Department of Biochemistry and Biophysics, Science for Life Laboratory, Stockholm University, 10691 Stockholm, Sweden.
Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, 69120 Heidelberg, Germany.
Proc Natl Acad Sci U S A. 2022 Dec 13;119(50):e2210669119. doi: 10.1073/pnas.2210669119. Epub 2022 Dec 8.
Pentameric ligand-gated ion channels (pLGICs) perform electrochemical signal transduction in organisms ranging from bacteria to humans. Among the prokaryotic pLGICs, there is architectural diversity involving N-terminal domains (NTDs) not found in eukaryotic relatives, exemplified by the calcium-sensitive channel (DeCLIC) from a deltaproteobacterium, which has an NTD in addition to the canonical pLGIC structure. Here, we have characterized the structure and dynamics of DeCLIC through cryoelectron microscopy (cryo-EM), small-angle neutron scattering (SANS), and molecular dynamics (MD) simulations. In the presence and absence of calcium, cryo-EM yielded structures with alternative conformations of the calcium-binding site. SANS profiles further revealed conformational diversity at room temperature beyond that observed in static structures, shown through MD to be largely attributable to rigid-body motions of the NTD relative to the protein core, with expanded and asymmetric conformations improving the fit of the SANS data. This work reveals the range of motion available to the DeCLIC NTD and calcium-binding site, expanding the conformational landscape of the pLGIC family. Further, these findings demonstrate the power of combining low-resolution scattering, high-resolution structural, and MD simulation data to elucidate interfacial interactions that are highly conserved in the pLGIC family.
五聚体配体门控离子通道（pLGICs）在从细菌到人类的生物体中进行电化学信号转导。在原核 pLGIC 中，存在涉及到真核同源物中不存在的 N 端结构域（NTD）的结构多样性，以钙敏感性通道（DeCLIC）为例，它来自δ变形菌，除了经典的 pLGIC 结构外，还具有 NTD。在这里，我们通过冷冻电镜（cryo-EM）、小角中子散射（SANS）和分子动力学（MD）模拟对 DeCLIC 的结构和动力学进行了表征。在存在和不存在钙的情况下，cryo-EM 产生了具有替代钙结合位点构象的结构。SANS 谱进一步揭示了在室温下除静态结构观察到的构象多样性，通过 MD 表明这主要归因于 NTD 相对于蛋白质核心的刚体运动，扩展和不对称构象改善了 SANS 数据的拟合。这项工作揭示了 DeCLIC NTD 和钙结合位点的运动范围，扩展了 pLGIC 家族的构象景观。此外，这些发现证明了结合低分辨率散射、高分辨率结构和 MD 模拟数据的力量，以阐明在 pLGIC 家族中高度保守的界面相互作用。
