کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1981597 | 1539419 | 2015 | 10 صفحه PDF | دانلود رایگان |
• Deinococcal Ssb variants with only N or C-terminal OB fold were constructed.
• The ssDNA binding affinity decreased in the order SsbFL > SsbC > SsbNC > SsbN.
• SsbNC or SsbN formed a binary complex with SsbC to enhance ssDNA binding.
• Individual Ssb variants modulated DNA relaxation by topoisomerase I.
• Only SsbFL facilitated strand exchange with cognate RecA.
Single-stranded DNA binding protein (Ssb) of Deinococcus radiodurans comprises N- and C-terminal oligonucleotide/oligosaccharide binding (OB) folds connected by a beta hairpin connector. To assign functional roles to the individual OB folds, we generated three Ssb variants: SsbN (N-terminal without connector), SsbNC (N-terminal with connector) and SsbC (C-terminal), each harboring one OB fold. Both SsbN and SsbNC displayed weak single-stranded DNA (ssDNA) binding activity, compared to the full-length Ssb (SsbFL). The level of ssDNA binding activity displayed by SsbC was intermediate between SsbFL and SsbN. SsbC and SsbFL predominantly existed as homo-dimers while SsbNC/SsbN formed different oligomeric forms. In vitro, SsbNC or SsbN formed a binary complex with SsbC that displayed enhanced ssDNA binding activity. Unlike SsbFL, Ssb variants were able to differentially modulate topoisomerase-I activity, but failed to stimulate Deinococcal RecA-promoted DNA strand exchange. The results suggest that the C-terminal OB fold is primarily responsible for ssDNA binding. The N-terminal OB fold binds weakly to ssDNA but is involved in multimerization.
Journal: FEBS Open Bio - Volume 5, 2015, Pages 378–387