TY - JOUR T1 - The Sodium Sialic Acid Symporter From Has Altered Substrate Specificity. JF - Front Chem Y1 - 2018 A1 - North, Rachel A A1 - Wahlgren, Weixiao Y A1 - Remus, Daniela M A1 - Scalise, Mariafrancesca A1 - Kessans, Sarah A A1 - Dunevall, Elin A1 - Claesson, Elin A1 - Soares da Costa, Tatiana P A1 - Perugini, Matthew A A1 - Ramaswamy, S A1 - Allison, Jane R A1 - Indiveri, Cesare A1 - Friemann, Rosmarie A1 - Dobson, Renwick C J AB -

Mammalian cell surfaces are decorated with complex glycoconjugates that terminate with negatively charged sialic acids. Commensal and pathogenic bacteria can use host-derived sialic acids for a competitive advantage, but require a functional sialic acid transporter to import the sugar into the cell. This work investigates the sodium sialic acid symporter (SiaT) from (SiaT). We demonstrate that SiaT rescues an strain lacking its endogenous sialic acid transporter when grown on the sialic acids -acetylneuraminic acid (Neu5Ac) or -glycolylneuraminic acid (Neu5Gc). We then develop an expression, purification and detergent solubilization system for SiaT and demonstrate that the protein is largely monodisperse in solution with a stable monomeric oligomeric state. Binding studies reveal that SiaT has a higher affinity for Neu5Gc over Neu5Ac, which was unexpected and is not seen in another SiaT homolog. We develop a homology model and use comparative sequence analyses to identify substitutions in the substrate-binding site of SiaT that may explain the altered specificity. SiaT is shown to be electrogenic, and transport is dependent upon more than one Na ion for every sialic acid molecule. A functional sialic acid transporter is essential for the uptake and utilization of sialic acid in a range of pathogenic bacteria, and developing new inhibitors that target these transporters is a valid mechanism for inhibiting bacterial growth. By demonstrating a route to functional recombinant SiaT, and developing the and assay systems, our work underpins the design of inhibitors to this transporter.

VL - 6 ER - TY - JOUR T1 - Substrate-bound outward-open structure of a Na-coupled sialic acid symporter reveals a new Na site. JF - Nat Commun Y1 - 2018 A1 - Wahlgren, Weixiao Y A1 - Dunevall, Elin A1 - North, Rachel A A1 - Paz, Aviv A1 - Scalise, Mariafrancesca A1 - Bisignano, Paola A1 - Bengtsson-Palme, Johan A1 - Goyal, Parveen A1 - Claesson, Elin A1 - Caing-Carlsson, Rhawnie A1 - Andersson, Rebecka A1 - Beis, Konstantinos A1 - Nilsson, Ulf J A1 - Farewell, Anne A1 - Pochini, Lorena A1 - Indiveri, Cesare A1 - Grabe, Michael A1 - Dobson, Renwick C J A1 - Abramson, Jeff A1 - Ramaswamy, S A1 - Friemann, Rosmarie AB -

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.

VL - 9 IS - 1 ER -