Document Detail


A novel loss-of-function mutation in the proton-coupled folate transporter from a patient with hereditary folate malabsorption reveals that Arg 113 is crucial for function.
MedLine Citation:
PMID:  18559978     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Hereditary folate malabsorption (HFM) patients harbor inactivating mutations including R113S in the proton-coupled folate transporter (PCFT), an intestinal folate transporter with optimal activity at acidic pH. Here we identified and characterized a novel R113C mutation residing in the highly conserved first intracellular loop of PCFT. Stable transfectants overexpressing a Myc-tagged wild-type (WT) and mutant R113C PCFT displayed similar transporter targeting to the plasma membrane. However, whereas WT PCFT transfectants showed a 22-fold increase in [(3)H]folic acid influx at pH 5.5, R113C or mock transfectants showed no increase. Moreover, WT PCFT transfectants displayed a 50% folic acid growth requirement concentration of 7 nM, whereas mock and R113C transfectants revealed 24- to 27-fold higher values. Consistently, upon fluorescein-methotrexate labeling, WT PCFT transfectants displayed a 50% methotrexate displacement concentration of 50 nM, whereas mock and R113C transfectants exhibited 12- to 14-fold higher values. Based on the crystal structure of the homologous Escherichia coli glycerol-3-phosphate transporter, we propose that the cationic R113 residue of PCFT is embedded in a hydrophobic pocket formed by several transmembrane helices that may be part of a folate translocation pore. These findings establish a novel loss of function mutation in HFM residing in an intracellular loop of PCFT crucial for folate transport.
Authors:
Inbal Lasry; Bluma Berman; Rachel Straussberg; Yael Sofer; Hanna Bessler; Mohamad Sharkia; Fabian Glaser; Gerrit Jansen; Stavit Drori; Yehuda G Assaraf
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Publication Detail:
Type:  Case Reports; In Vitro; Journal Article; Research Support, Non-U.S. Gov't     Date:  2008-06-17
Journal Detail:
Title:  Blood     Volume:  112     ISSN:  1528-0020     ISO Abbreviation:  Blood     Publication Date:  2008 Sep 
Date Detail:
Created Date:  2008-08-26     Completed Date:  2008-09-30     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  7603509     Medline TA:  Blood     Country:  United States    
Other Details:
Languages:  eng     Pagination:  2055-61     Citation Subset:  AIM; IM    
Affiliation:
The Fred Wyszkowski Cancer Research Laboratory, Department of Biology, Technion-Israel Institute of Technology, Haifa, Israel.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Animals
Arginine / chemistry
Base Sequence
Binding Sites / genetics
CHO Cells
Carrier Proteins / chemistry,  genetics*,  metabolism
Cell Membrane / metabolism
Child
Consanguinity
Cricetinae
Cricetulus
DNA Primers / genetics
Folic Acid / metabolism*
Homozygote
Humans
Hydrogen-Ion Concentration
Malabsorption Syndromes / genetics*,  metabolism*
Methotrexate / metabolism
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Point Mutation*
Protein Conformation
Receptors, Cell Surface / chemistry,  deficiency,  genetics*,  metabolism
Recombinant Proteins / chemistry,  genetics,  metabolism
Sequence Homology, Amino Acid
Transfection
Chemical
Reg. No./Substance:
0/Carrier Proteins; 0/DNA Primers; 0/Receptors, Cell Surface; 0/Recombinant Proteins; 0/folate-binding protein; 59-05-2/Methotrexate; 59-30-3/Folic Acid; 74-79-3/Arginine

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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