Millisecond dynamics of an unlabeled amino acid transporter
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...
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Published in | Nature communications Vol. 11; no. 1; p. 5016 |
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Main Authors | , , , , |
Format | Journal Article |
Language | English |
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06.10.2020
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Abstract | 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 Glt
Ph
, a prokaryotic EAAT homologue, with millisecond temporal resolution. We find that Glt
Ph
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 Glt
Ph
, a prokaryotic EAAT homologue, with millisecond temporal resolution. |
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AbstractList | 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 Glt
Ph
, a prokaryotic EAAT homologue, with millisecond temporal resolution. We find that Glt
Ph
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. 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 Glt Ph , a prokaryotic EAAT homologue, with millisecond temporal resolution. We find that Glt Ph 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 Glt Ph , a prokaryotic EAAT homologue, with millisecond temporal resolution. |
ArticleNumber | 5016 |
Author | Matin, Tina R. Scheuring, Simon Huysmans, Gerard H. M. Heath, George R. Boudker, Olga |
Author_xml | – sequence: 1 givenname: Tina R. surname: Matin fullname: Matin, Tina R. organization: Department of Anesthesiology, Weill Cornell Medicine, Department of Physiology and Biophysics, Weill Cornell Medicine – sequence: 2 givenname: George R. orcidid: 0000-0001-6431-2191 surname: Heath fullname: Heath, George R. organization: Department of Anesthesiology, Weill Cornell Medicine, Department of Physiology and Biophysics, Weill Cornell Medicine – sequence: 3 givenname: Gerard H. M. orcidid: 0000-0003-4156-8252 surname: Huysmans fullname: Huysmans, Gerard H. M. organization: Department of Physiology and Biophysics, Weill Cornell Medicine – sequence: 4 givenname: Olga surname: Boudker fullname: Boudker, Olga organization: Department of Physiology and Biophysics, Weill Cornell Medicine, Howard Hughes Medical Institute, Weill Cornell Medicine – sequence: 5 givenname: Simon orcidid: 0000-0003-3534-069X surname: Scheuring fullname: Scheuring, Simon email: sis2019@med.cornell.edu organization: Department of Anesthesiology, Weill Cornell Medicine, Department of Physiology and Biophysics, Weill Cornell Medicine |
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Snippet | Excitatory amino acid transporters (EAATs) are important in many physiological processes and crucial for the removal of excitatory amino acids from the... Excitatory amino acid transporters (EAATs) are crucial for the removal of excitatory amino acids from the synaptic cleft. Here authors combined high-speed... |
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Title | Millisecond dynamics of an unlabeled amino acid transporter |
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