title: Eukaryotic DNA polymerases require an iron-sulfur cluster for the formation of active complexes.
abstract: The eukaryotic replicative DNA polymerases (Pol α, δ and ɛ) and the major DNA mutagenesis enzyme Pol ζ contain two conserved cysteine-rich metal-binding motifs (CysA and CysB) in the C-terminal domain (CTD) of their catalytic subunits. Here we demonstrate by in vivo and in vitro approaches the presence of an essential [4Fe-4S] cluster in the CysB motif of all four yeast B-family DNA polymerases. Loss of the [4Fe-4S] cofactor by cysteine ligand mutagenesis in Pol3 destabilized the CTD and abrogated interaction with the Pol31 and Pol32 subunits. Reciprocally, overexpression of accessory subunits increased the amount of the CTD-bound Fe-S cluster. This implies an important physiological role of the Fe-S cluster in polymerase complex stabilization. Further, we demonstrate that the Zn-binding CysA motif is required for PCNA-mediated Pol δ processivity. Together, our findings show that the function of eukaryotic replicative DNA polymerases crucially depends on different metallocenters for accessory subunit recruitment and replisome stability.
authors: Institut für Zytobiologie und Zytopathologie, Philipps-Universität Marburg, Marburg, Germany.
authors: Daili J A
authors: DJ
authors: Netz
authors: Carrie M
authors: CM
authors: Stith
authors: Martin
authors: M
authors: Stümpfig
authors: Gabriele
authors: G
authors: Köpf
authors: Daniel
authors: D
authors: Vogel
authors: Heide M
authors: HM
authors: Genau
authors: Joseph L
authors: JL
authors: Stodola
authors: Roland
authors: R
authors: Lill
authors: Peter M J
authors: PM
authors: Burgers
authors: Antonio J
authors: AJ
authors: Pierik
mesh_terms: D020134:Catalytic Domain; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000378:metabolism*; D007501:Iron / Q000378:metabolism; D008958:Models, Molecular; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013455:Sulfur / Q000378:metabolism
chemical_list: D013455:Sulfur; D007501:Iron; D004259:DNA-Directed DNA Polymerase