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Proc Natl Acad Sci U S A
2009 Dec 08;10649:20752-7. doi: 10.1073/pnas.0908570106.
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Inward-facing conformation of glutamate transporters as revealed by their inverted-topology structural repeats.
Crisman TJ
,
Qu S
,
Kanner BI
,
Forrest LR
.
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Glutamate transporters regulate synaptic concentrations of this neurotransmitter by coupling its flux to that of sodium and other cations. Available crystal structures of an archeal homologue of these transporters, GltPh, resemble an extracellular-facing state, in which the bound substrate is occluded only by a small helical hairpin segment called HP2. However, a pathway to the cytoplasmic side of the membrane is not clearly apparent. We previously modeled an alternate state of a transporter from the neurotransmitter:sodium symporter family, which has an entirely different fold, solely on the presence of inverted-topology structural repeats. In GltPh, we identified two distinct sets of inverted-topology repeats and used these repeats to model an inward-facing conformation of the protein. To test this model, we introduced pairs of cysteines into the neuronal glutamate transporter EAAC1, at positions that are >27 A apart in the crystal structures of GltPh, but approximately = 10 A apart in the inward-facing model. Transport by these mutants was activated by pretreatment with the reducing agent dithithreitol. Subsequent treatment with the oxidizing agent copper(II)(1,10-phenantroline)(3) abolished this activation. The inhibition of transport was potentiated under conditions thought to promote the inward-facing conformation of the transporter. By contrast, the inhibition was reduced in the presence of the nontransportable substrate analogue D,L-threo-beta-benzyloxyaspartate, which favors the outward-facing conformation. Other conformation-sensitive accessibility measurements are also accommodated by our inward-facing model. These results suggest that the inclusion of inverted-topology repeats in transporters may provide a general solution to the requirement for two symmetry-related states in a single protein.
Bendahan,
Arginine 447 plays a pivotal role in substrate interactions in a neuronal glutamate transporter.
2000, Pubmed,
Xenbase
Bendahan,
Arginine 447 plays a pivotal role in substrate interactions in a neuronal glutamate transporter.
2000,
Pubmed
,
Xenbase
Borre,
Arginine 445 controls the coupling between glutamate and cations in the neuronal transporter EAAC-1.
2004,
Pubmed
Boudker,
Coupling substrate and ion binding to extracellular gate of a sodium-dependent aspartate transporter.
2007,
Pubmed
Brocke,
Proximity of two oppositely oriented reentrant loops in the glutamate transporter GLT-1 identified by paired cysteine mutagenesis.
2002,
Pubmed
Faham,
The crystal structure of a sodium galactose transporter reveals mechanistic insights into Na+/sugar symport.
2008,
Pubmed
Forrest,
Mechanism for alternating access in neurotransmitter transporters.
2008,
Pubmed
Groeneveld,
Rigidity of the subunit interfaces of the trimeric glutamate transporter GltT during translocation.
2007,
Pubmed
Grunewald,
Conformational changes monitored on the glutamate transporter GLT-1 indicate the existence of two neurotransmitter-bound states.
1995,
Pubmed
Gu,
Molecular simulations elucidate the substrate translocation pathway in a glutamate transporter.
2009,
Pubmed
Huang,
Dynamics of the extracellular gate and ion-substrate coupling in the glutamate transporter.
2008,
Pubmed
Jardetzky,
Simple allosteric model for membrane pumps.
1966,
Pubmed
Kanai,
Primary structure and functional characterization of a high-affinity glutamate transporter.
1992,
Pubmed
,
Xenbase
Kanner,
Binding order of substrates to the sodium and potassium ion coupled L-glutamic acid transporter from rat brain.
1982,
Pubmed
Kanner,
Gate movements in glutamate transporters.
2007,
Pubmed
Kavanaugh,
Mutation of an amino acid residue influencing potassium coupling in the glutamate transporter GLT-1 induces obligate exchange.
1997,
Pubmed
Koch,
Small-scale molecular motions accomplish glutamate uptake in human glutamate transporters.
2005,
Pubmed
,
Xenbase
Krishnamurthy,
Unlocking the molecular secrets of sodium-coupled transporters.
2009,
Pubmed
Kunkel,
Rapid and efficient site-specific mutagenesis without phenotypic selection.
1987,
Pubmed
Leighton,
Structural rearrangements at the translocation pore of the human glutamate transporter, EAAT1.
2006,
Pubmed
Levy,
Stoichiometry of the glial glutamate transporter GLT-1 expressed inducibly in a Chinese hamster ovary cell line selected for low endogenous Na+-dependent glutamate uptake.
1998,
Pubmed
Mim,
Two conformational changes are associated with glutamate translocation by the glutamate transporter EAAC1.
2007,
Pubmed
Pines,
Glutamate 404 is involved in the substrate discrimination of GLT-1, a (Na+ + K+)-coupled glutamate transporter from rat brain.
1995,
Pubmed
Pines,
Counterflow of L-glutamate in plasma membrane vesicles and reconstituted preparations from rat brain.
1990,
Pubmed
Qu,
Substrates and non-transportable analogues induce structural rearrangements at the extracellular entrance of the glial glutamate transporter GLT-1/EAAT2.
2008,
Pubmed
Reyes,
Transport mechanism of a bacterial homologue of glutamate transporters.
2009,
Pubmed
Ryan,
The chloride permeation pathway of a glutamate transporter and its proximity to the glutamate translocation pathway.
2004,
Pubmed
,
Xenbase
Sali,
Comparative protein modelling by satisfaction of spatial restraints.
1993,
Pubmed
Shlaifer,
Conformationally sensitive reactivity to permeant sulfhydryl reagents of cysteine residues engineered into helical hairpin 1 of the glutamate transporter GLT-1.
2007,
Pubmed
Shrivastava,
Time-resolved mechanism of extracellular gate opening and substrate binding in a glutamate transporter.
2008,
Pubmed
Thompson,
CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice.
1994,
Pubmed
Yernool,
Structure of a glutamate transporter homologue from Pyrococcus horikoshii.
2004,
Pubmed
Zerangue,
Flux coupling in a neuronal glutamate transporter.
1996,
Pubmed
,
Xenbase
Zomot,
Proximity of transmembrane domains 1 and 3 of the gamma-aminobutyric acid transporter GAT-1 inferred from paired cysteine mutagenesis.
2005,
Pubmed
