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Substrate-Specific Inactivation of Human AAG by Tumor-Associated Single Nucleotide Polymorphic Variants
Olga A Kladova1, Timofey E Tyugashev1, Artemiy S Bakman1
1Knorre Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of Russian Academy of Sciences, Novosibirsk 630090, Russia.
International Journal of Molecular Sciences
|August 13, 2026
Summary
Human alkyladenine DNA glycosylase (AAG) gene variants impact DNA repair. Three common SNPs showed reduced stability but altered enzyme activity, reprogramming substrate specificity and potentially serving as cancer biomarkers.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Human alkyladenine DNA glycosylase (AAG) repairs damaged purines in DNA.
- Single nucleotide polymorphisms (SNPs) in the AAG gene are common, but their functional effects are largely unknown.
- Three high-deleterious-potential AAG SNPs (P94L, V158M, E293K) were previously identified.
Purpose of the Study:
- To biochemically and structurally characterize the functional impact of three common AAG SNPs (P94L, V158M, E293K).
- To assess the effects of these SNPs on enzyme thermal stability, DNA binding, and catalytic activity against distinct substrates.
- To elucidate the structural consequences of these non-active-site mutations using molecular dynamics simulations.
Main Methods:
- Biochemical assays to measure thermal stability (melting temperature) and DNA binding affinity.
- Enzyme kinetics assays to determine catalytic activity against 1, N6-ethenoadenosine (εA) and hypoxanthine (Hx).
- Molecular dynamics (MD) simulations to investigate structural changes and DNA interactions.
Main Results:
- All three AAG mutants (P94L, V158M, E293K) exhibited reduced thermal stability.
- DNA binding affinities of the mutants were comparable to wild-type (WT) AAG.
- Mutants displayed differential loss of catalytic activity: P94L was inactive, V158M showed altered substrate specificity, and E293K was also substrate-specific.
- MD simulations revealed distinct structural alterations, including loop modification, active site cleft narrowing, and disruption of salt bridges.
Conclusions:
- Non-active-site SNPs in AAG can significantly alter enzyme function and substrate specificity.
- These AAG variants reprogrammed catalytic activity, demonstrating complex functional consequences.
- AAG SNPs may serve as potential biomarkers for cancer risk and response to alkylating chemotherapy.
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