A Short Carboxyl-terminal Tail is Required for ssDNA-binding, Higher Order Structural Organization and Stability of the Mitochondrial Single Strand Annealing Protein Mgm101.

Department of Biochemistry and Molecular Biology, State University of New York Upstate Medical University, Syracuse, NY 13210, USA.
Molecular biology of the cell (Impact Factor: 4.47). 03/2013; 24(10). DOI: 10.1091/mbc.E13-01-0006
Source: PubMed


Mgm101 is a Rad52-type Single Strand Annealing Protein (SSAP) required for mitochondrial DNA (mtDNA) repair and maintenance. Structurally, Mgm101 forms large oligomeric rings. Here, we determined the function(s) of a 32-amino acid carboxyl-terminal tail (Mgm101(238-269)) conserved in the Mgm101-family of proteins. Mutagenic analysis showed that Lys253, Trp257, Arg259 and Tyr268 are essential for mtDNA maintenance. Mutations in Lys251, Arg252, Lys260 and Tyr266 affect mtDNA stability at 37°C and under oxidative stress. The Y268A mutation severely affects ssDNA-binding without altering the ring structure. Mutations in the Lys251-Arg252-Lys253 positive triad also affect ssDNA-binding. Moreover, we found that the C-tail alone is sufficient to mediate ssDNA-binding. Finally, we found that the W257A and R259A mutations dramatically affect the conformation and oligomeric state of Mgm101. These structural alterations correlated with protein degradation in vivo. The data thus indicate that the C-tail of Mgm101, likely displayed on the ring surface, is required for ssDNA-binding, higher order structural organization and protein stability. We speculate that an initial electrostatic and base stacking interaction with ssDNA could remodel ring organization. This may facilitate the formation of nucleoprotein filaments competent for mtDNA repair. These findings could have broad implications for understanding how SSAPs promote DNA repair and genome maintenance.

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