Voltage dependence of L-arginine transport by hCAT-2A and hCAT-2B expressed in oocytes from Xenopus laevis.
Membrane potential and currents were investigated with the two-electrode voltage-clamp technique in Xenopus laevis oocytes expressing hCAT-2A or hCAT-2B, the splice variants of the human cationic amino acid transporter hCAT-2. Both hCAT-2A- and hCAT-2B-expressing oocytes exhibited a negative extracellular L-arginine concentration ([L-Arg](o))-sensitive membrane potential, additive to the K(+) diffusion potential, when cells were incubated in Leibovitz medium (containing 1.45 mM L-Arg and 0.25 mM L-lysine). The two carrier proteins produced inward and outward currents, which were dependent on the L-Arg gradient and membrane potential. Ion substitution experiments showed that the hCAT-induced currents were independent of external Na(+), K(+), Ca(2+), or Mg(2+). The apparent Michaelis-Menten constant values at -60 mV, obtained from plots of L-Arg-induced currents against [L-Arg](o), were 0.97 and 0.13 mM in oocytes expressing hCAT-2A and hCAT-2B, respectively; maximal currents amounted to -194 +/- 8 and -84 +/- 2 nA, respectively. At saturating [L-Arg](o), the current-voltage relationships of hCAT-2A-expressing oocytes became steeper, yielding an additional conductance up to 2 microS/oocyte, whereas those of hCAT-2B-expressing oocytes were simply shifted to the right, resulting in voltage-independent difference currents. The distinct electrochemical properties of the two isoforms of hCAT-2 are assumed to contribute differentially to the membrane transport and the maintenance of cationic amino acids in various tissues.
Nawrath
H
H
Department of Pharmacology, Johannes Gutenberg University, 55101 Mainz, Germany. nawrath@mail.uni-mainz.de
Wegener
J W
JW
Rupp
J
J
Habermeier
A
A
Closs
E I
EI
D026923:Amino Acid Transport Systems, Basic; D000818:Animals; D001120:Arginine / Q000378:metabolism*; D001692:Biological Transport / Q000502:physiology; D002352:Carrier Proteins / Q000378:metabolism*; D004563:Electrochemistry; D004594:Electrophysiology; D005260:Female; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism; D020033:Protein Isoforms / Q000378:metabolism; D014982:Xenopus laevis
D026923:Amino Acid Transport Systems, Basic; D002352:Carrier Proteins; D008565:Membrane Proteins; D020033:Protein Isoforms; D001120:Arginine