Functional Characterization of Two Novel Mammalian Electrogenic Proton-dependent Amino Acid Cotransporters

• Published in: The Journal of biological chemistry 2002-06, Vol.277 (25), p.22966-22973
• Main Authors: Boll, Michael, Foltz, Martin, Rubio-Aliaga, Isabel, Kottra, Gabor, Daniel, Hannelore
• Format: Article
• Language: English
• Subjects: Alanine - chemistry; Amino Acid Sequence; Amino Acid Transport Systems, Neutral; Amino Acids - chemistry; Amino Acids - metabolism; Animals; Biological Transport; Blotting, Northern; Carrier Proteins - chemistry; Carrier Proteins - metabolism; Cloning, Molecular; DNA, Complementary - metabolism; Electrophysiology; Epithelial Cells - metabolism; Glycine - chemistry; Green Fluorescent Proteins; HeLa Cells; Humans; Hydrogen-Ion Concentration; Ions; Kinetics; Luminescent Proteins - metabolism; Membrane Proteins - chemistry; Membrane Proteins - metabolism; Membrane Transport Proteins; Molecular Sequence Data; Oocytes - metabolism; Patch-Clamp Techniques; Proline - chemistry; Protein Binding; Protein Structure, Tertiary; RNA, Complementary - metabolism; RNA, Messenger - metabolism; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Substrate Specificity; Sulfate Transporters; Symporters; Tissue Distribution; Transfection; Xenopus laevis
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M200374200

We cloned two cDNAs encoding proton/amino acid cotransporters, designated as mPAT1 and mPAT2, from murine tissues. They were identified by sequence similarity to the amino acid/auxin permease family member of lower eukaryotes. We functionally characterized both transporters by flux studies and electrophysiology after expression in Xenopus laevis oocytes. Both mPAT1 and mPAT2 induced a pH-dependent electrogenic transport activity for small amino acids (glycine, alanine, and proline) that is altered by membrane potential. Direct evidence for amino acid/H+-symport was shown by intracellular acidification, and a flux coupling stoichiometry for proline/H+-symport of 1:1 was determined for both transporters. Besides small apolar l-amino acids, the transporters also recognize their d-enantiomers and selected amino acid derivatives such as γ-aminobutyric acid. The mPAT1 transporter, the murine orthologue of the recently cloned rat LYAAT-1 transporter, can be considered as a low affinity system when compared with mPAT2. The mRNA of mPAT1 is highly expressed in small intestine, colon, kidney, and brain; the mPAT2-mRNA is mainly found in heart and lung. Phenotypically, the PAT1 transporter possesses the same functional characteristics as the previously described proton-dependent amino acid transport process in apical membranes of intestinal and renal epithelial cells.