Direct Interaction between Uracil-DNA Glycosylase and a Proliferating Cell Nuclear Antigen Homolog in the CrenarchaeonPyrobaculum aerophilum

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 crenarchaeonPyrobaculum aerophilum (Topt = 100 °C). We demonstrate that the Pa-UDGa-Pa-PCNA1 complex is thermostable, and two hydrophobic amino acid residues onPa-UDGa (Phe191 and Leu192) 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 thatPa-UDGa may be a functional analog of hUNG2 for PCNA-dependent postreplicative removal of misincorporated uracil. 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 crenarchaeonPyrobaculum aerophilum (Topt = 100 °C). We demonstrate that the Pa-UDGa-Pa-PCNA1 complex is thermostable, and two hydrophobic amino acid residues onPa-UDGa (Phe191 and Leu192) 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 thatPa-UDGa may be a functional analog of hUNG2 for PCNA-dependent postreplicative removal of misincorporated uracil. proliferating cell nuclear antigen uracil-DNA glycosylase P. aerophilum UDGa family B DNA polymerase P. aerophilum Pol B3 flap endonuclease glutathione S-transferase nickel-nitrilotriacetic acid P. aerophilum human major nuclear uracil-DNA glycosylase Proliferating cell nuclear antigen (PCNA)1 is essential for life. It is a processivity factor for DNA polymerase, forming a toroidal-shaped trimer acting as a sliding clamp on duplex DNA (1Kong X.P. Onrust R. O'Donnell M. Kuriyan J. Cell. 1992; 69: 425-437Abstract Full Text PDF PubMed Scopus (645) Google Scholar, 2Krishna T.S. Kong X.P. Gary S. Burgers P.M. Kuriyan J. Cell. 1994; 79: 1233-1243Abstract Full Text PDF PubMed Scopus (766) Google Scholar, 3Moarefi I. Jeruzalmi D. Turner J. O'Donnell M. Kuriyan J. J. Mol. Biol. 2000; 296: 1215-1223Crossref PubMed Scopus (139) Google Scholar, 4Matsumiya S. Ishino Y. Morikawa K. Protein Sci. 2001; 10: 17-23Crossref PubMed Scopus (134) Google Scholar). Its function requires another protein, the clamp loader replication factor C, to load it onto the circular DNAs (5Pisani F.M., De Felice M. Carpentieri F. Rossi M. J. Mol. Biol. 2000; 301: 61-73Crossref PubMed Scopus (49) Google Scholar, 6Oyama T. Ishino Y. Cann I.K. Ishino S. Morikawa K. Mol. Cell. 2001; 8: 455-463Abstract Full Text Full Text PDF PubMed Scopus (64) Google Scholar, 7Jeruzalmi D. Yurieva O. Zhao Y. Young M. Stewart J. Hingorani M. O'Donnell M. Kuriyan J. Cell. 2001; 106: 417-428Abstract Full Text Full Text PDF PubMed Scopus (210) Google Scholar, 8Jeruzalmi D. O'Donnell M. Kuriyan J. Cell. 2001; 106: 429-441Abstract Full Text Full Text PDF PubMed Scopus (253) Google Scholar, 9Cann I.K. Ishino S. Yuasa M. Daiyasu H. Toh H. Ishino Y. J. Bacteriol. 2001; 183: 2614-2623Crossref PubMed Scopus (62) Google Scholar). PCNA is present in eukaryotes and its functional analog, the β subunit of DNA polymerase III holoenzyme, is present in bacteria (10Hingorani M.M. O'Donnell M. Curr. Biol. 2000; 10: R25-R29Abstract Full Text Full Text PDF PubMed Scopus (105) Google Scholar, 11Bruck I. O'Donnell M. Genome Biol. 2001; 2: 3001.1-3001.3Crossref Google Scholar). More than a dozen classes of eukaryotic PCNA-binding proteins have been shown to interact with the PCNA sliding clamp, linking PCNA to several important biological processes beyond DNA replication, such as DNA repair and cell cycle regulation (12Kelman Z. Hurwitz J. Trends Biochem. Sci. 1998; 23: 236-238Abstract Full Text Full Text PDF PubMed Scopus (136) Google Scholar, 13Tsurimoto T. Front. Biosci. 1999; 4: D849-D858Crossref PubMed Google Scholar, 14Krude T. Curr. Biol. 1999; 9: R394-R396Abstract Full Text Full Text PDF PubMed Scopus (15) Google Scholar, 15Kolodner R.D. Marsischky G.T. Curr. Opin. Genet. Dev. 1999; 9: 89-96Crossref PubMed Scopus (733) Google Scholar, 16Warbrick E. Bioessays. 