Electronic and Protein Structural Dynamics of a Photosensory Histidine Kinase
Electronic and Protein Structural Dynamics of a Photosensory Histidine Kinase
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DOI：
10.1021/bi100527a
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发表时间：
2010-06-15
期刊：
影响因子：
2.9
通讯作者：
Crosson, Sean
中科院分区：
文献类型：
--
作者：
Alexandre, Maxime T. A.;Purcell, Erin B.;Crosson, Sean
新月柄杆菌编码一种双组分信号蛋白LovK，其含有N-末端光敏LOV结构域，该结构域与C-末端组氨酸激酶偶联。LovK结合黄素辅因子，经历可逆的光循环，并响应可见光显示调节的ATP酶和自磷酸化活性。飞秒至纳秒可见吸收光谱表明全长LovK和孤立的LOV域之间的一致性，在光依赖性半胱氨酰-C4（a）加合物形成和破裂的机制和动力学，而稳态吸收和荧光线窄化（FLN）光谱揭示LovK黄素辅因子的电子结构的独特功能。与其他传感器组氨酸激酶一致，ATP以微摩尔亲和力特异性结合LovK。然而，ATP结合到LovK的组氨酸激酶结构域上，通过差示傅里叶变换红外（FTIR）光谱法评估，对全局蛋白质结构没有明显影响。半胱氨酰加合物的形成导致LovK的结构仅发生微小变化，如通过差示FTIR所确定的。这项研究提供了深入了解LOV介导的信号转导的背景下，全长组氨酸激酶的结构基础。特别是，这些数据提供了证据的模型，其中小的变化，在三级/四级结构的LovK，在N-末端LOV传感域的光子检测触发，足以调节组氨酸激酶活性。
The bacterium Caulobucter crescent us encodes a two-component signaling protein, LovK, that contains an N-terminal photosensory LOV domain coupled to a C-terminal histidine kinase. LovK binds a flavin cofactor, undergoes a reversible photocycle, and displays regulated ATPase and autophosphorylation activity in response to visible light. Femtosecond to nanosecond visible absorption spectroscopy demonstrates congruence between full-length LovK and isolated LOV domains in the mechanism and kinetics of light-dependent cysteinyl-C4(a) adduct formation and rupture, while steady-state absorption and fluorescence line narrowing (FLN) spectroscopies reveal unique features in the electronic structure of the LovK flavin cofactor. In agreement with other sensor histidine kinases, ATP binds specifically to LovK with micromolar affinity. However, ATP binding to the histidine kinase domain of LovK has no apparent effect on global protein structure as assessed by differential Fourier transform infrared (FTIR) spectroscopy. Cysteinyl adduct formation results in only minor changes in the structure of LovK as determined by differential FTIR. This study provides insight into the structural underpinnings of LOV-mediated signal transduction in the context of a full-length histidine kinase. In particular, the data provide evidence for a model in which small changes in the tertiary/quaternary structure of LovK, as triggered by photon detection in the N-terminal LOV sensory domain, are sufficient to regulate histidine kinase activity.