Effect of single-stranded DNA-binding proteins on the helicase and primase activities of the bacteriophage T7 gene 4 protein.
Gene 4 protein (gp4) of bacteriophage T7 provides two essential functions at the T7 replication fork, primase and helicase activities. Previous studies have shown that the single-stranded DNA-binding protein of T7, encoded by gene 2.5, interacts with gp4 and modulates its multiple functions. To further characterize the interactions between gp4 and gene 2.5 protein (gp2.5), we have examined the effect of wild-type and altered gene 2.5 proteins as well as Escherichia coli single-stranded DNA-binding (SSB) protein on the ability of gp4 to synthesize primers, hydrolyze dTTP, and unwind duplex DNA. Wild-type gp2.5 and E. coli SSB protein stimulate primer synthesis and DNA-unwinding activities of gp4 at low concentrations but do not significantly affect single-stranded DNA-dependent hydrolysis of dTTP. Neither protein inhibits the binding of gp4 to single-stranded DNA. The variant gene 2.5 proteins, gp2.5-F232L and gp2.5-Delta26C, inhibit primase, dTTPase, and helicase activities proportional to their increased affinities for DNA. Interestingly, wild-type gp2.5 stimulates the unwinding activity of gp4 except at very high concentrations, whereas E. coli SSB protein is highly inhibitory at relative low concentrations.
He
Zheng-Guo
ZG
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Richardson
Charles C
CC
D017123:Bacteriophage T7 / Q000201:enzymology*; D001483:Base Sequence; D002384:Catalysis; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004265:DNA Helicases / Q000378:metabolism*; D019915:DNA Primase / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D014764:Viral Proteins / Q000378:metabolism
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D009841:Oligonucleotides; D014764:Viral Proteins; C085913:gene 2.5 protein, Enterobacteria phage T7; D004247:DNA; D019915:DNA Primase; D004265:DNA Helicases