JBC, Vol. 252, Issue 7, 2304-2310, Apr, 1977
Initiation of synthesis of messenger RNA of deoxynucleotide kinase by oligoribonucleotides
P. J. Natale and J. M. Buchanan
The effects of nucleoside triphosphates and oligoribonucleotides on the
initiation of synthesis of messenger RNA of the T4 phage-specific enzyme,
deoxynucleotide kinase, have been studied. The procedure involved
incubation of T4 DNA, purified RNA polymerase from Escherichia coli, and
selected nucleotide compounds during a brief period to permit initiation of
RNA synthesis. Further initiation was arrested by the addition of
ribampicin, and completion of the transcription of the newly initiated RNA
was permitted to take place in the presence of the full complement of
nucleoside triphosphates. After translation of the messenger RNA into
phage-specific enzymes, the measured activities of the latter whe first
incubation period. The effectiveness of individual nucleoside
triphosphates, when present singly or in combination during the initiation
period, was compared to that when all four nucleoside triphosphates were
available. ATP alone was extremely effective as an initiator of the
synthesis of the messenger RNA for deoxynucleotide kinase. The addition of
UTP to ATP not only enhanced the magnitude of initiation but also affected
the kinetics of ATP interaction with T4 DNA and RNA polymerase during the
initiation period. Several oligoribonucleotides including a series ApA to
ApApApA, UpU to UpUpUpU, and the heteropolymers, Ap1pU and ApApApU, were
tested as initiators of kinase mRNA synthesis. A sequence of nucleotides in
the promoter region of T4 DNA for the deoxynucleotide kinase gene has been
proposed as a result of these experiments.
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