Research Center for Molecular Mechanisms of Aging and Age-Related Diseases, Moscow Institute of Physics and Technology, 141700 Dolgoprudny, Russia.
Institute of Personalized Medicine, Sechenov University, 119146 Moscow, Russia.
Int J Mol Sci. 2020 Apr 28;21(9):3110. doi: 10.3390/ijms21093110.
Membrane-embedded sensor histidine kinases (HKs) and chemoreceptors are used ubiquitously by bacteria and archaea to percept the environment, and are often crucial for their survival and pathogenicity. The proteins can transmit the signal from the sensor domain to the catalytic kinase domain reliably over the span of several hundreds of angstroms, and regulate the activity of the cognate response regulator proteins, with which they form two-component signaling systems (TCSs). Several mechanisms of transmembrane signal transduction in TCS receptors have been proposed, dubbed (swinging) piston, helical rotation, and diagonal scissoring. Yet, despite decades of studies, there is no consensus on whether these mechanisms are common for all TCS receptors. Here, we extend our previous work on nitrate/nitrite sensor kinase NarQ. We determined a crystallographic structure of the sensor-TM-HAMP fragment of the R50S mutant, which, unexpectedly, was found in a ligand-bound-like conformation, despite an inability to bind nitrate. Subsequently, we reanalyzed the structures of the ligand-free and ligand-bound NarQ and NarX sensor domains, and conducted extensive molecular dynamics simulations of ligand-free and ligand-bound wild type and mutated NarQ. Based on the data, we show that binding of nitrate to NarQ causes, first and foremost, helical rotation and diagonal scissoring of the α-helices at the core of the sensor domain. These conformational changes are accompanied by a subtle piston-like motion, which is amplified by a switch in the secondary structure of the linker between the sensor and TM domains. We conclude that helical rotation, diagonal scissoring, and piston are simply different degrees of freedom in coiled-coil proteins and are not mutually exclusive in NarQ, and likely in other nitrate sensors and TCS proteins as well.
膜嵌入传感器组氨酸激酶 (HKs) 和化学感受器被细菌和古菌广泛用于感知环境，通常对它们的生存和致病性至关重要。这些蛋白质可以在数百埃的跨度内可靠地将信号从传感器结构域传递到催化激酶结构域，并调节同源响应调节剂蛋白的活性，它们与这些蛋白形成了双组分信号系统 (TCS)。已经提出了 TCS 受体中几种跨膜信号转导机制，分别称为 (摆动) 活塞、螺旋旋转和对角切割。然而，尽管经过了几十年的研究，对于这些机制是否对所有 TCS 受体都通用，仍然没有共识。在这里，我们扩展了之前关于硝酸盐/亚硝酸盐传感器激酶 NarQ 的工作。我们确定了 R50S 突变体的传感器-TM-HAMP 片段的晶体结构，出乎意料的是，尽管不能结合硝酸盐，但它被发现处于配体结合样构象。随后，我们重新分析了配体自由和配体结合的 NarQ 和 NarX 传感器结构域的结构，并对配体自由和配体结合的野生型和突变型 NarQ 进行了广泛的分子动力学模拟。基于这些数据，我们表明，硝酸盐与 NarQ 的结合首先引起传感器结构域核心处的α-螺旋的螺旋旋转和对角切割。这些构象变化伴随着活塞样运动的微妙变化，这种变化被传感器和 TM 结构域之间的连接体二级结构的转变放大。我们得出的结论是，螺旋旋转、对角切割和活塞只是卷曲螺旋蛋白的不同自由度，在 NarQ 中并不相互排斥，而且在其他硝酸盐传感器和 TCS 蛋白中也可能如此。
