An N-terminal fragment of the gene 4 helicase/primase of bacteriophage T7 retains primase activity in the absence of helicase activity.
Primase and helicase activities of bacteriophage T7 are present in a single polypeptide coded by gene 4. Because the amino terminal region of the gene 4 protein contributes to primase activity, we constructed a truncated gene 4 encoding the N-terminal 271-aa residues. The truncated protein, purified from cells overexpressing the protein, is a dimer in solution; the full-length protein is a hexamer. Although the fragment is devoid of dTTPase and helicase activities, it catalyzes template-directed synthesis of di-, tri-, and tetranucleotides. The rates for tetraribonucleotide synthesis and for dinucleotide extension on a 20-nucleotide template are similar for the full-length and truncated proteins. However, the activity of the primase fragment is unaffected by dTTP whereas the primase activity of the full-length protein is stimulated >14-fold. The primase fragment is defective in the interaction with T7 DNA polymerase in that primer synthesis cannot be coupled to DNA synthesis.
Frick
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DN
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Baradaran
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K
Richardson
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CC
D017123:Bacteriophage T7 / Q000201:enzymology* / Q000235:genetics*; D004265:DNA Helicases / Q000235:genetics* / Q000378:metabolism; D019915:DNA Primase / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D014764:Viral Proteins / Q000235:genetics
D014764:Viral Proteins; D019915:DNA Primase; D004265:DNA Helicases