Hereditary folate malabsorption: A positively charged amino acid at position 113 of the proton-coupled folate transporter (PCFT/SLC46A1) is required for folic acid binding |
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Authors: | Inbal Lasry Fabian Glaser Yehuda G Assaraf |
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Institution: | a The Fred Wyszkowski Cancer Research Laboratory, Dept. of Biology, Technion-Israel Institute of Technology, Haifa 32000, Israel b Bioinformatics Knowledge Unit, The Lorry I. Lokey Interdisciplinary Center for Life Sciences and Engineering, Technion, Haifa 32000, Israel c Department of Rheumatology, VU University Medical Center, Amsterdam, The Netherlands |
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Abstract: | The proton-coupled folate transporter (PCFT/SLC46A1) mediates intestinal folate uptake at acidic pH. Some loss of folic acid (FA) transport mutations in PCFT from hereditary folate malabsorption (HFM) patients cluster in R113, thereby suggesting a functional role for this residue. Herein, unlike non-conservative substitutions, an R113H mutant displayed 80-fold increase in the FA transport Km while retaining parental Vmax, hence indicating a major fall in folate substrate affinity. Furthermore, consistent with the preservation of 9% of parental transport activity, R113H transfectants displayed a substantial decrease in the FA growth requirement relative to mock transfectants. Homology modeling based on the crystal structures of the Escherichia coli transporter homologues EmrD and glycerol-3-phosphate transporter revealed that the R113H rotamer properly protrudes into the cytoplasmic face of the minor cleft normally occupied by R113. These findings constitute the first demonstration that a basic amino acid at position 113 is required for folate substrate binding. |
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Keywords: | HFM hereditary folate malabsorption PCFT proton-coupled folate transporter FA folic acid MTX methotrexate |
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