Structural basis of sequence-specific cytosine deamination by double-stranded DNA deaminase toxin DddA

Yin, Lulu; Shi, Ke; Aihara, Hideki

doi:10.1038/s41594-023-01034-3

Structural basis of sequence-specific cytosine deamination by double-stranded DNA deaminase toxin DddA

Journal Article · Mon Jul 17 00:00:00 EDT 2023 · Nature Structural & Molecular Biology

DOI:https://doi.org/10.1038/s41594-023-01034-3· OSTI ID:2423640

Yin, Lulu ^[1]; Shi, Ke ^[1]; ^[1]

University of Minnesota, Minneapolis, MN (United States)

The interbacterial deaminase toxin DddA catalyzes cytosine-to-uracil conversion in double-stranded (ds) DNA and enables CRISPR-free mitochondrial base editing, but the molecular mechanisms underlying its unique substrate selectivity have remained elusive. Here, we report crystal structures of DddA bound to a dsDNA substrate containing the 5'-TC target motif. These structures show that DddA binds to the minor groove of a sharply bent dsDNA and engages the target cytosine extruded from the double helix. DddA Phe1375 intercalates in dsDNA and displaces the 5' (-1) thymine, which in turn replaces the target (0) cytosine and forms a noncanonical T–G base pair with the juxtaposed guanine. This tandem displacement mechanism allows DddA to locate a target cytosine without flipping it into the active site. Biochemical experiments demonstrate that DNA base mismatches enhance the DddA deaminase activity and relax its sequence selectivity. On the basis of the structural information, we further identified DddA mutants that exhibit attenuated activity or altered substrate preference. Our studies may help design new tools useful in genome editing or other applications.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES). Scientific User Facilities (SUF); US National Institutes of Health

Grant/Contract Number:: AC02-06CH11357

OSTI ID:: 2423640

Journal Information:: Nature Structural & Molecular Biology, Journal Name: Nature Structural & Molecular Biology Journal Issue: 8 Vol. 30; ISSN 1545-9993

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United States

Language:: English

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Structural basis of sequence-specific cytosine deamination by double-stranded DNA deaminase toxin DddA

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