Single-stranded DNA-binding protein of Deinococcus radiodurans: a biophysical characterization.
The highly conserved bacterial single-stranded DNA-binding (SSB) proteins play an important role in DNA replication, repair and recombination and are essential for the survival of the cell. They are functional as tetramers, in which four OB(oligonucleotide/oligosaccharide binding)-folds act as DNA-binding domains. The protomer of the SSB protein from the extremely radiation-resistant organism Deinococcus radiodurans (DraSSB) has twice the size of the other bacterial SSB proteins and contains two OB-folds. Using analytical ultracentrifugation, we could show that DraSSB forms globular dimers with some protrusions. These DraSSB dimers can interact with two molecules of E.coli DNA polymerase III chi subunit. In fluorescence titrations with poly(dT) DraSSB bound 47-54 nt depending on the salt concentration, and fluorescence was quenched by more than 75%. A distinct low salt binding mode as for EcoSSB was not observed for DraSSB. Nucleic acid binding affinity, rate constant and association mechanism are quite similar for EcoSSB and DraSSB. In a complementation assay in E.coli, DraSSB took over the in vivo function of EcoSSB. With DraSSB behaving almost identical to EcoSSB the question remains open as to why dimeric SSB proteins have evolved in the Thermus group of bacteria.
Witte
Gregor
G
Department of Biophysical Chemistry, Medical School Hannover Hannover, Germany.
Urbanke
Claus
C
Curth
Ute
U
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004258:DNA Polymerase III / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D034301:Deinococcus / Q000737:chemistry / Q000235:genetics*; D019281:Dimerization; D029968:Escherichia coli Proteins / Q000235:genetics; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C476430:SSB protein, E coli; D004258:DNA Polymerase III