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Cited by 3CRISPR and Genetic EngineeringRNA regulation and diseaseDNA and Nucleic Acid Chemistry

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Structural basis for sequence context-independent single-stranded DNA cytosine deamination by the bacterial toxin SsdA

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Yin, Lulu, Chen, Yanjun, Shi, Ke, Barreto Duran, Emilia, Harris, Reuben S., Aihara, Hideki·2025·Nature Communications
DOI 10.1038/s41467-025-63943-9PMC PMC12494967

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Abstract

Abstract DNA deaminase toxins are involved in interbacterial antagonism and the generation of genetic diversity in surviving bacterial populations. These enzymes have also been adopted as genome engineering tools. The single-stranded (ss)DNA deaminase SsdA represents the bacterial deaminase toxin family-2 (BaDTF2) and it deaminates ssDNA cytosines with little sequence context dependence, which contrasts with the AID/APOBEC family of sequence-selective ssDNA cytosine deaminases. Here we report the crystal structure of SsdA in complex with a ssDNA substrate. The structure reveals a unique mode of substrate binding, in which a cluster of aromatic residues of SsdA engages ssDNA in a V-shaped conformation sharply bent across the target cytosine. The bases 5’ or 3’ to the target cytosine are stacked linearly and make few sequence-specific protein contacts, thus explaining the broad substrate selectivity of SsdA. Unexpectedly, SsdA contains a β-amino acid isoaspartate, which is important for enzymatic activity and may contribute to the stability of SsdA as a toxin. Structure- function studies helped to design SsdA mutants active in human cells, which could lead to future applications in genome engineering.

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Metadata

Authors
Yin, L., Chen, Y., Shi, K., Barreto Duran, E., Harris, R. S., & Aihara, H.
Affiliations
  • University of Minnesota (US)
  • The University of Texas at San Antonio Health Science Center (US)
  • Howard Hughes Medical Institute (US)
Corresponding author
Aihara, Hideki
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2025-10-03
Volume
16
Issue
1
Pages
8841-8841
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-025-63943-9
PMC
PMC12494967
OpenAlex
W4414793805
Cited by
3
References
1
Citations by year
2026: 2 · 2025: 1
FWCI
0.90
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by-nc-nd)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • PubMed Central (publishedVersion, cc-by-nc-nd)
  • PubMed Central (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • U.S. Department of Health & Human Services | National Institutes of Health (R35-GM118047, P01-CA234228)
  • Cancer Prevention and Research Institute of Texas (RR220053)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
Keywords
crystal structure, substrate binding, π-π stacking, genome editing, base editing, APOBEC, AID

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