The gene 59 protein of bacteriophage T4. Characterization of protein-protein interactions with gene 32 protein, the T4 single-stranded DNA binding protein.
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.
Morrical
S W
SW
Department of Biochemistry, University of Vermont College of Medicine, Burlington, Vermont 05405, USA.
Beernink
H T
HT
Dash
A
A
Hempstead
K
K
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
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