4X2S

Crystal structure of 276S/M395R-GltPh in inward-facing conformation

Summary for 4X2S

• Entry DOI: 10.2210/pdb4x2s/pdb
• Descriptor: 425aa long hypothetical proton glutamate symport protein, ASPARTIC ACID, SODIUM ION (3 entities in total)
• Functional Keywords: amino acid secondary transporters, sodium coupled aspartate transporter, transport protein
• Biological source: Pyrococcus horikoshii OT3
• Total number of polymer chains: 3
• Total formula weight: 134234.55

Authors

Akyuz, N.,Boudker, O. (deposition date: 2014-11-26, release date: 2015-02-04, Last modification date: 2023-09-27)

Primary citation

Akyuz, N.,Georgieva, E.R.,Zhou, Z.,Stolzenberg, S.,Cuendet, M.A.,Khelashvili, G.,Altman, R.B.,Terry, D.S.,Freed, J.H.,Weinstein, H.,Boudker, O.,Blanchard, S.C.
Transport domain unlocking sets the uptake rate of an aspartate transporter.
Nature, 518:68-73, 2015

PubMed Abstract: Glutamate transporters terminate neurotransmission by clearing synaptically released glutamate from the extracellular space, allowing repeated rounds of signalling and preventing glutamate-mediated excitotoxicity. Crystallographic studies of a glutamate transporter homologue from the archaeon Pyrococcus horikoshii, GltPh, showed that distinct transport domains translocate substrates into the cytoplasm by moving across the membrane within a central trimerization scaffold. Here we report direct observations of these 'elevator-like' transport domain motions in the context of reconstituted proteoliposomes and physiological ion gradients using single-molecule fluorescence resonance energy transfer (smFRET) imaging. We show that GltPh bearing two mutations introduced to impart characteristics of the human transporter exhibits markedly increased transport domain dynamics, which parallels an increased rate of substrate transport, thereby establishing a direct temporal relationship between transport domain motion and substrate uptake. Crystallographic and computational investigations corroborated these findings by revealing that the 'humanizing' mutations favour structurally 'unlocked' intermediate states in the transport cycle exhibiting increased solvent occupancy at the interface between the transport domain and the trimeric scaffold.

PubMed: 25652997
DOI: 10.1038/nature14158

Experimental method

X-RAY DIFFRACTION (4.21 Å)