2000; 22: 997-1006Crossref PubMed Scopus (353) Google Scholar, 17Gulbis J.M. Kelman Z. Hurwitz J. O'Donnell M. Kuriyan J. Cell. 1996; 87: 297-306Abstract Full Text Full Text PDF PubMed Scopus (655) Google Scholar, 18Shamoo Y. Steitz T.A. Cell. 1999; 99: 155-166Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar). In many cases, PCNA binding partners interact with PCNA through a conserved motif identified as QXX(L/M/I)XX(F/Y/H)(F/Y) that is usually located near either the amino or the carboxyl terminus. One important example of a eukaryotic PCNA-binding protein involved in DNA repair is human major nuclear uracil-DNA glycosylase (hUNG2), which removes uracil from misincorporated dUMP residues in an immediate postreplicative process (19Otterlei M. Warbrick E. Nagelhus T.A. Haug T. Slupphaug G. Akbari M. Aas P.A. Steinsbekk K. Bakke O. Krokan H.E. EMBO J. 1999; 18: 3834-3844Crossref PubMed Scopus (300) Google Scholar, 20Krokan H.E. Otterlei M. Nilsen H. Kavli B. Skorpen F. Andersen S. Skjelbred C. Akbari M. Aas P.A. Slupphaug G. Prog. Nucleic Acid Res. Mol. Biol. 2001; 68: 365-386Crossref PubMed Google Scholar). hUNG2 interacts with PCNA through its PCNA binding site, 4QKTLYSFF11, which is located near the amino terminus of hUNG2. Recently PCNA sequence homologs have been identified in Archaea (21De Felice M. Sensen C.W. Charlebois R.L. Rossi M. Pisani F.M. J. Mol. Biol. 1999; 291: 47-57Crossref PubMed Scopus (49) Google Scholar,22Cann I.K. Ishino S. Hayashi I. Komori K. Toh H. Morikawa K. Ishino Y. J. Bacteriol. 1999; 181: 6591-6599Crossref PubMed Google Scholar). So far, each of the 10 completely sequenced archaeal genomes contains at least one putative PCNA homolog (23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar). There is a distinction found between the two major subdomains of the Archaea, Crenarchaeota and Euryarchaeota (23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar). Whereas each euryarchaeal genome tends to have one PCNA homolog, each crenarchaeal genome has two or three putative PCNA homologs (21De Felice M. Sensen C.W. Charlebois R.L. Rossi M. Pisani F.M. J. Mol. Biol. 1999; 291: 47-57Crossref PubMed Scopus (49) Google Scholar, 23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar, 24Fitz-Gibbon S.T. Ladner H. Kim U.J. Stetter K.O. Simon M.I. Miller J.H. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 984-989Crossref PubMed Scopus (197) Google Scholar). Biochemical studies have been conducted with several of the archaeal PCNA homologs, including a PCNA homolog from the euryarchaeote Pyrococcus furiosus and two PCNA homologs from the crenarchaeote Sulfolobus solfataricus (21De Felice M. Sensen C.W. Charlebois R.L. Rossi M. Pisani F.M. J. Mol. Biol. 1999; 291: 47-57Crossref PubMed Scopus (49) Google Scholar, 22Cann I.K. Ishino S. Hayashi I. Komori K. Toh H. Morikawa K. Ishino Y. J. Bacteriol. 1999; 181: 6591-6599Crossref PubMed Google Scholar). These studies have confirmed that all of them are processivity factors for their corresponding DNA polymerases. In Archaea, in addition to the PCNA clamp loader (replication factor C), two classes of archaeal proteins have so far been identified as PCNA-binding proteins, based on in vitro binding study and crystal structure analysis. They are family B DNA polymerase (Pol B) (22Cann I.K. Ishino S. Hayashi I. Komori K. Toh H. Morikawa K. Ishino Y. J. Bacteriol. 1999; 181: 6591-6599Crossref PubMed Google Scholar) and flap endonuclease (FEN) (25Hwang K.Y. Baek K. Kim H.Y. Cho Y. Nat. Struct. Biol. 1998; 5: 668-670Crossref PubMed Scopus (17) Google Scholar, 26Hosfield D.J. Mol C.D. Shen B. Tainer J.A. Cell. 1998; 95: 135-146Abstract Full Text Full Text PDF PubMed Scopus (226) Google Scholar, 27Hosfield D.J. Frank G. Weng Y. Tainer J.A. Shen B. J. Biol. Chem. 1998; 273: 27154-27161Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar, 28Rao H.G. Rosenfeld A. Wetmur J.G. J. Bacteriol. 