title: The gene 59 protein of bacteriophage T4. Characterization of protein-protein interactions with gene 32 protein, the T4 single-stranded DNA binding protein.
abstract: The gene 59 protein (gp59) of bacteriophage T4 stimulates the activities of gene 41 protein (gp41), the T4 replicative DNA helicase, by promoting the assembly of gp41 onto single-stranded (ss)-DNA molecules that are covered with cooperatively bound gene 32 protein (gp32). This helicase-ssDNA assembly process, which is important for the reconstitution of the primosome component of the T4 DNA replication fork, appears to require both gp59-gp41 and gp59-gp32 protein-protein interactions. In this study we characterize the physical and functional interactions of gp59 with gp32, the T4 ssDNA-binding protein. Experimental results presented herein indicate: 1) that gp59 binds specifically to both free and ssDNA-bound gp32 molecules; and 2) that in both cases binding involves contacts between gp59 and the acidic C-terminal domain of gp32 (the so-called "A-domain"). We further show that single-stranded DNA molecules coated with (gp32-A), a truncated form of gp32 lacking the A-domain, are refractory to gp59-dependent helicase assembly. The data indicate that specific contacts between gp59 molecules and the A-domains of gp32 molecules are essential for gp59-dependent assembly of gp41 onto gp32-ssDNA complexes. Our results are consistent with a model in which gp59 binds to gp32 molecules within the gp32-ssDNA complex and therein forms a target site for helicase-ssDNA assembly.
authors.affiliation: Department of Biochemistry, University of Vermont College of Medicine, Burlington, Vermont 05405, USA.
authors.forename: S W
authors.initials: SW
authors.lastname: Morrical
authors.forename: H T
authors.initials: HT
authors.lastname: Beernink
authors.forename: A
authors.initials: A
authors.lastname: Dash
authors.forename: K
authors.initials: K
authors.lastname: Hempstead
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D017122:Bacteriophage T4 / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D002846:Chromatography, Affinity; D004265:DNA Helicases / Q000378:metabolism; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005814:Genes, Viral; D011485:Protein Binding; D011995:Recombination, Genetic; D014764:Viral Proteins / Q000378:metabolism*; D015678:Viral Structural Proteins / Q000235:genetics; D014779:Virus Replication
chemical_list: D004277:DNA, Single-Stranded; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D014764:Viral Proteins; D015678:Viral Structural Proteins; C091225:gene 59 protein, Enterobacteria phage T4; C060035:gp32 protein, Enterobacteria phage T4; D000251:Adenosine Triphosphatases; D004265:DNA Helicases