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Binding to PCNA in Euryarchaeal DNA Replication requires two PIP motifs for DNA polymerase D and one PIP motif for DNA polymerase B

Abstract : Replicative DNA polymerases possess a canonical C-terminal proliferating cell nuclear antigen (PCNA)-binding motif termed the PCNA-interacting protein (PIP) box. We investigated the role of the PIP box on the functional interactions of the two DNA polymerases, PabPol B (family B) and PabPol D (family D), from the hyperthermophilic euryarchaeon Pyrococcus abyssi, with its cognate PCNA. The PIP box was essential for interactions of PabPol B with PCNA, as shown by surface plasmon resonance and primer extension studies. In contrast, binding of PabPol D to PCNA was affected only partially by removing the PIP motif. We identified a second palindromic PIP box motif at the N-terminus of the large subunit of PabPol D that was required for the interactions of PabPol D with PCNA. Thus, two PIP motifs were needed for PabPol D for binding to PabPCNA. Moreover, the C-terminus of PabPCNA was essential for stimulation of PabPol D activity but not for stimulation of PabPol B activity. Neither DNA polymerase interacted with the PabPCNA interdomain connecting loop. Our data suggest that distinct processes are involved in PabPol D and PabPol B binding to PCNA, raising the possibility that Archaea require two mechanisms for recruiting replicative DNA polymerases at the replication fork.
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https://hal.univ-brest.fr/hal-00609928
Contributor : Violaine Garguilo Connect in order to contact the contributor
Submitted on : Wednesday, July 20, 2011 - 3:12:39 PM
Last modification on : Monday, October 11, 2021 - 2:22:06 PM

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Benoît Castrec, Christophe Rouillon, Ghislaine Henneke, Didier Flament, Joël Querellou, et al.. Binding to PCNA in Euryarchaeal DNA Replication requires two PIP motifs for DNA polymerase D and one PIP motif for DNA polymerase B. Journal of Molecular Biology, Elsevier, 2009, 394 (2), pp.209-218. ⟨10.1016/j.jmb.2009.09.044⟩. ⟨hal-00609928⟩

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