title: Direct interaction between uracil-DNA glycosylase and a proliferating cell nuclear antigen homolog in the crenarchaeon Pyrobaculum aerophilum.
abstract: Proliferating cell nuclear antigen (PCNA) acts as a sliding clamp on duplex DNA. Its homologs, present in Eukarya and Archaea, are part of protein complexes that are indispensable for DNA replication and DNA repair. In Eukarya, PCNA is known to interact with more than a dozen different proteins, including a human major nuclear uracil-DNA glycosylase (hUNG2) involved in immediate postreplicative repair. In Archaea, only three classes of PCNA-binding proteins have been reported previously: replication factor C (the PCNA clamp loader), family B DNA polymerase, and flap endonuclease. In this study, we report a direct interaction between a uracil-DNA glycosylase (Pa-UDGa) and a PCNA homolog (Pa-PCNA1), both from the hyperthermophilic crenarchaeon Pyrobaculum aerophilum (T(opt) = 100 degrees C). We demonstrate that the Pa-UDGa-Pa-PCNA1 complex is thermostable, and two hydrophobic amino acid residues on Pa-UDGa (Phe(191) and Leu(192)) are shown to be crucial for this interaction. It is interesting to note that although Pa-UDGa has homologs throughout the Archaea and bacteria, it does not share significant sequence similarity with hUNG2. Nevertheless, our results raise the possibility that Pa-UDGa may be a functional analog of hUNG2 for PCNA-dependent postreplicative removal of misincorporated uracil.
authors.affiliation: Department of Microbiology and Molecular Genetics and the Molecular Biology Institute, University of California, Los Angeles, California 90095, USA.
authors.forename: Hanjing
authors.initials: H
authors.lastname: Yang
authors.forename: Ju-Huei
authors.initials: JH
authors.lastname: Chiang
authors.forename: Sorel
authors.initials: S
authors.lastname: Fitz-Gibbon
authors.forename: Michel
authors.initials: M
authors.lastname: Lebel
authors.forename: Alessandro A
authors.initials: AA
authors.lastname: Sartori
authors.forename: Joseph
authors.initials: J
authors.lastname: Jiricny
authors.forename: Malgorzata M
authors.initials: MM
authors.lastname: Slupska
authors.forename: Jeffrey H
authors.initials: JH
authors.lastname: Miller
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D015153:Blotting, Western; D004247:DNA / Q000737:chemistry*; D045647:DNA Glycosylases; D019951:DNA Polymerase beta / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000737:chemistry*; D010957:Plasmids / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000494:pharmacology; D013696:Temperature; D019624:Thermoproteaceae / Q000737:chemistry / Q000378:metabolism*; D051981:Uracil-DNA Glycosidase
chemical_list: D000596:Amino Acids; D018809:Proliferating Cell Nuclear Antigen; D011993:Recombinant Fusion Proteins; D012965:Sodium Chloride; D004247:DNA; D005982:Glutathione Transferase; D019951:DNA Polymerase beta; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase