Two different reactions involved in the primer/template-independent polymerization of dATP and dTTP by Taq DNA polymerase.
Taq and Tth DNA polymerases catalyzed polymerization of dATP and dTTP into poly d(A-T) without requiring added primer/template (Hanaki et al., Biochem. Biophys. Res. Commun. 238, 113-118), while the Stoffel fragment of Taq DNA polymerase and delta Tth DNA 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 to N-ethylmaleimide than the elongation reaction, probably the formation of the oligonucleotide preceded the elongation. But when the substrates were depleted, Taq DNA 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 of Taq DNA 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.
Hanaki
K
K
Department of Bacteriology, Faculty of Medicine, University of Tokyo, Japan.
Odawara
T
T
Nakajima
N
N
Shimizu
Y K
YK
Nozaki
C
C
Mizuno
K
K
Muramatsu
T
T
Kuchino
Y
Y
Yoshikura
H
H
D004247:DNA / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D017931:DNA Primers / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D003838:Deoxyadenine Nucleotides / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D008970:Molecular Weight; D009694:Nucleic Acid Precursors / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D009841:Oligonucleotides / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D011108:Polymers / Q000378:metabolism*; D019914:Taq Polymerase / Q000378:metabolism*; D013696:Temperature; D013698:Templates, Genetic; D013942:Thymine Nucleotides / Q000378:metabolism*
D017931:DNA Primers; D003838:Deoxyadenine Nucleotides; D009694:Nucleic Acid Precursors; D009841:Oligonucleotides; D011108:Polymers; D013942:Thymine Nucleotides; D004247:DNA; D019914:Taq Polymerase; D004259:DNA-Directed DNA Polymerase; D005033:Ethylmaleimide; C024157:thymidine 5'-triphosphate