Structural basis for aphidicolin inhibition of human processive DNA synthesis
- 1King Abdullah University of Science and Technology
Abstract
Eukaryotic DNA replication requires RNA--DNA primer synthesis by Pol and processive lagging- and leading-strand synthesis by Pol {delta} and Pol {epsilon} respectively, tethered to the PCNA clamp. Aphidicolin, a fungal diterpenoid, is a selective inhibitor of these B-family polymerases and a tool for inducing controlled replication stress, yet its structural mechanism has been defined only for the catalytic core of Pol . Here we report cryo-EM structures of human Pol {delta} and Pol {epsilon} holoenzymes bound to primer-template DNA, PCNA and aphidicolin at 3.0 [A] and 3.1 [A], respectively. In both, aphidicolin occupies the incoming dNTP pocket and flips the templating base outward, locking the fingers open and arresting synthesis while preserving the holoenzyme architecture. Aphidicolin occupies an architecturally conserved pocket in Pol , Pol {delta} and Pol {epsilon}, yet Pol {delta} is approximately 4-5-fold more sensitive than Pol {epsilon} in reconstituted PCNA-dependent assays. We attribute this to a grip beyond the templating base that anchors the template strand in Pol {delta} but engages it only loosely in Pol {epsilon}, allowing the strand to escape the aphidicolin trap. Selectivity therefore resides not in the inhibitor pocket but in template handling, defining a route to next-generation B-family polymerase inhibitors.
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BibTeX
@misc{danazumi2026structural,
title = {{Structural basis for aphidicolin inhibition of human processive DNA synthesis}},
author = {A. U. Danazumi and M. Tehseen and L. Zhao and S. Alshehri and V.-S. Raducanu and S. M. Hamdan and A. De Biasio},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.19.760038},
url = {https://www.biorxiv.org/content/10.64898/2026.10.19.760038v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Structural basis for aphidicolin inhibition of human processive DNA synthesis
AU - A. U. Danazumi
AU - M. Tehseen
AU - L. Zhao
AU - S. Alshehri
AU - V.-S. Raducanu
AU - S. M. Hamdan
AU - A. De Biasio
PY - 2026
DA - 2026/10/19
PB - bioRxiv
DO - 10.64898/2026.10.19.760038
UR - https://www.biorxiv.org/content/10.64898/2026.10.19.760038v1
AB - Eukaryotic DNA replication requires RNA--DNA primer synthesis by Pol and processive lagging- and leading-strand synthesis by Pol {delta} and Pol {epsilon} respectively, tethered to the PCNA clamp. Aphidicolin, a fungal diterpenoid, is a selective inhibitor of these B-family polymerases and a tool for inducing controlled replication stress, yet its structural mechanism has been defined only for the catalytic core of Pol . Here we report cryo-EM structures of human Pol {delta} and Pol {epsilon} holoenzymes bound to primer-template DNA, PCNA and aphidicolin at 3.0 [A] and 3.1 [A], respectively. In both, aphidicolin occupies the incoming dNTP pocket and flips the templating base outward, locking the fingers open and arresting synthesis while preserving the holoenzyme architecture. Aphidicolin occupies an architecturally conserved pocket in Pol , Pol {delta} and Pol {epsilon}, yet Pol {delta} is approximately 4-5-fold more sensitive than Pol {epsilon} in reconstituted PCNA-dependent assays. We attribute this to a grip beyond the templating base that anchors the template strand in Pol {delta} but engages it only loosely in Pol {epsilon}, allowing the strand to escape the aphidicolin trap. Selectivity therefore resides not in the inhibitor pocket but in template handling, defining a route to next-generation B-family polymerase inhibitors.
ER -