1998; 180: 5406-5412Crossref PubMed Google Scholar, 29Matsui E. Kawasaki S. Ishida H. Ishikawa K. Kosugi Y. Kikuchi H. Kawarabayashi Y. Matsui I. J. Biol. Chem. 1999; 274: 18297-18309Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar), both proteins known to interact with PCNA in eukaryotes. The proposed putative PCNA-binding motifs in these archaeal PCNA-binding proteins are quite similar to the conserved PCNA-binding motif identified in eukaryotic PCNA-binding proteins (22Cann I.K. Ishino S. Hayashi I. Komori K. Toh H. Morikawa K. Ishino Y. J. Bacteriol. 1999; 181: 6591-6599Crossref PubMed Google Scholar, 27Hosfield D.J. Frank G. Weng Y. Tainer J.A. Shen B. J. Biol. Chem. 1998; 273: 27154-27161Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar). However, these putative PCNA binding sites have not been verified by mutation analysis. In this study, we report identification of another archaeal PCNA-binding protein, Pyrobaculum aerophilum uracil-DNA glycosylase (Pa-UDGa), and the biochemical confirmation of its interaction with PCNA via the PCNA-binding motif. P. aerophilum is a hyperthermophile with an optimal growth temperature of 100 °C and a member of the crenarchaeal subdomain of Archaea (30Völkl P. Huber R. Drobner E. Rachel R. Burggraf S. Trincone A. Stetter K.O. Appl. Environ. Microbiol. 1993; 59: 2918-2926Crossref PubMed Google Scholar). The biochemical characterization of uracil-DNA glycosylase activity of Pa-UDGa was recently published (31Sartori A.A. Schar P. Fitz-Gibbon S. Miller J.H. Jiricny J. J. Biol. Chem. 2001; 276: 29979-29986Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar). Analysis of the complete genome sequence of P. aerophilum revealed two putative PCNA homologs (24Fitz-Gibbon S.T. Ladner H. Kim U.J. Stetter K.O. Simon M.I. Miller J.H. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 984-989Crossref PubMed Scopus (197) Google Scholar),Pa-PCNA1 and Pa-PCNA2, as expected for a crenarchaeote (23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar). We demonstrate that Pa-UDGa preferentially binds to Pa-PCNA1, similar to two otherP. aerophilum PCNA-binding proteins,Pa-FEN and P. aerophilum DNA polymerase B3 (Pa-Pol B3). Pa-UDGa's ability to bind toPa-PCNA1 resembles the eukaryotic PCNA-binding protein hUNG2, which belongs to a distinctly different UDG family due to low amino acid sequence similarity to Pa-UDGa. Our results raise the possibility that Pa-UDGa may be a functional analog of hUNG2 for PCNA-dependent postreplicative removal of misincorporated uracil. P. aerophilumgenomic DNA was prepared as described previously (32Fitz-Gibbon S. Choi A.J. Miller J.H. Stetter K.O. Simon M.I. Swanson R. Kim U.J. Extremophiles. 1997; 1: 36-51Crossref PubMed Scopus (58) Google Scholar). The coding regions for Pa-UDGa (PAE0651, Protein Data Bank accession number AAL62921) and Pa-FEN (PAE0698, Protein Data Bank accession number AAL62961) were amplified by PCR usingP. aerophilum genomic DNA as template with their corresponding primer pairs. The PCR products were cloned into a pCR2.1-TOPO vector using a TOPO TA cloning kit (Invitrogen). The primer information can be obtained upon request. The full-length Pa-UDGa gene was amplified by PCR using pCR2.1TOPOPa-UDGa as template, cloned into a pGEX-2TK vector (Amersham Biosciences) at the BamHI site to create a plasmid that expresses a fusion protein of glutathioneS-transferase (GST) and Pa-UDGa. The full-lengthPa-FEN gene was amplified by PCR using pCR2.1TOPOPa-FEN as template and cloned into a pGEX-2TK vector (Amersham Biosciences) at the EcoRI site to create a plasmid that expresses GST-Pa-FEN fusion protein. TwoPa-Pol B3 (PAE2109, Protein Data Bank accession numberAAL63952) fragments containing the carboxyl-terminal amino acid residues and amino acid residues were amplified by PCR using P. aerophilum genomic DNA as template, cloned into a pGEX-2TK vector (Amersham Biosciences) between the BamHI and EcoRI sites to create two that B3 amino fusion and B3 amino The full-length Protein Data Bank accession number was amplified by PCR using P. aerophilum genomic cloned into a vector between and sites to create a plasmid that expresses the The full-length gene Protein Data Bank accession number was amplified by PCR using P. aerophilum genomic cloned into a vector between the and sites to create a plasmid that expresses the The PCNA M. P. