Analysis of the quality of crystallographic data and the limitations of structural models

Arkhipova, Valentina; Guskov, Albert; Slotboom, Dirk Jan

doi:10.1085/jgp.201711852

Cited by 14 publications

(12 citation statements)

References 63 publications

(126 reference statements)

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“…The glutamate transporter (SLC1) family of solute transporters is structurally well-characterized with 39 available structures of four different family members: the prokaryotic sodium-dependent aspartate transporters Glt Ph and Glt Tk , the human sodium-and potassium-dependent glutamate transporter EAAT1 (Excitatory Amino Acid Transporter 1), and the human neutral amino acid exchanger ASCT2 (Alanine Serine Cysteine Transporter 2) ( Table 1 and reviewed in [26]) . While Glt Ph is the prototypical elevator transporter, ASCT2 is the first SLC1 member, for which four key conformations have been resolved structurally: outward-open, outward-occluded [27], inward-open [28] and inward-occluded [29].…”

Section: Fixed Barrier Elevator With One Gatementioning

confidence: 99%

Elevator-type mechanisms of membrane transport

Garaeva

Slotboom

2020

Biochemical Society Transactions

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Membrane transporters are integral membrane proteins that mediate the passage of solutes across lipid bilayers. These proteins undergo conformational transitions between outward- and inward-facing states, which lead to alternating access of the substrate-binding site to the aqueous environment on either side of the membrane. Dozens of different transporter families have evolved, providing a wide variety of structural solutions to achieve alternating access. A sub-set of structurally diverse transporters operate by mechanisms that are collectively named ‘elevator-type’. These transporters have one common characteristic: they contain a distinct protein domain that slides across the membrane as a rigid body, and in doing so it ‘drags” the transported substrate along. Analysis of the global conformational changes that take place in membrane transporters using elevator-type mechanisms reveals that elevator-type movements can be achieved in more than one way. Molecular dynamics simulations and experimental data help to understand how lipid bilayer properties may affect elevator movements and vice versa.

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Section: Fixed Barrier Elevator With One Gatementioning

confidence: 99%

Elevator-type mechanisms of membrane transport

Garaeva

Slotboom

2020

Biochemical Society Transactions

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“…Prokaryotic glutamate transporters homologs share high structural similarity with EAATs and neutral amino acid transporters ASCTs, which all belong to the solute carrier family 1 (SLC1A) 7 . Structural studies of the archaeal homologs Glt Ph from Pyrococcus horikoshii and Glt Tk from Thermococcus kodakarensis have provided the structural basis for understanding of the transport mechanism [8][9][10][11][12][13] . Both Glt Ph and Glt Tk couple uptake of one aspartate molecule to symport of three sodium ions 12,14 .…”

mentioning

confidence: 99%

Structural ensemble of a glutamate transporter homologue in lipid nanodisc environment

2020

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Glutamate transporters are cation-coupled secondary active membrane transporters that clear the neurotransmitter L-glutamate from the synaptic cleft. These transporters are homotrimers, with each protomer functioning independently by an elevator-type mechanism, in which a mobile transport domain alternates between inward-and outward-oriented states. Using single-particle cryo-EM we have determined five structures of the glutamate transporter homologue Glt Tk , a Na + -L-aspartate symporter, embedded in lipid nanodiscs. Dependent on the substrate concentrations used, the protomers of the trimer adopt a variety of asymmetrical conformations, consistent with the independent movement. Six of the 15 resolved protomers are in a hitherto elusive state of the transport cycle in which the inwardfacing transporters are loaded with Na + ions. These structures explain how substrate-leakage is preventeda strict requirement for coupled transport. The belt protein of the lipid nanodiscs bends around the inward oriented protomers, suggesting that membrane deformations occur during transport.

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“…In the TBOA:Glt Ph crystal structure, HP2 is an OFC open conformation, moving from its position in the aspartate-bound complex towards the loop between TM3 and TM4 ( Figure 2C). This HP2 movement in the TBOA-bound complex exposes the substrate-binding site to the extracellular solution, suggesting that, in the apo state, HP2 adopts an open conformation before aspartate binding [22]. The structure of substrate-free transporter for the Glt Tk homologue revealed an OFC with occluded binding site and closed HP2 [15].…”

Section: Modulationmentioning

confidence: 98%

“…Potassium binding to the IFC has been proposed to enable translocation of the substrate-free transport domain to the OFC [19]. Verdon and colleagues claimed to identify a potassium-binding site in Glt Ph [19], but subsequent structure analysis led to this being disregarded [22].…”

Section: Potassium Counter-transportmentioning

confidence: 99%

“…The best structurally characterized members of SLC1 transporters are archaeal homologues Glt Ph and Glt Tk [12][13][14][15][16][17][18][19][20][21] (Table 1). They share ∼36% sequence identity with eukaryotic EAATs [22] with many conserved residues involved in substrate and ion binding and transport [13]. The progress in crystallographic studies of the glutamate transporter family, including the structure of human EAAT1 [23], has revealed much about the mechanism of glutamate transporters.…”

Section: Introductionmentioning

confidence: 99%

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Glutamate transporters: a broad review of the most recent archaeal and human structures

Pavić

Holmes

Postis

et al. 2019

Biochemical Society Transactions

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Glutamate transporters play important roles in bacteria, archaea and eukaryotes. Their function in the mammalian central nervous system is essential for preventing excitotoxicity, and their dysregulation is implicated in many diseases, such as epilepsy and Alzheimer's. Elucidating their transport mechanism would further the understanding of these transporters and promote drug design as they provide compelling targets for understanding the pathophysiology of diseases and may have a direct role in the treatment of conditions involving glutamate excitotoxicity. This review outlines the insights into the transport cycle, uncoupled chloride conductance and modulation, as well as identifying areas that require further investigation.*These authors contributed equally to this work.

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Analysis of the quality of crystallographic data and the limitations of structural models

Abstract: Arkhipova et al. caution that the limitations of structural models be taken into account when interpreting crystallographic data.

Cited by 14 publications

References 63 publications

Elevator-type mechanisms of membrane transport

Elevator-type mechanisms of membrane transport

Structural ensemble of a glutamate transporter homologue in lipid nanodisc environment

Glutamate transporters: a broad review of the most recent archaeal and human structures

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