Active transport of L-proline in the protozoan parasite Trypanosoma brucei brucei

Biochem J. 1993 Apr 1;291 ( Pt 1)(Pt 1):297-301. doi: 10.1042/bj2910297.


The characteristics of L-proline transport in the procyclic form of Trypanosoma brucei were studied by using L-[14C]proline and a quick separation technique by centrifugation through an oil mixture. L-Proline uptake displayed typical Michaelis-Menten kinetics, with a Km of 19 microM and a maximum transport velocity of 17 nmol/min per 10(8) cells at 27 degrees C. The maximum concentration gradient factor obtained after 1 min of incubation was 270-fold in 0.02 mM proline. Cells permeabilized with 80 microM digitonin were still able to accumulate 14C label, but to a lower extent. The temperature-dependence of proline uptake gave an apparent activation energy of 74.9 kJ.mol-1. In competition studies with a 10-fold excess of structural analogues, L-alanine, L-cysteine and L-azetidine-2-carboxylate were found to inhibit L-proline uptake. Variation of pH or addition of the protonophore carbonyl cyanide m-chlorophenylhydrazone ('CCCP') did not affect proline transport, showing that it is not driven by a protonmotive force. The absence of Na+, with or without monensin, did not affect proline transport. The absence of K+ and the addition of the Na+,K(+)-ATPase inhibitor ouabain had no significant effect on proline uptake activity. The thiol-modifying reagent iodoacetate (10 mM) decreased proline uptake by half. KCN (1 mM) inhibited proline uptake to a lesser extent, and the degree of inhibition was proportional to the intracellular ATP concentration. Preliminary experiments on proline transport in plasma-membrane vesicles of the cells, using a filtration technique, showed an uptake of proline (0.67 nmol/mg of protein) by the vesicles, but only in the presence of intravesicular ATP. The results thus obtained suggest that the proline carrier system in T. brucei is ATP-driven and independent of Na+, K+ or H+ co-transport.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Alanine / pharmacology
  • Animals
  • Azetidinecarboxylic Acid / pharmacology
  • Binding, Competitive
  • Biological Transport, Active / drug effects
  • Cell Membrane Permeability / drug effects
  • Cysteine / pharmacology
  • Digitonin / pharmacology
  • Hydrogen-Ion Concentration
  • Hydroxyproline / pharmacology
  • Iodoacetates / pharmacology
  • Iodoacetic Acid
  • Kinetics
  • Potassium Cyanide / pharmacology
  • Proline / metabolism*
  • Temperature
  • Thermodynamics
  • Trypanosoma brucei brucei / metabolism*


  • Iodoacetates
  • Azetidinecarboxylic Acid
  • Adenosine Triphosphate
  • Proline
  • Cysteine
  • Digitonin
  • Potassium Cyanide
  • Alanine
  • Hydroxyproline
  • Iodoacetic Acid