the top region of AT is in close juxtaposition to TRAP in the AT-TRAP complex. In vivo studies confirmed the importance of these residues on the top of AT in regulating TRAP mediated gene regulation.
色氨酸RNA结合衰减蛋白（TRAP）可响应许多革兰氏阳性菌细胞内游离L-色氨酸水平的变化，调节色氨酸生物合成（trp）基因的表达。当通过结合色氨酸被激活时，TRAP会与几个参与色氨酸代谢的基因的mRNA结合，并下调这些基因的转录或翻译。抗TRAP（AT）是TRAP的拮抗剂，它与色氨酸激活的TRAP结合，阻止其与RNA靶标结合，从而上调trp基因的表达。晶体结构表明，AT是一种锥形三聚体（AT(3)），三个亚基的N端残基聚集在锥顶，并且这些三聚体可以进一步组装成十二聚体（AT(12)）结构。通过丙氨酸扫描诱变，我们发现了四个残基，它们都位于AT(3)的“顶部”区域，对于与TRAP结合至关重要。荧光标记实验进一步表明，在AT-TRAP复合物中，AT的顶部区域与TRAP紧密相邻。体内研究证实了AT顶部这些残基在调节TRAP介导的基因调控中的重要性。
