Substrate-bound outward-open structure of a Na+-coupled sialic acid symporter reveals a new Na+ site

Weixiao Yuan Wahlgren ,Rachel A North,Mariafrancesca Scalise,Johan Bengtsson‐Palme ,Adrian Paz ,Paola Bisignano,Parveen Goyal,Jeff Abramson,Elin Claesson,Rebecka Andersson,Konstantinos Beis,Michael Grabe,Anne Farewell,S Ramaswamy,Elin Dunevall,Lorena Pochini,Rhawnie Caing-Carlsson,Cesare Indiveri,Renwick C J Dobson,Ulf J Nilsson,Rosmarie Friemann

doi:10.1038/s41467-018-04045-7

Abstract

Many pathogenic bacteria utilise sialic acids as an energy source or use them as an external coating to evade immune detection. As such, bacteria that colonise sialylated environments deploy specific transporters to mediate import of scavenged sialic acids. Here, we report a substrate-bound 1.95 Å resolution structure and subsequent characterisation of SiaT, a sialic acid transporter from Proteus mirabilis. SiaT is a secondary active transporter of the sodium solute symporter (SSS) family, which use Na+ gradients to drive the uptake of extracellular substrates. SiaT adopts the LeuT-fold and is in an outward-open conformation in complex with the sialic acid N-acetylneuraminic acid and two Na+ ions. One Na+ binds to the conserved Na2 site, while the second Na+ binds to a new position, termed Na3, which is conserved in many SSS family members. Functional and molecular dynamics studies validate the substrate-binding site and demonstrate that both Na+ sites regulate N-acetylneuraminic acid transport.

Highlights

Campus Drive West Stanford, Stanford, CA 94305-5126, USA
Our results suggest that this subgroup of the sodium solute symporter (SSS) family utilise the binding energy of a second Na+ ion to allosterically stabilise the substrate without directly coordinating it as observed for endogenous ligand binding to LeuT20 and inhibitor binding to dDAT42 and SERT43
The simulations indicate that binding a second ion further pre-organises the binding site to increase substrate binding affinity, and it may play an important role in stabilising the outward-facing conformation

Summary

Introduction

Campus Drive West Stanford, Stanford, CA 94305-5126, USA. These authors contributed : Weixiao Y. The electron density in this site corresponds to Neu5Ac in its β-anomeric form (Fig. 1c) as seen in the Haemophilus influenzae periplasmic binding protein (SiaP) of the sialic acid TRAP system[29]. This is a common feature observed among interactions between viruses and sialic acid coated glycan molecules[34,35], as well as in otherwise polar active sites of proteins and enzymes that use Neu5Ac as a substrate[29,36].

Results

Conclusion