Mechanistic insight into the assembly of the HerA-NurA helicase-nuclease DNA end resection complex

• Published in: Nucleic acids research 2017-11, Vol.45 (20), p.12025-12038
• Main Authors: Ahdash, Zainab, Lau, Andy M, Byrne, Robert Thomas, Lammens, Katja, Stüetzer, Alexandra, Urlaub, Henning, Booth, Paula J, Reading, Eamonn, Hopfner, Karl-Peter, Politis, Argyris
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - chemistry; Adenosine Triphosphate - metabolism; Archaeal Proteins - chemistry; Archaeal Proteins - genetics; Archaeal Proteins - metabolism; Binding Sites - genetics; Deoxyribonucleases - chemistry; Deoxyribonucleases - genetics; Deoxyribonucleases - metabolism; DNA Breaks, Double-Stranded; DNA Helicases - chemistry; DNA Helicases - genetics; DNA Helicases - metabolism; DNA Repair; DNA, Archaeal - chemistry; DNA, Archaeal - genetics; DNA, Archaeal - metabolism; Models, Molecular; Nucleic Acid Conformation; Protein Binding; Protein Domains; Protein Multimerization; Structural Biology; Sulfolobus solfataricus - enzymology; Sulfolobus solfataricus - genetics; Sulfolobus solfataricus - metabolism
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/gkx890

The HerA-NurA helicase-nuclease complex cooperates with Mre11 and Rad50 to coordinate the repair of double-stranded DNA breaks. Little is known, however, about the assembly mechanism and activation of the HerA-NurA. By combining hybrid mass spectrometry with cryo-EM, computational and biochemical data, we investigate the oligomeric formation of HerA and detail the mechanism of nucleotide binding to the HerA-NurA complex from thermophilic archaea. We reveal that ATP-free HerA and HerA-DNA complexes predominantly exist in solution as a heptamer and act as a DNA loading intermediate. The binding of either NurA or ATP stabilizes the hexameric HerA, indicating that HerA-NurA is activated by substrates and complex assembly. To examine the role of ATP in DNA translocation and processing, we investigated how nucleotides interact with the HerA-NurA. We show that while the hexameric HerA binds six nucleotides in an 'all-or-none' fashion, HerA-NurA harbors a highly coordinated pairwise binding mechanism and enables the translocation and processing of double-stranded DNA. Using molecular dynamics simulations, we reveal novel inter-residue interactions between the external ATP and the internal DNA binding sites. Overall, here we propose a stepwise assembly mechanism detailing the synergistic activation of HerA-NurA by ATP, which allows efficient processing of double-stranded DNA.