Structural basis of histidine kinase autophosphorylation deduced by integrating genomics, molecular dynamics, and mutagenesis
Structural basis of histidine kinase autophosphorylation deduced by integrating genomics, molecular dynamics, and mutagenesis
复制标题
DOI：
10.1073/pnas.1201301109
复制
发表时间：
2012-06-26
影响因子：
11.1
通讯作者：
Szurmant, Hendrik
中科院分区：
文献类型：
--
作者：
Dago, Angel E.;Schug, Alexander;Szurmant, Hendrik
信号转导蛋白，如细菌传感器组氨酸激酶，旨在多种构象之间的转换，往往是由不稳定的瞬态相互作用，使所有功能状态的结构表征困难。本研究探讨了传感器组氨酸激酶，HisKA和HATP酶的两个催化结构域的非活性和信号激活的构象状态。将直接偶联分析（一种全局统计推断方法）应用于来自蛋白质数据库的> 13，000个这样的结构域，以鉴定两个结构域之间的残基接触。这些接触指导结构组装的域使用MAGMA，一种先进的分子动力学对接方法。MAGMA产生的活性构象结构同时容纳了序列衍生的残基接触和ATP催化的组氨酸接触。该结构的有效性在生物学上通过枯草芽孢杆菌传感器组氨酸激酶KinA中的接触位置的突变和通过在失活的KinA（HisKA）：KinD（HATP酶）杂合蛋白中的活性的恢复来证实。这些数据表明，结合到传感器结构域的信号通过在HisKA结构域的C-末端螺旋的末端引起局部应变和解旋来激活传感器组氨酸激酶。这使两个结构域的非活性构象的接触位置不稳定，通过先前的晶体结构分析和本文所述的序列分析鉴定，诱导活性构象的形成。这项研究表明，相互作用的蛋白质和蛋白质结构域的不稳定的瞬时复合物的结构是可访问的，通过应用这种交叉验证技术的组合。
Signal transduction proteins such as bacterial sensor histidine kinases, designed to transition between multiple conformations, are often ruled by unstable transient interactions making structural characterization of all functional states difficult. This study explored the inactive and signal-activated conformational states of the two catalytic domains of sensor histidine kinases, HisKA and HATPase. Direct coupling analyses, a global statistical inference approach, was applied to > 13,000 such domains from protein databases to identify residue contacts between the two domains. These contacts guided structural assembly of the domains using MAGMA, an advanced molecular dynamics docking method. The active conformation structure generated by MAGMA simultaneously accommodated the sequence derived residue contacts and the ATP-catalytic histidine contact. The validity of this structure was confirmed biologically by mutation of contact positions in the Bacillus subtilis sensor histidine kinase KinA and by restoration of activity in an inactive KinA(HisKA): KinD(HATPase) hybrid protein. These data indicate that signals binding to sensor domains activate sensor histidine kinases by causing localized strain and unwinding at the end of the C-terminal helix of the HisKA domain. This destabilizes the contact positions of the inactive conformation of the two domains, identified by previous crystal structure analyses and by the sequence analysis described here, inducing the formation of the active conformation. This study reveals that structures of unstable transient complexes of interacting proteins and of protein domains are accessible by applying this combination of cross-validating technologies.