Intracellular accumulation of L-Arg, kinetics of transport, and potassium leak conductance in oocytes from Xenopus laevis expressing hCAT-1, hCAT-2A, and hCAT-2B.
Cationic amino acid transporters play an important role in the intracellular supply of L-Arg and the generation of nitric oxide. Since the transport of L-Arg is voltage-dependent, we aimed at determining the intracellular L-Arg concentration and describing the transport of L-Arg in terms of Michaelis-Menten kinetics, taking into account membrane voltage. The human isoforms of the cationic amino acid transporters, hCAT-1, hCAT-2A, and hCAT-2B, were expressed in oocytes from Xenopus laevis and studied with the voltage clamp technique and in tracer experiments. We found that L-Arg was concentrated intracellularly by all hCAT isoforms and that influx and efflux, in the steady state of exchange, were nearly mirror images. Conductance measurements at symmetric concentrations of L-Arg (inside/outside) allowed us to determine KM and Vmax. The empty transporter of hCAT-2B featured an unexpected potassium conductance, which was inhibited by L-Arg.
Rotmann
Alexander
A
Department of Pharmacology, University of Mainz, Obere Zahlbacherstr 67, D-55101 Mainz, Germany.
Closs
Ellen I
EI
Liewald
Jana F
JF
Nawrath
Hermann
H
D026923:Amino Acid Transport Systems, Basic; D000818:Animals; D001120:Arginine / Q000737:chemistry / Q000378:metabolism*; D001692:Biological Transport; D027201:Cationic Amino Acid Transporter 1 / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D027202:Cationic Amino Acid Transporter 2 / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D007700:Kinetics; D008564:Membrane Potentials; D009865:Oocytes / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism*; D014982:Xenopus laevis / Q000235:genetics / Q000378:metabolism*
D026923:Amino Acid Transport Systems, Basic; D027201:Cationic Amino Acid Transporter 1; D027202:Cationic Amino Acid Transporter 2; C420825:SLC7A1 protein, human; C414177:SLC7A2 protein, human; D001120:Arginine; D011188:Potassium