Substrate translocation and inhibition in human dicarboxylate transporter NaDC3

Abstract

The human high-affinity sodium–dicarboxylate cotransporter (NaDC3) imports various substrates into the cell as tricarboxylate acid cycle intermediates, lipid biosynthesis precursors and signaling molecules. Understanding the cellular signaling process and developing inhibitors require knowledge of the structural basis of the dicarboxylate specificity and inhibition mechanism of NaDC3. To this end, we determined the cryo-electron microscopy structures of NaDC3 in various dimers, revealing the protomer in three conformations: outward-open Co, outward-occluded Coo and inward-open Ci. A dicarboxylate is first bound and recognized in Co and how the substrate interacts with NaDC3 in Coo likely helps to further determine the substrate specificity. A phenylalanine from the scaffold domain interacts with the bound dicarboxylate in the Coo state and modulates the kinetic barrier to the transport domain movement. Structural comparison of an inhibitor-bound structure of NaDC3 to that of the sodium-dependent citrate transporter suggests ways for making an inhibitor that is specific for NaDC3. The authors show cryo-electron microscopy structures of the human high-affinity sodium–dicarboxylate cotransporter, responsible for dicarboxylate import into the cell, in complex with various substrates and in different states establish the basis of substrate recognition, differentiation and release, as well as regulation of transport domain movement.