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus (210) Google Scholar) was into a vector between and EcoRI was into a vector between the BamHI and sites to create a plasmid that expresses the binding the of Pa-UDGa was into vector to create a plasmid that expresses an Pa-UDGa protein. The was into a vector to create a plasmid that expresses the of the The amino acid fragments of Pa-UDGa and were amplified by PCR using pCR2.1TOPOPa-UDGa as The products were into a pGEX-2TK vector (Amersham Biosciences) at the BamHI site to create fusion protein The Pa-UDGa was by PubMed Scopus Google Scholar) using pCR2.1TOPOPa-UDGa as The obtained PCR products were cloned into a pGEX-2TK vector (Amersham Biosciences) at site to create an plasmid for and the were verified by DNA using a DNA kit The Pa-UDGa was by The PCR products were cloned into a pCR2.1-TOPO vector using a TOPO TA cloning kit (Invitrogen). containing the with two amino acid was into a pGEX-2TK vector (Amersham Biosciences) at the BamHI site to create an plasmid for and the were verified by DNA using a DNA kit The Pa-FEN was by The PCR products were cloned into a pCR2.1-TOPO vector using a TOPO TA cloning kit (Invitrogen). containing the full-lengthPa-FEN with two amino acid was into a pGEX-2TK vector (Amersham Biosciences) at the EcoRI site to create an plasmid for Pa-FEN and the were verified by DNA using a DNA kit of plasmid Pa-PCNA1, or were to 100 of with and The of Pa-PCNA1, Pa-PCNA2, or PCNA was with growth for at The plasmid expresses the protein by the gene to the of were in and with and by The cell were by at The protein was by protein The cell were as at of was obtained by of of cell at °C for 10 by at for The was at protein the was obtained for the binding by a similar as described E. plasmid was for the of cell a was to the cell to a in its the E. was as the to all fusion proteins and the protein. The cell were prepared in as described Protein of the cell was by protein were with the for at with with fusion proteins were in at the of each fusion protein for the a of proteins was by in by and by The of each fusion protein for the was for each based on the of proteins by were to to the of were with PCNA at °C for with of the proteins were by in of by and to for homologs were using the with and by (Amersham Pa-UDGa protein was in and the cell was prepared by in B with were to the cell and at °C for with the the were to A. The containing Pa-UDGa were with the in its or with E. cell containing vector °C a for the a containing the and of Pa-UDGa was with of at the were into two one was at °C in a for with of at the was on with of A. were at proteins were by in by and by a member of the archaeal subdomain P. aerophilum contains two putative PCNA homologs, and Pa-PCNA2, identified by amino acid sequence 24Fitz-Gibbon S.T. Ladner H. Kim U.J. Stetter K.O. Simon M.I. Miller J.H. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 984-989Crossref PubMed Scopus (197) Google amino acid sequence The amino acid sequence between the two putative homologs and the eukaryotic or archaeal from to putative homologs the conserved motif located near the carboxyl which may interact with the clamp replication factor C (23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar). of PCNA homologs in that the homologs in into two with a the of the crenarchaeal (23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar). and were cloned on the vector and as proteins in E. and and homologs for in vitro binding we of the of proteins