Role of bacteriophage T7 DNA primase in the initiation of DNA strand synthesis

Abstract

Bacteriophage T7 DNA primase (gene-4 protein, 66,000 daltons) enables T7 DNA polymerase to initiate the synthesis of DNA chains on single-stranded templates. An initial step in the process of chain initiation is the formation of an oligoribonucleotide primer by T7 primase. The enzyme, in the presence of natural SS DNA, Mg++ (or Mn++), ATP and CTP (or a mixture of all 4 rNTPs), catalyzes the synthesis of di-, tri-, and tetraribonucleotides all starting at the 5' terminus with pppA. In a subsequent step requiring both T7 DNA polymerase and primase, the short oligoribonucleotides (predominantly pppA-C-C-AOH) are extended by covalent addition of deoxyribonucleotides. With the aid of primase, T7 DNA polymerase can also utilize efficiently a variety of synthetic tri-, tetra-, or pentanucleotides as chain initiators. T7 primase apparently plays an active role in primer extension by stabilizing the short primer segments in a duplex state on the template DNA.