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<p>DinG-like proteins are members of the XPD-family SF2 helicases and are widely distributed in bacteria, yet they exhibit remarkable diversity in domain architecture and biological function. A distinct subgroup of DinG homologs harboring an N-terminal PD-(D/E)XK-type nuclease domain, designated EndoDinGs, has remained uncharacterized. Here, we report the first structural and functional characterization of this subgroup using <i>Finegoldia magna</i> DinG (FmaDinG) as a representative example. FmaDinG displays Mn<sup>2+</sup>-preferred endonuclease activity together with Mg<sup>2+</sup>-dependent 5′–3′ unidirectional helicase activity. We determined the crystal structure of the FmaDinG–ssDNA–ADPNP complex at 2.26 Å resolution. Structural and biochemical analyses reveal a direct role of the Arch domain in DNA engagement and suggest a gating mechanism within the nuclease domain that regulates DNA entry, a feature that may be specific to the EndoDinG subgroup. Key residues required for nuclease and helicase activities were identified by site-directed mutagenesis. Together, these results support a mechanistic model in which EndoDinGs such as FmaDinG function as coordinated nuclease–helicase machines for DNA processing.</p>

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Structural and biochemical characterization of a novel DinG containing an endonuclease domain

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年7月15日

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  • Shen Li,
  • Tianwen Gao,
  • Wanshan Hao,
  • Hang Chen,
  • Xiaolei Hu,
  • Rong Qi,
  • Cheng Chen,
  • Kaiying Cheng

摘要

DinG-like proteins are members of the XPD-family SF2 helicases and are widely distributed in bacteria, yet they exhibit remarkable diversity in domain architecture and biological function. A distinct subgroup of DinG homologs harboring an N-terminal PD-(D/E)XK-type nuclease domain, designated EndoDinGs, has remained uncharacterized. Here, we report the first structural and functional characterization of this subgroup using Finegoldia magna DinG (FmaDinG) as a representative example. FmaDinG displays Mn2+-preferred endonuclease activity together with Mg2+-dependent 5′–3′ unidirectional helicase activity. We determined the crystal structure of the FmaDinG–ssDNA–ADPNP complex at 2.26 Å resolution. Structural and biochemical analyses reveal a direct role of the Arch domain in DNA engagement and suggest a gating mechanism within the nuclease domain that regulates DNA entry, a feature that may be specific to the EndoDinG subgroup. Key residues required for nuclease and helicase activities were identified by site-directed mutagenesis. Together, these results support a mechanistic model in which EndoDinGs such as FmaDinG function as coordinated nuclease–helicase machines for DNA processing.