Steady-state kinetic characterization of the mouse B(0)AT1 sodium-dependent neutral amino acid transporter

Pflugers Arch. 2005 Nov;451(2):338-48. doi: 10.1007/s00424-005-1455-x. Epub 2005 Aug 26.

Abstract

The members of the neurotransmitter transporter family SLC6A exhibit a high degree of structural homology; however differences arise in many aspects of their transport mechanisms. In this study we report that mouse B(0)AT1 (mouse Slc6a19) mediates the electrogenic transport of a broad range of neutral amino acids but not of the chemically similar substrates transported by other SLC6A family members. Cotransport of L: -Leu and Na(+) generates a saturable, reversible, inward current with Michaelis-Menten kinetics (Hill coefficient approximately 1) yielding a K(0.5) for L: -Leu of 1.16 mM and for Na(+) of 16 mM at a holding potential of -50 mV. Changing the membrane voltage influences both substrate binding and substrate translocation. Li(+) can substitute partially for Na(+) in the generation of L: -Leu-evoked inward currents, whereas both Cl(-) and H(+) concentrations influence its magnitude. The simultaneous measurement of charge translocation and L: -Leu uptake in the same cell indicates that B(0)AT1 transports one Na(+) per neutral amino acid. This appears to be accomplished by an ordered, simultaneous mechanism, with the amino acid binding prior to the Na(+), followed by the simultaneous translocation of both co-substrates across the plasma membrane. From this kinetic analysis, we conclude that the relatively constant [Na(+)] along the renal proximal tubule both drives the uptake of neutral amino acids via B(0)AT1 thermodynamically and ensures that, upon binding, these are translocated efficiently into the cell.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Transport Systems / drug effects
  • Amino Acid Transport Systems / genetics
  • Amino Acid Transport Systems / physiology*
  • Amino Acid Transport Systems, Neutral / genetics
  • Amino Acid Transport Systems, Neutral / physiology*
  • Amino Acids / metabolism*
  • Animals
  • Biological Transport / drug effects
  • Chlorides / pharmacology
  • Electrophysiology
  • Female
  • Genetic Vectors / genetics
  • Gluconates / pharmacology
  • Glycine / metabolism
  • Hydrogen-Ion Concentration
  • Kidney Tubules / enzymology
  • Kinetics
  • Leucine / metabolism
  • Lithium / metabolism
  • Lithium / pharmacology
  • Mice
  • Oocytes / metabolism
  • Patch-Clamp Techniques
  • Phenylalanine / metabolism
  • Sodium / metabolism
  • Transfection
  • Xenopus laevis

Substances

  • Amino Acid Transport Systems
  • Amino Acid Transport Systems, Neutral
  • Amino Acids
  • Chlorides
  • Gluconates
  • SLC6A19 protein, mouse
  • Phenylalanine
  • Lithium
  • Sodium
  • Leucine
  • gluconic acid
  • Glycine