Identification of a Novel Binding Motif in Pyrococcus furiosus DNA Ligase for the Functional Interaction with Proliferating Cell Nuclear Antigen

Kiyonari, Shinichi; Takayama, Kohei; Nishida, Hirokazu; Ishino, Yuji

doi:10.1074/jbc.m603403200

Cited by 35 publications

(43 citation statements)

References 38 publications

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“…It is not clear why the stimulation effect of PCNA on the activity is relatively low at this stage; however, the effect is the same as the case of human UNG2 and PCNA, as reported earlier (16). In our previous study of PfuLig-PfuPCNA interaction, stimulation effect of PCNA on the ligation reaction was especially distinct at high salt reaction conditions, which are more similar to the physiological condition in the P. furiosus cells (35). In the case of PfuUDG, however, the glycosylase activity was drastically decreased at high salt concentrations (over 0.3 M KCl or potassium glutamate) both in the presence and absence of PfuPCNA (data not shown).…”

Section: Resultsmentioning

confidence: 69%

“…The uracil excision efficiency is plotted as a function of the enzyme concentration. Ϫ7 M) and the PfuPolBPfuPCNA interaction (9.9 ϫ 10 Ϫ8 M) determined by our SPR analysis (35,36). These observations suggested that PfuUDG can interact with PfuPCNA via an unidentified binding motif.…”

Section: Resultsmentioning

confidence: 70%

“…In addition, a vast excess of PCNA is required for the functional interaction in vitro, as shown in our previous study (35). This discrepancy could be explained by the difficulty of loading PCNA onto the DNA fragment in the assay mixture.…”

Section: Resultsmentioning

confidence: 75%

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Physical and Functional Interactions between Uracil-DNA Glycosylase and Proliferating Cell Nuclear Antigen from the Euryarchaeon Pyrococcus furiosus

Kiyonari

Uchimura

Shirai

et al. 2008

Journal of Biological Chemistry

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Uracil-DNA glycosylase (UDG) is an important repair enzyme in all organisms to remove uracil bases from DNA. Recent biochemical studies have revealed that human nuclear UDG (UNG2) forms a multiprotein complex in replication foci and initiates the base excision repair pathway by interacting with proliferating cell nuclear antigen (PCNA). Here, we show the physical and functional interactions between UDG and PCNA from the hyperthermophilic euryarchaeon, Pyrococcus furiosus. The physical interaction between the two proteins was identified by a surface plasmon resonance analysis. Furthermore, the uracil glycosylase activity of P. furiosus UDG is stimulated by P. furiosus PCNA (PfuPCNA) in vitro. This stimulatory effect was observed only when wild type PfuPCNA, but not a monomeric PCNA mutant, was present in the reaction. Mutational analyses revealed that our predicted PCNA-binding region (AKTLF) in P. furiosus UDG is actually important for the interaction with PfuPCNA. This is the first report describing the functional interaction between archaeal UDG and PCNA.

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Section: Resultsmentioning

confidence: 69%

Section: Resultsmentioning

confidence: 70%

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Physical and Functional Interactions between Uracil-DNA Glycosylase and Proliferating Cell Nuclear Antigen from the Euryarchaeon Pyrococcus furiosus

Kiyonari

Uchimura

Shirai

et al. 2008

Journal of Biological Chemistry

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“…The hLigI protein also bears a PIP-box in the N-terminal domain (11,12), whereas the corresponding domain is missing in archaeal DNA ligases. Recently, a functional PCNA binding motif of PfuLig, -QKSFF-, was found in a loop within the middle of the DBD, rather than the terminus of the enzyme (13).…”

mentioning

confidence: 99%

Mechanism of replication machinery assembly as revealed by the DNA ligase–PCNA–DNA complex architecture

