Methotrexate recognition by the human reduced folate carrier SLC19A1.

Abstract

The human Reduced Folate Carrier (hRFC) is essential for folate uptake into mammalian cells. As a critical substrate in nucleotide biosynthesis and amino acid metabolism, folate is required for cell division. Consequently, cancerous cells rely heavily upon folate for their growth, so specific targeting by anti-folate pharmaceuticals is highly sought after. One such anti-folate chemotherapeutic is methotrexate (MTX), which requires hRFC for uptake into the cell and variants of which can lead to drug resistance and folate deficiency. Unravelling the transport mechanism of hRFC, as well as the basis for its interaction with folates and anti-folate drugs has important implications for chemotherapeutic development and personalized medicine. In our companion poster [“Cryo-EM structure determination of the human reduced folate carrier SLC19A1 in complex with methotrexate”] we detail the biochemical and structural techniques required for elucidating the structure of MTX-bound hRFC (65 kDa). Here we present transport assay data and MD simulations that build upon the structure to identify the features of hRFC that make it capable of transporting a wide range of anions, while also probing the molecular determinants of substrate specificity. This multifaceted study provides not only insight into the transport mechanism of hRFC but also a basis for rational chemotherapeutic drug design.