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2004-03-19

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Please use this identifier to cite or link to this item: https://hdl.handle.net/2445/181312

Thiol modification of cysteine 327 in the eighth transmembrane domain of the light subunit xCT of the heteromeric cystine/glutamate antiporter suggests close proximity to the substrate binding site/permeation pathway

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https://doi.org/10.1074/jbc.M309866200

Abstract

We measured sensitivity to thiol modification of the heteromeric glutamate/cystine transporter 4F2hc-xCT expressed in Xenopus oocytes. p-Chloromercuribenzoate (pCMB) and p-chloromercuribenzenesulfonate (pCMBS) rapidly blocked transport activity. Cys(327), located in the middle of the eighth transmembrane domain of the light subunit (xCT), was found to be the main target of inactivation. Cysteine, an impermeant reducing reagent, reversed pCMB and pCMBS effects only when applied from the extracellular medium. l-Glutamate and l-cystine, but not l-arginine, protected from the inactivation with an IC(50) similar to the K(m). Protection was not temperature-dependent, suggesting that it did not depend on large substrate-induced conformational changes. Mutation of Cys(327) to Ala and Ser slightly modified the K(m) and a C327L mutant abolished transport function without compromising transporter expression at the plasma membrane. The results indicate that Cys(327) is a functionally important residue accessible to the aqueous extracellular environment and is structurally linked to the permeation pathway and/or the substrate binding site.

Subject

Aminoàcids, Proteïnes portadores, Proteïnes de membrana

Subject (English)

Amino acids, Carrier proteins, Membrane proteins

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Articles publicats en revistes (Ciències Fisiològiques)
Articles publicats en revistes (Biologia Cel·lular, Fisiologia i Immunologia)
Articles publicats en revistes (Bioquímica i Biomedicina Molecular)

Citation

JIMÉNEZ VIDAL, Maite, et al. Thiol modification of cysteine 327 in the eighth transmembrane domain of the light subunit xCT of the heteromeric cystine/glutamate antiporter suggests close proximity to the substrate binding site/permeation pathway. Journal of Biological Chemistry. 2004. Vol. 279, num. 12, pags. 11214-11221. ISSN 0021-9258. [consulted: 13 of August of 2026]. Available at: https://hdl.handle.net/2445/181312

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