Two Different Reactions Involved in the Primer/Template-Independent Polymerization of dATP and dTTP byTaqDNA Polymerase

Taqand Tth DNA polymerases catalyzed polymerization of dATP and dTTP into poly d(A-T) without requiring added primer/template (Hanakiet al., Biochem. Biophys. Res. Commun.238, 113–118), while the Stoffel fragment ofTaqDNA polymerase and ΔTthDNA polymerase with respective deletions of ca. 290 and 250 N-terminal amino acids did not. The primer/template-independent polymerization appeared to proceed via two reactions, the slow process of formation of 16–19 nt long oligo d(A-T) without primer/template and the rapid process of elongation of the oligo d(A-T) by self-priming. As the former step was more sensitive toN-ethylmaleimide than the elongation reaction, probably the formation of the oligonucleotide preceded the elongation. But when the substrates were depleted,TaqDNA polymerase degraded the high molecular weigh d(A-T) polymer to the oligomers which were resistant to the further digestion by the 5′ → 3′ exonuclease activity ofTaqDNA polymerase. Probably, the elongation and the degradation reactions proceeded simultaneously, the former process being faster than the latter in the presence of enough dATP and dTTP.