Reconstitution into liposomes and functional characterization of the carnitine transporter from renal cell plasma membrane

Biochim Biophys Acta. 2004 Feb 10;1661(1):78-86. doi: 10.1016/j.bbamem.2003.12.001.


The carnitine transporter was solubilized from rat renal apical plasma membrane (brush-border membrane) with C12E8 and reconstituted into liposomes by removing the detergent from mixed micelles by hydrophobic chromatography on Amberlite XAD-4. The reconstitution was optimised with respect to the protein concentration, the detergent/phospholipid ratio and the number of passages through a single Amberlite column. The reconstituted carnitine transporter catalysed a first-order antiport reaction (carnitine/carnitine or carnitine/substrate) stimulated by external, not internal, Na+, with a positive cooperativity. Na+ was co-transported with carnitine. Optimal activity was found between pH 5.5 and pH 6.0. The sulfhydryl reagents MTSES, MTSET and mercurials strongly inhibited the transport. Substrate analogues inhibited the transport; the most effective were acylcarnitines and betaine, followed by dimethylglicine, tetraethylammonium and arginine. Besides carnitine, only acylcarnitines and betaine were efficiently translocated. The Km for carnitine on the external and internal side of the transporter was 0.08 and 1.2 mM, respectively. The transporter is asymmetrical and it is unidirectionally inserted into the proteoliposomal membrane with an orientation corresponding to that of the native membrane. The reconstituted carnitine transporter corresponds, very probably, to the OCTN2 protein.

MeSH terms

  • Animals
  • Biological Transport
  • Carnitine / metabolism
  • Carrier Proteins / antagonists & inhibitors
  • Carrier Proteins / chemistry
  • Carrier Proteins / metabolism*
  • Hydrogen-Ion Concentration
  • Intracellular Membranes / chemistry
  • Intracellular Membranes / metabolism*
  • Kidney Tubules / metabolism*
  • Kidney Tubules / ultrastructure
  • Liposomes / chemistry*
  • Membrane Proteins / chemistry
  • Membrane Proteins / metabolism*
  • Microvilli / metabolism
  • Organic Cation Transport Proteins*
  • Proteolipids / metabolism
  • Rats
  • Sodium Chloride
  • Sodium Radioisotopes
  • Solute Carrier Family 22 Member 5
  • Tritium


  • Carrier Proteins
  • Liposomes
  • Membrane Proteins
  • Organic Cation Transport Proteins
  • Proteolipids
  • SLC22A5 protein, human
  • Slc22a5 protein, rat
  • Sodium Radioisotopes
  • Solute Carrier Family 22 Member 5
  • proteoliposomes
  • Tritium
  • Sodium Chloride
  • Carnitine