A functional chimeric DNA primase: the Cys4 zinc-binding domain of bacteriophage T3 primase fused to the helicase of bacteriophage T7.
Two colinear bacteriophage T7 gene 4 proteins provide helicase and primase functions in vivo. T7 primase differs from T7 helicase by an additional 63 residues at the amino terminus. This terminal domain contains a zinc-binding motif which mediates an interaction with the basic primase recognition sequence 3'-CTG-5'. We have generated a chimeric primase in which the 81 amino-terminal residues are derived from the primase of phage T3 and the 484 carboxyl-terminal residues are those of phage T7 helicase. The amino-terminal domain of T3 primase is 50% homologous with that of T7 primase. The resulting T3/T7 chimeric protein is a functional primase in vivo. While the primase activity of the purified protein is about one-third that of T7 primase, the recognition sites used and the oligoribonucleotides synthesized from these sites are identical. We conclude that the residues responsible for the interaction with the sequence 3'-CTG-5' are conserved between the chimeric and T7 proteins.
Hine
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AV
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115.
Richardson
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CC
D000595:Amino Acid Sequence; D017903:Bacteriophage T3 / Q000201:enzymology*; D017123:Bacteriophage T7 / Q000201:enzymology*; D001483:Base Sequence; D003001:Cloning, Molecular; D003545:Cysteine; D004265:DNA Helicases / Q000378:metabolism*; D019915:DNA Primase; D004261:DNA Replication; D005814:Genes, Viral; D016679:Genome, Viral; D007700:Kinetics; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000096:biosynthesis; D012316:RNA Nucleotidyltransferases / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D016335:Zinc Fingers
D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D019915:DNA Primase; D012316:RNA Nucleotidyltransferases; D004265:DNA Helicases; D003545:Cysteine