Initiation Complexes of Escherichia coli RNA Polymerase and T7 DNA

Complexes between Escherichia coli RNA polymerase and T7 DNA have been studied by three techniques measuring their stability either to 0.4 M (NH4)2SO4 or to rifampicin in 0.15 M KCl. In each case a nucleoside triphosphate was required for stabilisation, the order of efficiency in general being GTP > ATP > UTP ∼ 0. This did not correspond to the order with which the nucleotides initiate RNA synthesis (ATP > GTP ≫ CTP = UTP) contradicting the hypothesis of Anthony et al., which suggested that complexes formed in the presence of the 5′‐terminal nucleotide in the absence of polymerisation are stable to 0.4 M (NH4)2SO4. The complexes formed in the presence of [3H]GTP were shown to have tightly bound oligonucleotides. The degree of stabilisation produced by GTP could be reduced markedly by purification while in the case of ATP purification completely removed the ability to stabilise. These results suggest that the interpretation of previous experiments on the initiation of RNA synthesis may need to be revised.