Light-induced structural changes occur in the transmembrane helices of the Natronobacterium pharaonis HtrII transducer.
Light-induced structural changes occur in the transmembrane helices of the Natronobacterium pharaonis HtrII transducer.
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光诱导的结构变化发生在法老嗜碱杆菌 HtrII 传感器的跨膜螺旋中。
DOI：
10.1021/bi010985c
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发表时间：
2001
期刊：
影响因子：
2.9
通讯作者：
Spudich,JL
中科院分区：
文献类型：
--
作者：
Yang,CS;Spudich,JL
嗜盐杆菌HtrII（NpHtrII）传感器与其同源光敏感觉视紫红质受体NpSRII相互作用以介导趋光性反应。预测NpHtrII具有两个跨膜螺旋和一个大的胞质结构域，并形成同源二聚体。在其跨膜结构域中的38个位置处将单个半胱氨酸取代成工程化的无半胱氨酸NpHtrII。在有或没有NpSRII光活化的情况下测量单半胱氨酸突变体的氧化二硫键交联效率。在几个位置的快速交联率支持NpHtrII是一个二聚体时，在thehalobacteriumsalinarum膜功能性表达。第二个跨膜段（TM 2）中的13个位置显示出交联效率的显著光诱导增加，并且当建模为α-螺旋时，它们定义了横贯该段长度的单个面。该螺旋中显示光诱导效率降低的四个位置聚集在蛋白质的细胞质侧。其中一个单半胱氨酸突变体G83 C表现出失去了趋光性反应，对双突变体的分析表明，G83 C突变改变了NpHtrII的TM 2 − TM 2 '区域的暗结构。总之，结果揭示了NpHtrII的第二跨膜区段中的构象活性区域和沿TM 2长度的沿着面，该面在受体光活化后变得更可用于TM 2-TM 2 '交联。数据还确定，TM 2中的一个残基，Gly 83，对于维持NpHtrII的正确构象以用于从光活化受体到换能器的激酶结合区域的信号中继是至关重要的。
TheNatronobacterium pharaonisHtrII (NpHtrII) transducer interacts with its cognate photoactive sensory rhodopsin receptor, NpSRII, to mediate phototaxis responses. NpHtrII is predicted to have two transmembrane helices and a large cytoplasmic domain and to form a homodimer. Single cysteines were substituted into an engineered cysteine-less NpHtrII at 38 positions in its transmembrane domain. Oxidative disulfide cross-linking efficiencies of the monocysteine mutants were measured with or without photoactivation of NpSRII. The rapid cross-linking rates at several positions support that NpHtrII is a dimer when functionally expressed in theHalobacteriumsalinarummembrane. Thirteen positions in the second transmembrane segment (TM2) exhibited significant light-induced increases in cross-linking efficiency, and they define a single face traversing the length of the segment when modeled as an α-helix. Four positions in this helix showing light-induced decreases in efficiency are clustered on the cytoplasmic side of the protein. One of the monocysteine mutants, G83C, showed loss of phototaxis responses, and analysis of double mutants showed that the G83C mutation alters the dark structure of the TM2−TM2‘ region of NpHtrII. In summary, the results reveal conformationally active regions in the second transmembrane segment of NpHtrII and a face along the length of TM2 that becomes more available for TM2−TM2‘ cross-linking upon receptor photoactivation. The data also establish that one residue in TM2, Gly83, is critical for maintaining the proper conformation of NpHtrII for signal relay from the photoactivated receptor to the kinase-binding region of the transducer.