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Proc Natl Acad Sci U S A. 2012 Jan 31;109(5):E210-7. doi: 10.1073/pnas.1115581109. Epub 2012 Jan 9.

Mechanism of proton/substrate coupling in the heptahelical lysosomal transporter cystinosin.

Author information

1
Université Paris Descartes, Sorbonne Paris Cité, Centre National de la Recherche Scientifique, Unité Mixte de Recherche 8192, Centre Universitaire des Saints-Pères, 45 rue des Saints-Pères, F-75006 Paris, France.

Abstract

Secondary active transporters use electrochemical gradients provided by primary ion pumps to translocate metabolites or drugs "uphill" across membranes. Here we report the ion-coupling mechanism of cystinosin, an unusual eukaryotic, proton-driven transporter distantly related to the proton pump bacteriorhodopsin. In humans, cystinosin exports the proteolysis-derived dimeric amino acid cystine from lysosomes and is impaired in cystinosis. Using voltage-dependence analysis of steady-state and transient currents elicited by cystine and neutralization-scanning mutagenesis of conserved protonatable residues, we show that cystine binding is coupled to protonation of a clinically relevant aspartate buried in the membrane. Deuterium isotope substitution experiments are consistent with an access of this aspartate from the lysosomal lumen through a deep proton channel. This aspartate lies in one of the two PQ-loop motifs shared by cystinosin with a set of eukaryotic membrane proteins of unknown function and is conserved in about half of them, thus suggesting that other PQ-loop proteins may translocate protons.

PMID:
22232659
PMCID:
PMC3277178
DOI:
10.1073/pnas.1115581109
[Indexed for MEDLINE]
Free PMC Article

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