Biotinylation of single cysteine mutants of the glutamate transporter GLT-1 from rat brain reveals its unusual topology

Myriam Grunewald, Annie Bendahan, Baruch I. Kanner*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

138 Scopus citations

Abstract

In the central nervous system, (Na+ + K+)-coupled glutamate transporters restrict the neurotoxicity of this transmitter and limit the duration of synaptic excitation at some synapses. The various isotransporters exhibit a particularly high homology in an extended hydrophobic domain of ill-defined topology that contains several determinants involved in ion and transmitter binding. Here, we describe the determination of the membrane topology of the cloned astroglial glutamate transporter GLT-1. A series of functional transporters containing single cysteines was engineered. Their topological disposition was determined by using a biotinylated sulfhydryl reagent. The glutamate transporter has eight transmembrane domains long enough to span the membrane as α helices. Strikingly, between the seventh and eighth domains, a structure reminiscent of a pore loop and an outward- facing hydrophobic linker are positioned.

Original languageAmerican English
Pages (from-to)623-632
Number of pages10
JournalNeuron
Volume21
Issue number3
DOIs
StatePublished - Sep 1998

Bibliographical note

Funding Information:
This work was supported by grants from the US–Israel Binational Science Foundation and from the Israel Science Foundation, which is administered by the Israel Academy of Sciences and Humanities, the Charles Revson Foundation, and the Bernard Katz Minerva Center for Cell Biophysics. Expert secretarial help by Mrs. Beryl Levene is gratefully acknowledged.

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