Cryo-EM structure of Pol κ-DNA-PCNA holoenzyme and implications for polymerase switching in DNA lesion bypass

Claudia Lancey; Muhammad Tehseen; Masateru Takahashi; Mohamed A Sobhy; Timothy J Ragan; Ramon Crehuet; Samir M Hamdan; Alfredo De Biasio

doi:10.1101/2020.07.10.196956

bioRxiv · 10.1101/2020.07.10.196956

Cryo-EM structure of Pol κ-DNA-PCNA holoenzyme and implications for polymerase switching in DNA lesion bypass

Claudia Lancey·Muhammad Tehseen·Masateru Takahashi·Mohamed A Sobhy·Timothy J Ragan·Ramon Crehuet·Samir M Hamdan·Alfredo De Biasio

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Abstract

Replacement of the stalled replicative polymerase (Pol {delta}) at a DNA lesion by the error-prone DNA polymerase {kappa} (Pol {kappa}) restarts synthesis past the lesion to prevent genome instability. The switching from Pol {delta} to Pol {kappa} is mediated by the processivity clamp PCNA but the structural basis of this mechanism is unknown. We determined the Cryo-EM structures of human Pol {kappa}-DNA-PCNA complex and of a stalled Pol {delta}-DNA-PCNA complex at 3.9 and 4.7 [A] resolution, respectively. In Pol {kappa} complex, the C-terminus of the PAD domain docks the catalytic core to one PCNA protomer in an angled orientation, bending the DNA exiting Pol {kappa} active site through PCNA. In Pol {delta} complex, the DNA is disengaged from the active site but is retained by the thumb domain. We present a model for polymerase switching facilitated by Pol {kappa} recruitment to PCNA and Pol {kappa} conformational sampling to seize the DNA from stalled Pol {delta} assisted by PCNA tilting.

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Claudia Lancey, Muhammad Tehseen, Masateru Takahashi, Mohamed A Sobhy, Timothy J Ragan, Ramon Crehuet, Samir M Hamdan, Alfredo De Biasio. 2020-07-10. Cryo-EM structure of Pol κ-DNA-PCNA holoenzyme and implications for polymerase switching in DNA lesion bypass. https://doi.org/10.1101/2020.07.10.196956

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