Identification and properties of the Deinococcus grandis and Deinococcus proteolyticus single-stranded DNA binding proteins (SSB).

To study the biochemical properties of SSB's from Deinococcus grandis (DgrSSB) and Deinoc- occus proteolyticus (DprSSB), we have cloned the ssb genes obtained by PCR and have devel- oped Escherichia coli overexpression systems. The genes consist of an open reading frame of 891 (DgrSSB) and 876 (DprSSB) nucleotides encoding proteins of 296 and 291 amino acids with a cal - culated molecular mass of 32.29 and 31.33 kDa, respectively. The amino-acid sequence of DgrSSB exhibits 45%, 44% and 82% identity and the amino-acid sequence of DprSSB reveals 43%, 43% and 69% identity with Thermus aquaticus (TaqSSB), Thermus thermophilus (TthSSB) and Deinoc- occus radiodurans SSBs, respectively. We show that DgrSSB and DprSSB are similar to Thermus/ Deinococcus SSBs in their biochemical properties. They are functional as homodimers, with each monomer encoding two single-stranded DNA binding domains (OB-folds). In fluorescence titra - tions with poly(dT), both proteins bind single-stranded DNA with a binding site size of about 33 nt per homodimer. In a complementation assay in E. coli, DgrSSB and DprSSB took over the in vivo function of EcoSSB. Thermostability with half-lives of about 1 min at 65-67.5°C make DgrSSB and DprSSB similar to the known SSB of Deinococcus radiodurans (DraSSB).