Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston, Houston, TX.
E. Carrillo and S.A. Shaikh contributed equally to this work and are listed in alphabetical order.
J Gen Physiol. 2020 Jan 6;152(1). doi: 10.1085/jgp.201912451.
Fast excitatory synaptic transmission in the mammalian central nervous system is mediated by glutamate-activated α-amino-5-methyl-3-hydroxy-4-isoxazole propionate (AMPA) receptors. In neurons, AMPA receptors coassemble with transmembrane AMPA receptor regulatory proteins (TARPs). Assembly with TARP γ8 alters the biophysical properties of the receptor, producing resensitization currents in the continued presence of glutamate. Using single-channel recordings, we show that under resensitizing conditions, GluA2 AMPA receptors primarily transition to higher conductance levels, similar to activation of the receptors in the presence of cyclothiazide, which stabilizes the open state. To study the conformation associated with these states, we have used single-molecule FRET and show that this high-conductance state exhibits tighter coupling between subunits in the extracellular parts of the receptor. Furthermore, the dwell times for the transition from the tightly coupled state to the decoupled states correlate to longer open durations of the channels, thus correlating conformation and function at the single-molecule level.
在哺乳动物中枢神经系统中，快速兴奋性突触传递是由谷氨酸激活的α-氨基-5-甲基-3-羟基-4-异恶唑丙酸（AMPA）受体介导的。在神经元中，AMPA 受体与跨膜 AMPA 受体调节蛋白（TARPs）共组装。与 TARPγ8 的组装改变了受体的生物物理特性，在持续存在谷氨酸的情况下产生再敏化电流。使用单通道记录，我们表明在再敏化条件下，GluA2 AMPA 受体主要转变为更高的传导水平，类似于在环噻嗪存在下激活受体，其稳定开放状态。为了研究与这些状态相关的构象，我们使用了单分子 FRET，并表明这种高传导状态在受体的细胞外部分的亚基之间表现出更紧密的偶联。此外，从紧密偶联状态到去偶联状态的转变的停留时间与通道的更长的开放持续时间相关，因此在单分子水平上关联构象和功能。
