{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T12:53:35Z","timestamp":1791550415843,"version":"4.3.4"},"reference-count":52,"publisher":"American Society for Microbiology","issue":"8","license":[{"start":{"date-parts":[[2025,8,13]],"date-time":"2025-08-13T00:00:00Z","timestamp":1755043200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2025,8,13]],"date-time":"2025-08-13T00:00:00Z","timestamp":1755043200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32270043, 32100017"],"award-info":[{"award-number":["32270043, 32100017"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32200146"],"award-info":[{"award-number":["32200146"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004731","name":"Natural Science Foundation of Zhejiang Province","doi-asserted-by":"publisher","award":["LQ22C050002"],"award-info":[{"award-number":["LQ22C050002"]}],"id":[{"id":"10.13039\/501100004731","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004731","name":"Natural Science Foundation of Zhejiang Province","doi-asserted-by":"publisher","award":["LKLY25H200002"],"award-info":[{"award-number":["LKLY25H200002"]}],"id":[{"id":"10.13039\/501100004731","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["mBio"],"published-print":{"date-parts":[[2025,8,13]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title\/>\n                    <jats:p>\n                      Damage-inducible gene G (DinG), a bacterial homolog of SF2 helicase, has been extensively studied in\n                      <jats:italic toggle=\"yes\">Escherichia coli<\/jats:italic>\n                      . However, the structural and functional characteristics of DinG homologs fused with an N-terminal 3\u2032\u20135\u2032 exonuclease domain, such as\n                      <jats:italic toggle=\"yes\">Staphylococcus aureus<\/jats:italic>\n                      DinG (SaDinG), remain unexplored. In this study, we demonstrate that SaDinG possesses 3\u2032\u20135\u2032 exonuclease activity and exhibits 5\u2032\u20133\u2032 helicase activity on diverse DNA substrates, including splayed duplexes, 5\u2032-overhangs, double flaps, bubbles, and gapped duplexes, resolving prior ambiguities about its biochemical functions. Intriguingly, both enzymatic activities were inhibited by elevated ATP concentrations, suggesting a potential ATP-dependent regulatory mechanism\n                      <jats:italic toggle=\"yes\">in vivo<\/jats:italic>\n                      . We determined the crystal structures of SaDinG bound to ssDNA at ~3.2 \u00c5 resolution and identified key residues essential for its helicase and exonuclease activities through mutational analysis. Phenotypic studies revealed that a SaDinG deletion mutant exhibited heightened sensitivity to DNA crosslinking agents (mitomycin C and formaldehyde) but retained wild-type susceptibility to other DNA-damaging compounds. Complementation with either nuclease-dead or helicase-dead variants failed to restore crosslink resistance, indicating that both activities are indispensable for DNA crosslink repair. These results support a model in which SaDinG functions as a coordinated nuclease-helicase machine specifically adapted for DNA crosslink repair, with its dual enzymatic activities being tightly regulated by physiological ATP concentrations.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>\n                      DNA helicases and exonucleases play essential roles in genome maintenance; however, little is known about bacterial helicase-exonuclease fusion proteins. This study examines DNA helicases and exonucleases that play essential roles in genome maintenance; however, little is known about bacterial helicase-exonuclease fusion proteins. This study provides the first structural and functional characterization of\n                      <jats:italic toggle=\"yes\">Staphylococcus aureus<\/jats:italic>\n                      DinG (SaDinG), a unique enzyme that combines 5\u2032-3\u2032 helicase and 3\u2032-5\u2032 exonuclease activities. Our findings resolve previous uncertainties about SaDinG's function and reveal an ATP-dependent regulatory mechanism that modulates its activity. Additionally, we demonstrate that SaDinG is critical for bacterial resistance to DNA crosslinking agents. These insights not only expand our understanding of bacterial DNA repair but also suggest potential avenues for targeting DinG-like enzymes in antimicrobial strategies. Given the growing concerns over antibiotic resistance, understanding how bacteria maintain genome integrity under stress conditions is crucial. This work lays the foundation for further exploration of bacterial helicase-exonuclease systems and their role in genome stability and adaptive survival.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/mbio.00884-25","type":"journal-article","created":{"date-parts":[[2025,6,30]],"date-time":"2025-06-30T09:00:42Z","timestamp":1751274042000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":5,"title":["Structural and functional investigation of DinG containing a 3\u2032\u20135\u2032 exonuclease domain"],"prefix":"10.1128","volume":"16","author":[{"given":"Tianwen","family":"Gao","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/014v1mr15","id-type":"ROR","asserted-by":"publisher"}],"name":"Zhejiang Key Laboratory of Medical Epigenetics, Department of Immunology and Pathogen Biology, School of Basic Medical Sciences, Affiliated Hospital of Hangzhou Normal University, 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