from and the E. cell each homolog to The and were a and and eukaryotic PCNA protein was in the and was found to be the and and the and have of and the on the was and for The of on the was in two S. solfataricus (21De Felice M. Sensen C.W. Charlebois R.L. Rossi M. Pisani F.M. J. Mol. Biol. 1999; 291: 47-57Crossref PubMed Scopus (49) Google Scholar) and eukaryotic PCNA homologs P. Z. PubMed Scopus Google Scholar) for B3 was for binding to and Analysis of the B3 protein sequence revealed that its carboxyl-terminal contains a putative PCNA-binding motif which is similar to the putative PCNA-binding motifs of several archaeal Pol B homologs by Ishino and (22Cann I.K. Ishino S. Hayashi I. Komori K. Toh H. Morikawa K. Ishino Y. J. Bacteriol. 1999; 181: 6591-6599Crossref PubMed Google Scholar). S-transferase (GST) fusion proteins were for two fragments containing the carboxyl-terminal of B3 amino amino and In the interaction binding to either of the C, However, binding to was with both of the B3 and In a binding was C, and The binding toPa-PCNA1 by the carboxyl-terminal B3 for an in direct interaction between B3 and fusion protein with Pa-UDGa was for the study of in vitro direct binding of Pa-UDGa to two homologs using the interaction which was to be a PCNA-binding protein based on studies D.J. Frank G. Weng Y. Tainer J.A. Shen B. J. Biol. Chem. 1998; 273: 27154-27161Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar), was for the binding to and both and GST-Pa-FEN to and to was not with either fusion the binding and Pa-FEN to for a direct in interaction The of on the of a complex between Pa-UDGa was was in the of and was at than C The of temperature on the of the complex was using a and a in its binding was at °C and was a at °C and the the protein binding to the not of Pa-UDGa and Pa-FEN to the eukaryotic PCNA was was our not The interaction between Pa-UDGa was verified by the regions on Pa-UDGa for PCNA three fusion proteins that regions were and for binding activity using the interaction not bind and this fusion protein an not However, both fusion proteins containing the carboxyl-terminal of Pa-UDGa binding activity not shown for C and for These results demonstrate that the acid near carboxyl terminus is for PCNA Pa-UDGa with the identified acid were to amino acid residues for PCNA binding studies have shown that many PCNA-binding proteins the PCNA-binding motif and that the two hydrophobic amino acid residues this motif are involved in the interaction with PCNA I. B. Hurwitz J. S. Mol. Biol. 2001; PubMed Scopus Google Scholar). Analysis of the Pa-UDGa amino acid sequence revealed two putative PCNA-binding motifs near the carboxyl terminus The sequence for the is which contains the eukaryotic PCNA-binding sequence The sequence for the which contains two hydrophobic amino acid residues (Phe191 and Leu192) to the carboxyl terminus Pa-UDGa were for the binding each with two amino acid in the putative PCNA-binding motif or motif the in its putative PCNA-binding motif near its carboxyl terminus was in the the Pa-FEN to bind to The Pa-UDGa in the putative binding motif was of binding to C However, the binding was in the Pa-UDGa in the putative motif C and These results that and of and and of Pa-UDGa are for the binding of Pa-FEN and Pa-UDGa to the of archaeal PCNA-binding proteins, we the protein of the P. aerophilum genome with identified putative PCNA-binding motifs using R. Appl. Biosci. 