Biochemical properties of single-stranded DNA-binding protein from Mycobacterium smegmatis, a fast-growing mycobacterium and its physical and functional interaction with uracil DNA glycosylases.
The single-stranded DNA-binding proteins (SSBs) are vital to virtually all DNA functions. Here, we report on the biochemical properties of SSB from a fast-growing mycobacteria, Mycobacterium smegmatis, and the interaction of the homotetrameric SSBs with uracil DNA glycosylases (UDGs) from M. smegmatis (Msm), Mycobacterium tuberculosis (Mtu) and Escherichia coli (Eco). UDG is a crucial DNA repair enzyme, which removes the promutagenic uracil residues. MsmSSB stimulates activity of the homologous Msm UDG and of the heterologous Mtu-, and Eco-UDGs. On the contrary, while the MtuSSB stimulates the Mtu UDG, it inhibits the other two UDGs. Although the MsmSSB shares 84% identity with MtuSSB, the two are strikingly different, in that MsmSSB contains a glycine-rich segment (11 out of 13 residues) in the spacer connecting the N-terminal DNA-binding domain with the C-terminal acidic tail. While the DNA-binding properties of MsmSSB, such as its affinity to oligomeric DNA, requirement of minimum size DNA and the modes of interaction are indistinguishable from those of Eco-, and Mtu-SSBs, it is unclear if the glycine-rich segment confers structural advantage to MsmSSB, responsible for its stimulatory effect on all UDGs tested. More importantly, by using a small polypeptide inhibitor of UDGs, and the deletion mutants of SSBs, we suggest that the C-terminal acidic tail of the SSBs interacts within the DNA-binding groove of the UDGs, and propose a role for SSBs in the recruitment of UDGs to the damaged DNA.
Acharya
Narottam
N
Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore.
Varshney
Umesh
U
D000595:Amino Acid Sequence; D001665:Binding Sites; D045647:DNA Glycosylases; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D020102:Mycobacterium smegmatis / Q000378:metabolism*; D009699:N-Glycosyl Hydrolases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D051981:Uracil-DNA Glycosidase
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase