Abstract
The transport of amino acids across cell membranes is essential for metabolism, neuronal signalling, and immune system function. The amino acid polyamine organocation (APC) superfamily controls amino acid transport via mechanisms including amino acid exchange, facilitative diffusion, and sodium- or proton-coupled transport. Although many mammalian APC members functioning as exchangers and sodium-coupled systems have been identified, the mechanisms underlying pH-regulated amino acid transport in mammalian cells remain unclear. Here, we show that the plasma membrane amino acid transporter SLC7A4 is regulated by low extracellular pH and functions as a leucine transporter in human cells. Using Cryo-EM structures of the plant homologue, CAT4, from Arabidopsis thaliana in outward-open apo and L-ornithine-bound states, as well as transport assays and molecular dynamics simulations based on homology models of the human transporter, we identify residues responsible for amino acid selectivity that supports an allosteric mechanism linking ligand recognition to pH regulation. This mechanism is consistent with an evolutionary link to proton-coupled prokaryotic homologues. Overall, our findings provide a structural and functional basis for pH-gated leucine transport by the human SLC7A4 transporter and provides a framework for understanding amino acid selectivity within the wider SLC7 family.
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Data availability
Atomic coordinates for SLC7A4 have been deposited in the Protein Data Bank under accession codes: 9HJK (Apo with Syb), 9SP8 (L-Orn with Syb) and 9SQH (Apo without Syb). The cryo-EM maps have been deposited in the Electron Microscopy Data Bank (EMDB) under accession codes: EMD-52217, EMD-55065 and EMD-55110. Source data are provided with this paper.
Code availability
The processed molecular dynamics simulations trajectories have been uploaded to Zenodo: (https://doi.org/10.5281/zenodo.17184930).
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Acknowledgements
This research was supported by Wellcome awards (215519/Z/19/Z & 219531/Z/19/Z) to SN and UKRI BBSRC award BB/Z517215/1 to JLP and SN. Computing was supported via the Advanced Research Computing facility, Oxford, the EPSRC ARCHER2 UK National Supercomputing Service and JADE (EP/X035603/1), granted via the High-End Computing Consortium for Biomolecular Simulation (HECBioSim-https://www.hecbiosim.ac.uk), supported by EPSRC (EP/X035603/1) to PCB. DK was supported by a BBSRC studentship (BB/ M011224/1) and an Onassis Foundation PhD scholarship award (F ZO 035-1/2018-2019). AB was supported by a BBSRC studentship (BB/T008784/1). The authors gratefully acknowledge the Micron Bioimaging Facility for their support & assistance in this work.
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D.K., J.L.P. and S.N. conceived the project. D.K. and A.B. performed all cloning, protein preparation and transport assays. D.K., T.K. and Y.C.Z performed all cryo-EM sample processing, data collection and image analysis. D.K. and S.N. constructed the atomic models with assistance from Y.C.Z. D.K., S.L. and P.C.B. performed all molecular dynamics simulations and analysis. D.K. and S.N. wrote the manuscript and prepared figures with contributions and discussions from J.L.P.
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Kolokouris, D., Bothra, A., Kato, T. et al. Structural basis for pH-responsive amino acid transport via SLC7A4.. Nat Commun (2026). https://doi.org/10.1038/s41467-026-70956-5
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DOI: https://doi.org/10.1038/s41467-026-70956-5
