{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T14:27:12Z","timestamp":1785421632592,"version":"3.56.0"},"posted":{"date-parts":[[2026,1,18]]},"group-title":"Biochemistry","reference-count":90,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,1,18]],"date-time":"2026-01-18T00:00:00Z","timestamp":1768694400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"award":["GM115809"],"award-info":[{"award-number":["GM115809"]}],"id":[{"id":"https:\/\/ror.org\/04q48ey07","id-type":"ROR","asserted-by":"publisher"}]},{"award":["GM131754"],"award-info":[{"award-number":["GM131754"]}],"id":[{"id":"https:\/\/ror.org\/04q48ey07","id-type":"ROR","asserted-by":"publisher"}]},{"id":[{"id":"https:\/\/ror.org\/006w34k90","id-type":"ROR","asserted-by":"publisher"}]},{"id":[{"id":"https:\/\/ror.org\/00wm07d60","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,1,20]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  DNA sliding clamps are essential for processive DNA synthesis in all domains of life and are loaded by ATP-dependent clamp loaders that recognize recessed 3\u2032 ends. How clamp loaders function at nicks and small ssDNA gaps\u2014common intermediates during DNA repair\u2014remains incompletely understood. Here, we show that the bacterial\n                  <jats:italic>Escherichia coli<\/jats:italic>\n                  DnaX clamp loader employs a fundamentally different mechanism from its eukaryotic counterpart. Whereas eukaryotic RFC unwinds DNA at the recessed 3\u2032 end and stabilizes the 5\u2032-dsDNA at a dedicated shoulder site, the bacterial DnaX-complex neither unwinds DNA nor stably binds the 5\u2032-dsDNA in vitro. Instead, cryo-EM structures reveal that the \u03b2-clamp itself contains a conserved external DNA-binding site that enables sharp bending of gapped DNA by \u223c150\u00b0, promoting insertion of the 3\u2032-dsDNA into the clamp. This DNA-bending mechanism allows efficient \u03b2-clamp loading at nicks and small gaps and reveals a distinct bacterial strategy for clamp loading. Because small DNA gaps are frequently associated with DNA damage, clamps loaded at these sites are likely important for DNA repair.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>In brief<\/jats:title>\n                  <jats:p>Zheng et al. show that the bacterial clamp loader DnaX-complex uses a DNA-bending mechanism\u2014rather than DNA unwinding\u2014to load the \u03b2-clamp at nicks and small gaps, revealing a clamp-loading strategy distinct from eukaryotic RFC and relevant to DNA damage repair.<\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Highlights<\/jats:title>\n                  <jats:list list-type=\"bullet\">\n                    <jats:list-item>\n                      <jats:p>The bacterial DnaX clamp loader lacks a stable shoulder DNA-binding site<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Unlike eukaryotic RFC, DnaX does not unwind DNA at nicks and small gaps<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>The E. coli \u03b2-clamp contains a conserved external DNA-binding site absent in PCNA<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Sharp DNA bending enables \u03b2-clamp loading at nicks and small ssDNA gaps<\/jats:p>\n                    <\/jats:list-item>\n                  <\/jats:list>\n                <\/jats:sec>","DOI":"10.64898\/2026.01.17.700081","type":"posted-content","created":{"date-parts":[[2026,1,19]],"date-time":"2026-01-19T03:40:14Z","timestamp":1768794014000},"source":"Crossref","is-referenced-by-count":0,"title":["The\n                  <i>E. coli<\/i>\n                  DnaX clamp loader sharply bends DNA to load \u03b2-clamp at nicks and small gaps"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7139-4831","authenticated-orcid":false,"given":"Fengwei","family":"Zheng","sequence":"first","affiliation":[{"name":"Department of Structural Biology, Van Andel Institute, Grand Rapids, Michigan, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4728-6531","authenticated-orcid":false,"given":"Nina Y.","family":"Yao","sequence":"additional","affiliation":[{"name":"DNA Replication Laboratory and Howard Hughes Medical Institute, The Rockefeller University, New York, New York, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1882-2358","authenticated-orcid":false,"given":"Roxana E.","family":"Georgescu","sequence":"additional","affiliation":[{"name":"DNA Replication Laboratory and Howard Hughes Medical Institute, The Rockefeller University, New York, New York, 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