DNA-binding Domain within the Brh2 N Terminus Is the Primary Interaction Site for Association with DNA.
The C-terminal region of Brh2 (Brh2(CT)), the BRCA2 homolog in Ustilago maydis, is highly conserved and aligns with the DSS1/DNA-binding domain (DBD) of mammalian BRCA2, while the N-terminal region (Brh2(NT)) is poorly conserved and has no obvious functional domain except for the single Rad51-interacting BRC element. Paradoxically, Brh2(NT), but not Brh2(CT), complements the DNA repair and recombination deficiency of the brh2 mutant. We show here that Brh2(NT) exhibits an unexpected DNA binding activity with properties similar to that of the full-length protein. Deletion mapping localized the region responsible for the DNA binding activity to a stretch of residues between the BRC element and the canonical DBD. A heterologous DNA-binding domain from the large subunit of replication protein A substituted for the endogenous binding region within Brh2(NT) in supporting DNA repair. Rad51-promoted strand invasion was stimulated by Brh2(NT), but required the presence of the BRC element. The findings suggest a model in which Brh2(NT) serves as the principal site for association with DNA, while the Brh2(CT) provides a means for regulation.
Zhou
Qingwen
Q
Department of Microbiology and Immunology, Cornell University Weill Medical College, New York, New York 10021.
Kojic
Milorad
M
Holloman
William K
WK
D024682:BRCA2 Protein / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair / Q000502:physiology*; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D010449:Peptide Mapping; D017434:Protein Structure, Tertiary / Q000502:physiology; D051135:Rad51 Recombinase / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014588:Ustilago / Q000235:genetics / Q000378:metabolism*
D024682:BRCA2 Protein; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D051135:Rad51 Recombinase