Active transport of L-proline in Trypanosoma cruzi.
L-proline is the main energy source in insect vector stages of most trypanosomatids, including Trypanosoma cruzi epimastigotes. This is the first biochemical description of two active proline transporter systems in T. cruzi. Uptake of this amino acid occurred by a low affinity system B and a high affinity system A. System B consistently appeared more specific than System A when excess competing amino acids were used in transport inhibition assays. Furthermore, the high affinity system is 70% inhibited by L-tryptophan, but the low affinity system is not. Both systems were found to be insensitive to the intracellular proline concentration and D-proline did not inhibit L-proline uptake showing that both systems are stereospecific. Both systems were Na+ and K+ independant but dependant on energy since ATP depletion impairs L-proline uptake. The combined action of carbonyl cyanide p-trifluoromethoxyphenyl hydrazone (FCCP) and oligomycin, and the dependence of activity on pH, further differentiated between the two systems leading to the conclusion that the high affinity system is a H+ gradient-dependant transporter whereas the low affinity system depends directly on ATP.
Silber
Ariel M
AM
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Caixa Postal 26077, São Paulo 05513-970, Brazil.
Tonelli
Renata R
RR
Martinelli
Marcela
M
Colli
Walter
W
Alves
Maria Júlia M
MJ
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D001693:Biological Transport, Active; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011392:Proline / Q000378:metabolism*; D013816:Thermodynamics; D014349:Trypanosoma cruzi / Q000254:growth & development / Q000378:metabolism*
D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D000255:Adenosine Triphosphate; D011392:Proline