Cryo-EM structures reveal the H+/citrate symport mechanism ofDrosophilaINDY

• DOI: 10.26508/lsa.202402992
• Publication Date: 2025-01-30T17:20:19Z
• Authors: Subin Kim; Jun Gyou Park; Seung Hun Choi; Ji Won Kim; Mi Sun Jin

DrosophilaI’m Not Dead Yet (INDY) functions as a transporter for citrate, a key metabolite in the citric acid cycle, across the plasma membrane. Partial deficiency of INDY extends lifespan, akin to the effects of caloric restriction. In this work, we use cryo-electron microscopy to determine structures of INDY in the presence and absence of citrate and in complex with the well-known inhibitor 4,4′-diisothiocyano-2,2′-disulfonic acid stilbene (DIDS) at resolutions ranging from 2.7 to 3.6 Å. Together with functional data obtained in vitro, the INDY structures reveal the H+/citrate co-transport mechanism, in which aromatic residue F119 serves as a one-gate element. They also provide insight into how protein–lipid interactions at the dimerization interface affect the stability and function of the transporter, and how DIDS disrupts the transport cycle.