Mutational Analysis of the T4 Gp59 Helicase Loader Reveals Its Sites for Interaction with Helicase, Single-stranded Binding Protein, and DNA

Abstract

Background: Replication requires coordinated interactions among proteins/DNA. Results: Mutational analysis of T4 helicase loader, gp59, identifies domains that interact with DNA and replication factors. Conclusion: The high mobility group motif of gp59 interacts with the DNA fork; gp59 C-terminal surfaces interact with helicase and single-stranded binding protein.Significance: Processive replication may involve switching of gp59 between interactions with helicase and the single-stranded binding protein.

Models for the Binary Complex of Bacteriophage T4 Gp59 Helicase Loading Protein

“…The ternary complex models are consistent with both affinity analysis and mutagenesis studies of gp59 protein presented in a accompanying article (38). Native agarose gel electrophoresis results were in qualitative agreement with other studies (15,16), indicating binding of the gp59 protein to both the gp32 full-length and the gp32⌬B-truncated proteins, but not to gp32⌬A and gp32⌬〈B proteins.…”

“…The mutational analysis, described in the accompanying article (38), confirms the significance of the HMG-like domain previously used to generate the model for gp59 protein binding to pseudo-Y junction DNA, which proposed binding sites for the gp32 protein and the gp41 helicase. The mutational analysis indicates that gp41 helicase binds to the C-terminal region of the gp59 protein.…”

“…It appears that the gp32⌬B protein becomes more compact upon interaction with the gp59 protein. This model of the ternary complex combines the protein structure derived from the SAXS envelope with the mutational analysis of gp59 protein provided in our accompanying article (38).…”