Yamada Seiji, Akiyama Shuji, Sugimoto Hiroshi, Kumita Hideyuki, Ito Kazuki, Fujisawa Tetsuro, Nakamura Hiro, Shiro Yoshitsugu
Biometal Science Laboratory, RIKEN SPring-8 Center, Harima Institute, Hyogo 679-5148, Japan.
J Mol Biol. 2006 Sep 8;362(1):123-39. doi: 10.1016/j.jmb.2006.07.012. Epub 2006 Jul 15.
The structure of a histidine kinase (ThkA) complexed with a response regulator (TrrA) in the two-component regulatory system from hyperthermophile Thermotoga maritima was determined by a combination of X-ray crystallography at a resolution of 4.2 A and small-angle X-ray scattering (SAXS). The boundary of the three component domains (PAS-sensor, dimerization and catalytic domains) of ThkA and the bound TrrA molecule were unambiguously assigned in the electron density map at 4.2 A resolution. ThkA forms a dimer with crystallographic 2-fold symmetry and two monomeric TrrAs bind to the ThkA dimer. SAXS experiments also confirmed this association state in solution and specific binding between ThkA and TrrA (Kd=8.2x10(-11) M(-2)). The association interface between ThkA and TrrA contains the phosphotransfer His residue in the ThkA, indicative of an efficient receipt of the phosphoryl group. One Per-Arnt-Sim (PAS) domain does not interact with the other PAS domain, but with the catalytic domain of the same polypeptide chain and with one TrrA molecule. Observed inter-domain and inter-molecular interactions reveal a definite pathway of signal transduction in the kinase/regulator complex. In addition, we propose a responsible role of TrrA for the feedback regulation of sensing and/or kinase activities of ThkA.
嗜热栖热袍菌双组分调节系统中与响应调节蛋白（TrrA）复合的组氨酸激酶（ThkA）的结构，是通过分辨率为4.2 Å的X射线晶体学和小角X射线散射（SAXS）相结合的方法确定的。在4.2 Å分辨率的电子密度图中，明确确定了ThkA的三个组分结构域（PAS传感器、二聚化和催化结构域）以及结合的TrrA分子的边界。ThkA形成具有晶体学2倍对称性的二聚体，两个单体TrrA与ThkA二聚体结合。SAXS实验也证实了溶液中的这种结合状态以及ThkA与TrrA之间的特异性结合（解离常数Kd = 8.2×10⁻¹¹ M⁻²）。ThkA与TrrA之间的结合界面包含ThkA中的磷酸转移组氨酸残基，这表明磷酸基团能够有效接收。一个Per-Arnt-Sim（PAS）结构域不与另一个PAS结构域相互作用，而是与同一条多肽链的催化结构域以及一个TrrA分子相互作用。观察到的结构域间和分子间相互作用揭示了激酶/调节蛋白复合物中信号转导的明确途径。此外，我们提出TrrA在ThkA的传感和/或激酶活性的反馈调节中发挥作用。
