title: Interaction of T4 UvsW helicase and single-stranded DNA binding protein gp32 through its carboxy-terminal acidic tail.
abstract: Bacteriophage T4 UvsW helicase contains both unwinding and annealing activities and displays some functional similarities to bacterial RecG and RecQ helicases. UvsW is involved in several DNA repair pathways, playing important roles in recombination-dependent DNA repair and the reorganization of stalled replication forks. The T4 single-stranded DNA (ssDNA) binding protein gp32 is a central player in nearly all DNA replication and repair processes and is thought to facilitate their coordination by recruiting and regulating the various proteins involved. Here, we show that the activities of the UvsW protein are modulated by gp32. UvsW-catalyzed unwinding of recombination intermediates such as D-loops and static X-DNA (Holliday junction mimic) to ssDNA products is enhanced by the gp32 protein. The enhancement requires the presence of the protein interaction domain of gp32 (the acidic carboxy-terminus), suggesting that a specific interaction between UvsW and gp32 is required. In the absence of this interaction, the ssDNA annealing and ATP-dependent translocation activities of UvsW are severely inhibited when gp32 coats the ssDNA lattice. However, when UvsW and gp32 do interact, UvsW is able to efficiently displace the gp32 protein from the ssDNA. This ability of UvsW to remove gp32 from ssDNA may explain its ability to enhance the strand invasion activity of the T4 recombinase (UvsX) and suggests a possible new role for UvsW in gp32-mediated DNA transactions.
authors.affiliation: 414 Wartik Laboratories, Department of Chemistry, Pennsylvania State University, University Park, PA 16802, USA.
authors.forename: Senthil K
authors.initials: SK
authors.lastname: Perumal
authors.forename: Scott W
authors.initials: SW
authors.lastname: Nelson
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Benkovic
mesh_terms: D017122:Bacteriophage T4 / Q000201:enzymology*; D004265:DNA Helicases / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D008954:Models, Biological; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D004277:DNA, Single-Stranded; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D014764:Viral Proteins; C060035:gp32 protein, Enterobacteria phage T4; D004265:DNA Helicases; C107414:UvsW protein, Bacteriophage T4
keywords: DNA repair helicase; DNA replication; DSB; EDTA; FRET; PNK; double-strand break; double-stranded DNA; dsDNA; ethylenediaminetetraacetic acid; fluorescence resonance energy transfer; polynucleotide kinase; recombination; repair; single-stranded DNA; single-stranded DNA binding protein; single-stranded M13; ssDNA; ssM13