1994; 10: Google Scholar). In this Pa-UDGa was identified as a putative PCNA-binding protein. We in vitro biochemical of Pa-UDGa binding to two putative P. aerophilum PCNA homologs, and Pa-PCNA2, using the interaction Our results that Pa-UDGa preferentially binds toPa-PCNA1 to a protein the binding between Pa-UDGa and is as Pa-UDGa has binding to the hydrophobic amino acid and located near the carboxyl terminus of are crucial for PCNA binding these the between and two P. aerophilum proteins, Pa-FEN and and the carboxyl-terminal of for as a functional PCNA homolog in P. this we were to to the of the PCNA binding of Pa-UDGa due to the of a in P. However, identification of Pa-UDGa as an archaeal PCNA-binding protein and its amino acid residues for PCNA binding is the in its biological in The PCNA binding activity in Pa-UDGa from this study and in hUNG2 from studies (19Otterlei M. Warbrick E. Nagelhus T.A. Haug T. Slupphaug G. Akbari M. Aas P.A. Steinsbekk K. Bakke O. Krokan H.E. EMBO J. 1999; 18: 3834-3844Crossref PubMed Scopus (300) Google Scholar) the possibility that Pa-UDGa may be a functional analog of and hUNG2 to two UDG due to the low sequence similarity between them (31Sartori A.A. Schar P. Fitz-Gibbon S. Miller J.H. Jiricny J. J. Biol. Chem. 2001; 276: 29979-29986Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, M. Curr. Biol. 1999; 9: Full Text Full Text PDF PubMed Scopus (62) Google Scholar). In is an protein (31Sartori A.A. Schar P. Fitz-Gibbon S. Miller J.H. Jiricny J. J. Biol. Chem. 2001; 276: 29979-29986Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar) due to the of an J.A. De Y. A.A. Jiricny J. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar), hUNG2 is these both Pa-UDGa and hUNG2 have similar uracil glycosylase and both interact with PCNA through binding at either the carboxyl-terminal or the hUNG2, it is that Pa-UDGa may be a functional analog of hUNG2 for PCNA-dependent postreplicative removal of misincorporated uracil (19Otterlei M. Warbrick E. Nagelhus T.A. Haug T. Slupphaug G. Akbari M. Aas P.A. Steinsbekk K. Bakke O. Krokan H.E. EMBO J. 1999; 18: 3834-3844Crossref PubMed Scopus (300) Google Scholar, 20Krokan H.E. Otterlei M. Nilsen H. Kavli B. Skorpen F. Andersen S. Skjelbred C. Akbari M. Aas P.A. Slupphaug G. Prog. Nucleic Acid Res. Mol. Biol. 2001; 68: 365-386Crossref PubMed Google Scholar). this two or three putative PCNA homologs have been identified in the completely sequenced Crenarchaeota including P. S. and (21De Felice M. Sensen C.W. Charlebois R.L. Rossi M. Pisani F.M. J. Mol. Biol. 1999; 291: 47-57Crossref PubMed Scopus (49) Google Scholar, 23Iwai T. Kurosawa N. Itoh Y.H. Horiuchi T. Extremophiles. 2000; 4: 357-364Crossref PubMed Scopus (15) Google Scholar). The of more than one PCNA homolog may either functional or functional with S. solfataricus and B demonstrate that both homologs are processivity factors for the family B DNA polymerase (Pol their different (21De Felice M. Sensen C.W. Charlebois R.L. Rossi M. Pisani F.M. J. Mol. Biol. 1999; 291: 47-57Crossref PubMed Scopus (49) Google Scholar). Our study with and a functional between two to Pa-UDGa and Pa-FEN is only with the carboxyl-terminal B3 preferentially binds to we the possibility that the is our of its and binding to the carboxyl-terminal of our results that may be the major protein with similar to PCNA in eukaryotes. is and which proteins it interact to be Our not the possibility and may to functional homologs are present in Archaea and Eukarya D.J. Frank G. Weng Y. Tainer J.A. Shen B. J. Biol. Chem. 1998; 273: 27154-27161Abstract Full Text Full Text PDF PubMed Scopus (68) Google family are found in Archaea and M. Curr. Biol. 1999; 9: Full Text Full Text PDF PubMed Scopus (62) Google Scholar, M. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). The amino acid sequence of the carboxyl-terminal regions of and UDG homologs are shown in the conserved PCNA-binding motif was near the carboxyl terminus for all archaeal homologs 27Hosfield D.J. Frank G. Weng Y. Tainer J.A. Shen B. J. Biol. Chem. 1998; 273: 27154-27161Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar), carboxyl-terminal were for archaeal UDG homologs homologs PCNA-binding from S. from and from Biochemical be to these binding motifs and to the archaeal homologs bind

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