Structural basis of antifolate recognition and transport by PCFT

Folates (also known as vitamin B9) have a critical role in cellular metabolism as the starting point in the synthesis of nucleic acids, amino acids and the universal methylating agent S -adenylsmethionine 1 , 2 . Folate deficiency is associated with a number of developmental, immune and neurological disorders 3 – 5 . Mammals cannot synthesize folates de novo; several systems have therefore evolved to take up folates from the diet and distribute them within the body 3 , 6 . The proton-coupled folate transporter (PCFT) (also known as SLC46A1) mediates folate uptake across the intestinal brush border membrane and the choroid plexus 4 , 7 , and is an important route for the delivery of antifolate drugs in cancer chemotherapy 8 – 10 . How PCFT recognizes folates or antifolate agents is currently unclear. Here we present cryo-electron microscopy structures of PCFT in a substrate-free state and in complex with a new-generation antifolate drug (pemetrexed). Our results provide a structural basis for understanding antifolate recognition and provide insights into the pH-regulated mechanism of folate transport mediated by PCFT. Cryo-electron microscopy structures of PCFT in a substrate-free state and bound to the antifolate drug pemetrexed provide insights into how this protein recognizes folates and mediates their transport into cells.