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April 24th, 2025
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University of California, Riverside
biochemistry
biorxiv

Structural insight into the substrate binding of the AMT complex via an inhibitor-trapped state

Shao, Z.Open in Google Scholar•Yoon, S.Open in Google Scholar•Lu, J.Open in Google Scholar•Athavale, P.Open in Google Scholar•Liu, Y.Open in Google Scholar•Song, J.Open in Google Scholar

N6-adenine (6mA) DNA methylation plays an important role in gene regulation and genome stability. The 6mA methylation in Tetrahymena thermophila is mainly mediated by the AMT complex, comprised of the AMT1, AMT7, AMTP1 and AMTP2 subunits. To date, how this complex assembles on the DNA substrate remains elusive. Here we report the structure of the AMT complex bound to the OCR protein from bacteriophage T7, mimicking the AMT-DNA encounter complex. The AMT1-AMT7 heterodimer approaches OCR from one side, while AMTP1 N-terminal domain, assuming a homeodomain fold, binds to OCR from the other side, resulting in a saddle-shaped architecture reminiscent of what was observed for prokaryotic 6mA writers. Mutation of the AMT1, AMT7 and AMTP1 residues on the OCR-contact points led to impaired DNA methylation activity to various extents, supporting a role for these residues in DNA binding. Furthermore, structural comparison of the AMT1-AMT7 subunits with the evolutionarily related METTL3-METTL14 and AMT1-AMT6 complexes reveal sequence conservation and divergence on the region corresponding to the OCR-binding site, shedding light onto the substrate binding of the latter two complexes. Together, this study suggests a substrate binding-induced open-to-closed conformational transition of the AMT complex, with implications in its substrate binding and processive 6mA methylation.

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