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  Early Intermediates in the Transport Cycle of the Neuronal Excitatory Amino Acid Carrier EAAC1

Watzke, N., Bamberg, E., & Grewer, C. (2001). Early Intermediates in the Transport Cycle of the Neuronal Excitatory Amino Acid Carrier EAAC1. Journal of General Physiology, 117(6), 547-562. doi:10.1085/jgp.117.6.547.

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 Creators:
Watzke, Natalie1, Author              
Bamberg, Ernst1, Author              
Grewer, Christof1, Author              
Affiliations:
1Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society, ou_2068289              

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Free keywords: glutamate transporter; charge movement; patch clamp; caged compounds; rapid kinetics
 Abstract: Electrogenic glutamate transport by the excitatory amino acid carrier 1 (EAAC1) is associated with multiple charge movements across the membrane that take place on time scales ranging from microseconds to milliseconds. The molecular nature of these charge movements is poorly understood at present and, therefore, was studied in this report in detail by using the technique of laser-pulse photolysis of caged glutamate providing a 100-μs time resolution. In the inward transport mode, the deactivation of the transient component of the glutamate-induced coupled transport current exhibits two exponential components. Similar results were obtained when restricting EAAC1 to Na+ translocation steps by removing potassium, thus, demonstrating (1) that substrate translocation of EAAC1 is coupled to inward movement of positive charge and, therefore, electrogenic; and (2) the existence of at least two distinct intermediates in the Na+-binding and glutamate translocation limb of the EAAC1 transport cycle. Together with the determination of the sodium ion concentration and voltage dependence of the two-exponential charge movement and of the steady-state EAAC1 properties, we developed a kinetic model that is based on sequential binding of Na+ and glutamate to their extracellular binding sites on EAAC1 explaining our results. In this model, at least one Na+ ion and thereafter glutamate rapidly bind to the transporter initiating a slower, electroneutral structural change that makes EAAC1 competent for further, voltage-dependent binding of additional sodium ion(s). Once the fully loaded EAAC1 complex is formed, it can undergo a much slower, electrogenic translocation reaction to expose the substrate and ion binding sites to the cytoplasm.

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Language(s): eng - English
 Dates: 2001-04-132001-02-122001-04-162001-05-292001-06-01
 Publication Status: Published in print
 Pages: 16
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.1085/jgp.117.6.547
PMID: 11382805
 Degree: -

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Title: Journal of General Physiology
  Other : J. Gen. Physiol.
  Abbreviation : JGP
Source Genre: Journal
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Publ. Info: Rockefeller University Press
Pages: - Volume / Issue: 117 (6) Sequence Number: - Start / End Page: 547 - 562 Identifier: ISSN: 0022-1295
CoNE: https://pure.mpg.de/cone/journals/resource/954925413895