Uracil-DNA glycosylase efficiency is modulated by substrate rigidity

Uracil DNA-glycosylase (UNG) is a DNA repair enzyme that removes the highly mutagenic uracil lesion from DNA using a base flipping mechanism. Although this enzyme has evolved to remove uracil from diverse sequence contexts, UNG excision efficiency depends on DNA sequence. To provide the molecular ba...

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Bibliographic Details
Published in:Scientific reports 2023-03, Vol.13 (1), p.3915-3915, Article 3915
Main Authors: Orndorff, Paul B., Poddar, Souvik, Owens, Aerial M., Kumari, Nikita, Ugaz, Bryan T., Amin, Samrat, Van Horn, Wade D., van der Vaart, Arjan, Levitus, Marcia
Format: Article
Language:English
Subjects:
631/45/147
631/45/56
631/92/607/1159
Deformability
Deoxyribonucleic acid
DNA
DNA - chemistry
DNA glycosylase
DNA Repair
Efficiency
Enzymes
Flexibility
Fluorescence spectroscopy
Humanities and Social Sciences
Humans
Lesions
Molecular evolution
multidisciplinary
Mutagenesis
Mutation hot spots
NMR
Nuclear magnetic resonance
Nucleotide sequence
Rigidity
Science
Science (multidisciplinary)
Substrate preferences
Substrates
Uracil
Uracil-DNA glycosidase
Uracil-DNA Glycosidase - chemistry
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Summary:Uracil DNA-glycosylase (UNG) is a DNA repair enzyme that removes the highly mutagenic uracil lesion from DNA using a base flipping mechanism. Although this enzyme has evolved to remove uracil from diverse sequence contexts, UNG excision efficiency depends on DNA sequence. To provide the molecular basis for rationalizing UNG substrate preferences, we used time-resolved fluorescence spectroscopy, NMR imino proton exchange measurements, and molecular dynamics simulations to measure UNG specificity constants ( k cat / K M ) and DNA flexibilities for DNA substrates containing central AUT, TUA, AUA, and TUT motifs. Our study shows that UNG efficiency is dictated by the intrinsic deformability around the lesion, establishes a direct relationship between substrate flexibility modes and UNG efficiency, and shows that bases immediately adjacent to the uracil are allosterically coupled and have the greatest impact on substrate flexibility and UNG activity. The finding that substrate flexibility controls UNG efficiency is likely significant for other repair enzymes and has major implications for the understanding of mutation hotspot genesis, molecular evolution, and base editing.
ISSN:2045-2322
2045-2322
DOI:10.1038/s41598-023-30620-0