Evidence is presented that in Escherichia coli made permeable to nucleotides by exposure to toluene, the synthesis of a DNA chain complementary to the infecting single stranded DNA of bacteriophage phiX174 requires ATP as well as the four deoxyribonucleoside triphosphates. This synthesis results in the formation of the parental double stranded replicative form (RF) molecule. The ATP is not required simply to prevent degradation of the ribonucleoside or deoxyribonucleoside triphosphates; it can be partially substituted for by other ribonucleoside triphosphates. No single one of the known E. coli DNA polymerases appears to be uniquely responsible in vivo for the formation of the parental RF. Since phiX replicates well in strains lacking all, or almost all, of the in vitro activities of DNA polymerases I and II, neither of these two enzymes would seem essential; and in a temperature sensitive E. coli mutant [dnaE(ts)] deficient in DNA polmerase I activity and possessing a temperature sensitive DNA polymerase III, the viral single stranded DNA is efficiently incorporated into an RF molecule at the restrictive temperature. In contrast, both RF replication and progeny single stranded DNA synthesis are dependent upon DNA polymerase III activity.
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