小柯机器人

美国威斯康星大学麦迪逊分校Baron Chanda、Randall H. Goldsmith等研究人员合作发现，cAMP与封闭的起搏器离子通道的结合是非协同的。相关论文于2021年6月30日在线发表在《自然》杂志上。

研究人员表示，大脑和心脏中的电活动取决于起搏器离子通道（HCN）有节奏地产生动作电位，其活动受cAMP结合的调节。以前的工作已经发现了cAMP结合中正协同和负协同的证据，但这种批量测量的参数分辨率有限。由于无法直接监测单个配体分子在生理浓度下与膜受体的结合，使用单分子技术消除这种模糊性的努力受到了阻碍。

研究人员使用纳米光子零模波导来克服了这些挑战，从而直接解析了单个配体与多聚HCN1和HCN2离子通道的结合动力学。结果表明，尽管随后在每个位点构象异构化为翻转状态，但当孔关闭时，cAMP独立地与所有四个亚基结合。结合和异构化的不同动力学可能是每个亚型对cAMP的生理学不同反应的基础，并提供了配体诱导翻转状态模型的直接验证。这种观察多聚体蛋白在生理相关浓度下逐步结合的方法，可以直接探测其他完整膜蛋白和受体中单分子分辨率的结合变构。

附：英文原文

Title: cAMP binding to closed pacemaker ion channels is non-cooperative

Author: David S. White, Sandipan Chowdhury, Vinay Idikuda, Ruohan Zhang, Scott T. Retterer, Randall H. Goldsmith, Baron Chanda

Issue&Volume: 2021-06-30

Abstract: Electrical activity in the brain and heart depends on rhythmic generation of action potentials by pacemaker ion channels (HCN) whose activity is regulated by cAMP binding1. Previous work has uncovered evidence for both positive and negative cooperativity in cAMP binding2,3, but such bulk measurements suffer from limited parameter resolution. Efforts to eliminate this ambiguity using single-molecule techniques have been hampered by the inability to directly monitor binding of individual ligand molecules to membrane receptors at physiological concentrations. Here we overcome these challenges using nanophotonic zero-mode waveguides4 to directly resolve binding dynamics of individual ligands to multimeric HCN1 and HCN2 ion channels. We show that cAMP binds independently to all four subunits when the pore is closed, despite a subsequent conformational isomerization to a flip state at each site. The different dynamics in binding and isomerization are likely to underlie physiologically distinct responses of each isoform to cAMP5 and provide direct validation of the ligand-induced flip-state model6,7,8,9. This approach for observing stepwise binding in multimeric proteins at physiologically relevant concentrations can directly probe binding allostery at single-molecule resolution in other intact membrane proteins and receptors.

DOI: 10.1038/s41586-021-03686-x

Source: https://www.nature.com/articles/s41586-021-03686-x

Nature：《自然》，创刊于1869年。隶属于施普林格·自然出版集团，最新IF：69.504
官方网址：http://www.nature.com/
投稿链接：http://www.nature.com/authors/submit_manuscript.html