This study presents cryo-electron microscopy (cryo-EM) structures of human GLUT9 in complex with urate and its inhibitor apigenin (API) at resolutions of 3.5 Å and 3.3 Å, respectively. The structures reveal that GLUT9 adopts an inward-open conformation, with the substrate binding pocket facing the intracellular side. These findings elucidate the molecular basis for GLUT9's preference for urate over glucose and demonstrate that API acts as a competitive inhibitor by occupying the substrate binding site. The structures also highlight the interactions between GLUT9 and urate, including hydrogen bonds and hydrophobic interactions, which are critical for urate recognition. Additionally, the study shows that API binds to GLUT9 in the substrate binding pocket, stabilizing through polar and hydrophobic interactions. The results provide insights into the mechanism of urate transport and the inhibitory effect of API on GLUT9, offering a foundation for the development of specific inhibitors targeting GLUT9 for the treatment of gout and hyperuricemia. The study also identifies key residues in GLUT9 that are essential for urate recognition and transport, and highlights the importance of these residues in determining the transporter's substrate specificity. The findings contribute to a deeper understanding of the structural and functional properties of GLUT9, which is a key player in urate transport and the pathogenesis of gout.This study presents cryo-electron microscopy (cryo-EM) structures of human GLUT9 in complex with urate and its inhibitor apigenin (API) at resolutions of 3.5 Å and 3.3 Å, respectively. The structures reveal that GLUT9 adopts an inward-open conformation, with the substrate binding pocket facing the intracellular side. These findings elucidate the molecular basis for GLUT9's preference for urate over glucose and demonstrate that API acts as a competitive inhibitor by occupying the substrate binding site. The structures also highlight the interactions between GLUT9 and urate, including hydrogen bonds and hydrophobic interactions, which are critical for urate recognition. Additionally, the study shows that API binds to GLUT9 in the substrate binding pocket, stabilizing through polar and hydrophobic interactions. The results provide insights into the mechanism of urate transport and the inhibitory effect of API on GLUT9, offering a foundation for the development of specific inhibitors targeting GLUT9 for the treatment of gout and hyperuricemia. The study also identifies key residues in GLUT9 that are essential for urate recognition and transport, and highlights the importance of these residues in determining the transporter's substrate specificity. The findings contribute to a deeper understanding of the structural and functional properties of GLUT9, which is a key player in urate transport and the pathogenesis of gout.