Two DNA polymerases from Trypanosoma cruzi: biochemical characterization and effects of inhibitors.
1. Two DNA polymerases have been partially purified from Trypanosoma cruzi epimastigotes by DEAE-cellulose, phosphocellulose, and DNA agarose chromatography. 2. Both enzyme activities were characterized by several biochemical criteria. 3. They showed different sensitivity to KCl and displayed characteristic Mg2+ and Mn2+ requirements, although they exhibited almost identical primer-template utilization. 4. The preferred substrates were poly dC-oligo dG, activated calf thymus DNA, and poly dT-oligo rA. 5. Both enzyme fractions are not inhibited by aphidicolin while N-ethylmaleimide and phosphonacetic acid inhibited them to different extents. 6. ButylphenyldGTP strongly inhibited T. cruzi enzyme fraction I while it had no effect on enzyme fraction III. 7. This dGTP analog also inhibited the poly dT-directed polymerization of dAMP as described for other mammalian DNA polymerases. Kinetic studies indicated that butylphenyldGTP inhibited enzyme fraction I in a non-competitive fashion.
Departamento de Bioquímica, Facultad de Medicina, Universidad de Chile, Santiago.
C
C
Rojas
J
J
Venegas
S
S
Litvak
A
A
Solari
D000818:Animals; D002413:Cations, Divalent / Q000494:pharmacology; D002847:Chromatography, Agarose; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000302:isolation & purification; D019384:Nucleic Acid Synthesis Inhibitors; D014349:Trypanosoma cruzi / Q000201:enzymology*
D002413:Cations, Divalent; D019384:Nucleic Acid Synthesis Inhibitors; D004259:DNA-Directed DNA Polymerase