title: Characterization of the bacteriophage T4 gene 41 DNA helicase.
abstract: The T4 gene 41 protein and the gene 61 protein function together as a primase-helicase within the seven protein bacteriophage T4 multienzyme complex that replicates duplex DNA in vitro. We have previously shown that the 41 protein is a 5' to 3' helicase that requires a single-stranded region on the 5' side of the duplex to be unwound and is stimulated by the 61 protein (Venkatesan, M., Silver L. L., and Nossal, N. G. (1982) J. biol. Chem. 257, 12426-12434). The 41 protein, in turn, is required for pentamer primer synthesis by the 61 protein. We now show that the 41 protein helicase unwinds a partially duplex DNA molecule containing a performed fork more efficiently than a DNA molecule without a fork. Optimal helicase activity requires greater than 29 nucleotides of single-stranded DNA on the 3' side of the duplex (analogous to the leading strand template). This result suggests the 41 protein helicase interacts with the leading strand template as well as the lagging strand template as it unwinds the duplex region at the replication fork. As the single-stranded DNA on the 3' side of a short duplex (51 base pairs) is lengthened, the stimulation of the 41 protein helicase by the 61 protein is diminished. However, both the 61 protein and a preformed fork are essential for efficient unwinding of longer duplex regions (650 base pairs). These findings suggest that the 61 protein promotes both the initial unwinding of the duplex to form a fork and subsequent unwinding of longer duplexes by the 41 protein. A stable protein-DNA complex, detected by a gel mobility shift of phi X174 single-stranded DNA, requires both the 41 and 61 proteins and a rNTP (preferably rATP or rGTP, the nucleotides with the greatest effect on the helicase activity). In the accompanying paper, we report the altered properties of a proteolytic fragment of the 41 protein helicase and its effect on in vitro DNA synthesis in the T4 multienzyme replication system.
authors.affiliation: Section on Nucleic Acid Biochemistry, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland 20892.
authors.forename: R W
authors.initials: RW
authors.lastname: Richardson
authors.forename: N G
authors.initials: NG
authors.lastname: Nossal
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D010584:Bacteriophage phi X 174 / Q000235:genetics; D003570:Cytidine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D004265:DNA Helicases / Q000378:metabolism*; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D007700:Kinetics; D009690:Nucleic Acid Conformation; D013379:Substrate Specificity; D013604:T-Phages / Q000201:enzymology*; D014544:Uridine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D004277:DNA, Single-Stranded; D004279:DNA, Viral; D014764:Viral Proteins; D003570:Cytidine Triphosphate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D004259:DNA-Directed DNA Polymerase; D004265:DNA Helicases; D014544:Uridine Triphosphate