Structures and single-molecule analysis of bacterial motor nuclease AdnAB illuminate the mechanism of DNA double-strand break resection Journal Article
| Authors: | Jia, N.; Unciuleac, M. C.; Xue, C.; Greene, E. C.; Patel, D. J.; Shuman, S. |
| Article Title: | Structures and single-molecule analysis of bacterial motor nuclease AdnAB illuminate the mechanism of DNA double-strand break resection |
| Abstract: | Mycobacterial AdnAB is a heterodimeric helicase–nuclease that initiates homologous recombination by resecting DNA double-strand breaks (DSBs). The AdnA and AdnB subunits are each composed of an N-terminal motor domain and a C-terminal nuclease domain. Here we report cryoelectron microscopy (cryo-EM) structures of AdnAB in three functional states: in the absence of DNA and in complex with forked duplex DNAs before and after cleavage of the 5′ single-strand DNA (ssDNA) tail by the AdnA nuclease. The structures reveal the path of the 5′ ssDNA through the AdnA nuclease domain and the mechanism of 5′ strand cleavage; the path of the 3′ tracking strand through the AdnB motor and the DNA contacts that couple ATP hydrolysis to mechanical work; the position of the AdnA iron–sulfur cluster subdomain at the Y junction and its likely role in maintaining the split trajectories of the unwound 5′ and 3′ strands. Single-molecule DNA curtain analysis of DSB resection reveals that AdnAB is highly processive but prone to spontaneous pausing at random sites on duplex DNA. A striking property of AdnAB is that the velocity of DSB resection slows after the enzyme experiences a spontaneous pause. Our results highlight shared as well as distinctive properties of AdnAB vis-à-vis the RecBCD and AddAB clades of bacterial DSB-resecting motor nucleases. © 2019 National Academy of Sciences. All rights reserved. |
| Keywords: | controlled study; nonhuman; homologous recombination; in vitro study; dna strand breakage; double stranded dna; 5' untranslated region; gene interaction; nuclease; 3' untranslated region; adenosine triphosphate; single stranded dna; structure analysis; enzyme structure; hydrolysis; dna determination; cryoelectron microscopy; dna conformation; priority journal; article; dna end resection; dna curtain |
| Journal Title: | Proceedings of the National Academy of Sciences of the United States of America |
| Volume: | 116 |
| Issue: | 49 |
| ISSN: | 0027-8424 |
| Publisher: | National Academy of Sciences |
| Date Published: | 2019-12-03 |
| Start Page: | 24507 |
| End Page: | 24516 |
| Language: | English |
| DOI: | 10.1073/pnas.1913546116 |
| PUBMED: | 31740608 |
| PROVIDER: | scopus |
| PMCID: | PMC6900545 |
| DOI/URL: | |
| Notes: | Article -- Source: Scopus |
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