A conserved leucine occupies the empty substrate site of LeuT in the Na+-free return state

DOI: 10.1038/ncomms11673

Authors: Lina Malinauskaite (Aarhus University) , Saida Said (Aarhus University) , Caglanur Sahin (Aarhus University) , Julie Grouleff (Aarhus University) , Azadeh Shahsavar (Aarhus University) , Henriette Bjerregaard (Aarhus University) , Pernille Noer (Aarhus University) , Kasper Severinsen (Aarhus University) , Thomas Boesen (Aarhus University) , Birgit Schiøtt (Aarhus University) , Steffen Sinning (Aarhus University) , Poul Nissen (Aarhus University)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Nature Communications , VOL 7

State: Published (Approved)
Published: May 2016

Abstract: Bacterial members of the neurotransmitter:sodium symporter (NSS) family perform Na+-dependent amino-acid uptake and extrude H+ in return. Previous NSS structures represent intermediates of Na+/substrate binding or intracellular release, but not the inward-to-outward return transition. Here we report crystal structures of Aquifex aeolicus LeuT in an outward-oriented, Na+- and substrate-free state likely to be H+-occluded. We find a remarkable rotation of the conserved Leu25 into the empty substrate-binding pocket and rearrangements of the empty Na+ sites. Mutational studies of the equivalent Leu99 in the human serotonin transporter show a critical role of this residue on the transport rate. Molecular dynamics simulations show that extracellular Na+ is blocked unless Leu25 is rotated out of the substrate-binding pocket. We propose that Leu25 facilitates the inward-to-outward transition by compensating a Na+- and substrate-free state and acts as the gatekeeper for Na+ binding that prevents leak in inward-outward return transitions.

Journal Keywords: Membrane proteins; Structural biology

Subject Areas: Biology and Bio-materials, Chemistry


Instruments: I24-Microfocus Macromolecular Crystallography

Added On: 17/11/2016 14:32

Documents:
ncomms11673.pdf

Discipline Tags:

Health & Wellbeing Neurology Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)