Millisecond dynamics of an unlabeled amino acid transporter.
Millisecond dynamics of an unlabeled amino acid transporter.
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DOI：
10.1038/s41467-020-18811-z
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发表时间：
2020-10-06
影响因子：
16.6
通讯作者：
Scheuring S
中科院分区：
文献类型：
--
作者：
Matin TR;Heath GR;Huysmans GHM;Boudker O;Scheuring S
兴奋性氨基酸转运蛋白（EAAT）在许多生理过程中起着重要作用，并对兴奋性氨基酸从突触间隙中的清除至关重要。在这里，我们开发和应用高速原子力显微镜线扫描（HS-AFM-LS）结合自动化状态分配和转换分析，用于确定未标记的膜重组GltPh，原核EAAT同系物，毫秒级时间分辨率的运输动力学。我们发现，GltPh转运蛋白可以比以前报道的更快地运行，状态停留时间在50毫秒的范围内，并报告中间运输状态的动力学与高度之间的向外和向内的状态。转运域随机探针跨膜运动，和可逆的不成功的漂移到中间状态发生。所提出的方法和分析方法通常适用于研究转运蛋白动力学在系统相关的时间分辨率。兴奋性氨基酸转运蛋白（EAAT）是从突触间隙中清除兴奋性氨基酸的关键。在这里，作者结合了高速原子力显微镜线扫描与自动状态分配的确定运输动力学的GltPh，原核EAAT同系物，毫秒级的时间分辨率。
Excitatory amino acid transporters (EAATs) are important in many physiological processes and crucial for the removal of excitatory amino acids from the synaptic cleft. Here, we develop and apply high-speed atomic force microscopy line-scanning (HS-AFM-LS) combined with automated state assignment and transition analysis for the determination of transport dynamics of unlabeled membrane-reconstituted GltPh, a prokaryotic EAAT homologue, with millisecond temporal resolution. We find that GltPh transporters can operate much faster than previously reported, with state dwell-times in the 50 ms range, and report the kinetics of an intermediate transport state with height between the outward- and inward-facing states. Transport domains stochastically probe transmembrane motion, and reversible unsuccessful excursions to the intermediate state occur. The presented approach and analysis methodology are generally applicable to study transporter kinetics at system-relevant temporal resolution. Excitatory amino acid transporters (EAATs) are crucial for the removal of excitatory amino acids from the synaptic cleft. Here authors combined high-speed atomic force microscopy line-scanning with automated state assignment for the determination of transport dynamics of GltPh, a prokaryotic EAAT homologue, with millisecond temporal resolution.
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影响因子：
15
作者：
Grazioso, Giovanni;Limongelli, Vittorio;Parrinello, Michele
通讯作者：
Parrinello, Michele
DOI：
10.1073/pnas.211400898
发表时间：
2001-10-23
影响因子：
11.1
作者：
Ando, T;Kodera, N;Toda, A
通讯作者：
Toda, A
影响因子：
6.2
作者：
Benediktsson, Adrienne M.;Marrs, Glen S.;Tu, Jian Cheng;Worley, Paul F.;Roth-Stein, Jeffrey D.;Bergles, Dwight E.;Dailey, Michael E.
通讯作者：
Dailey, Michael E.
影响因子：
16.8
作者：
Hanelt, Inga;Wunnicke, Dorith;Slotboom, Dirk Jan
通讯作者：
Slotboom, Dirk Jan
影响因子：
64.8
作者：
Canul-Tec JC;Assal R;Cirri E;Legrand P;Brier S;Chamot-Rooke J;Reyes N
通讯作者：
Reyes N