Abstract
The N6-methyladenine (6mA) modification is an essential epigenetic marker and plays a crucial role in processes, such as DNA repair, replication, gene expression regulation, etc. YerA from Bacillus subtilis is considered a novel class of enzymes capable of catalyzing the deamination of 6mA to produce hypoxanthine. Despite the significance of this type of enzymes in bacterial self-defense systems and potential applications as a gene-editing tool, the substrate specificity, the catalytic mechanism and the physiological function of YerA are currently unclear due to the lack of structural information. In this study, we expressed the recombinant enzyme and conducted its reconstitution to yield the active form. Our deamination assays showed that N6-methyladenosine (N6-mAdo) served as a more favorable substrate than its base derivative 6mA. Here we report the high-resolution structure of the C-terminal region of YerA, which exhibited a compact architecture composed of two antiparallel b-sheets with no obvious close structural homologs in PDB. We also created docking models to investigate the ligand-binding pattern, and found that more favorable contacts of N6-mAdo with the enzyme binding pocket lead to its preference for 6mA. Lastly, structural comparison to the deaminase MAPDA allowed us to propose a plausible role for this C-terminal region: shielding the active site from solvent and protecting the intermediate during catalysis. Taken together, this study sheds light on the catalytic mechanism and evolutionary pathways of the promiscuous enzyme YerA, thereby contributing to our molecular understanding of epigenetic nucleoside metabolism.
Published Version
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