Mayanagi

Kiyonari

Saito

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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The 3D structure of the ternary complex, consisting of DNA ligase, the proliferating cell nuclear antigen (PCNA) clamp, and DNA, was investigated by single-particle analysis. This report presents the structural view, where the crescent-shaped DNA ligase with 3 distinct domains surrounds the central DNA duplex, encircled by the closed PCNA ring, thus forming a double-layer structure with dual contacts between the 2 proteins. The relative orientations of the DNA ligase domains, which remarkably differ from those of the known crystal structures, suggest that a large domain rearrangement occurs upon ternary complex formation. A second contact was found between the PCNA ring and the middle adenylation domain of the DNA ligase. Notably, the map revealed a substantial DNA tilt from the PCNA ring axis. This structure allows us to propose a switching mechanism for the replication factors operating on the PCNA ring.DNA replication ͉ electron microscopy ͉ single-particle analysis ͉ DNA sliding clamp ͉ protein-DNA complex D NA ligase plays essential roles in various DNA transactions, such as joining Okazaki fragments in DNA replication and nick-sealing at the final step of several DNA repair pathways, including nucleotide excision repair, base excision repair, and mismatch repair (1, 2). The enzyme catalyzes phosphodiester bond formation at the nicks generated within dsDNA, through the well-conserved 3-step reaction using either NAD ϩ or ATP as a cofactor (3). At the first step, the DNA ligase interacts with the nucleotide cofactor to form a covalent ligase-nucleoside monophosphate. At the second step, the bound AMP is transferred to the 5Ј terminus of the DNA, and finally, a phosphodiester bond is formed by a reaction between the 5Ј-DNAadenylate and the 3Ј-hydroxy group.To date, 3 crystal structures of ATP-dependent eukaryotictype DNA ligases have been reported. The structures of the 2 archaeal DNA ligases from Pyrococcus furiosus (PfuLig) and Sulfolobus solfataricus (SsoLig) were determined in the closed (4) and extended forms (5), respectively. The structure of the human DNA ligase 1 (hLigI) in complex with DNA (6) revealed that the enzyme entirely encircles the nicked DNA. All of these DNA ligases are in common composed of 3 domains, designated as the DNA binding domain (DBD), the adenylation domain (AdD), and the OB-fold domain (OBD), in the sequences from the N to C termini. Although the internal architectures of these domains are strikingly similar among the 3 DNA ligases, their relative domain orientations within each enzyme are quite different. Similar to many other replication factors, such as DNA polymerase and Flap endonuclease 1 (FEN1), DNA ligases exhibit the full activity by binding to proliferating cell nuclear antigen (PCNA). A small-angle X-ray scattering analysis revealed that the morphology of SsoLig in complex with PCNA coincides with the extended structure of SsoLig alone (5). However, the structure of the ternary Lig-PCNA-DNA complex remains unknown.PCNA interacts with various protein factors to contr...

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“…However, no typical motif sequence was found in either the N or C terminus of the peptide chain, where the PIP motif usually exists in many PCNA-binding proteins. Therefore, we predicted that the PCNA-binding site of TkoHef is located in the IDR, because PCNA-binding sites also exist in structurally flexible areas, such as looped-out regions, when present in the internal region of a peptide chain (38,40). We subjected TkoHef⌬ID to the same SPR analysis.…”

Section: Prediction Of the Idrs In The Hef/fancm Proteins-thementioning

confidence: 99%

Multiple Interactions of the Intrinsically Disordered Region between the Helicase and Nuclease Domains of the Archaeal Hef Protein

Ishino

Yamagami

Kitamura

et al. 2014

Journal of Biological Chemistry

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Background: Hef/FANCM participates in interstrand cross-link DNA repair. Results: The predicted intrinsically disordered region (IDR) in Hef was experimentally verified. Proliferating cell nuclear antigen and a RecJ-like protein interact with the IDR individually, but not simultaneously. Conclusion:The IDR in Hef interacts with multiple proteins. Significance: Hef may function in DNA repair by association of its IDR with multiple partners.

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Identification of a Novel Binding Motif in Pyrococcus furiosus DNA Ligase for the Functional Interaction with Proliferating Cell Nuclear Antigen

Cited by 35 publications

References 38 publications

Physical and Functional Interactions between Uracil-DNA Glycosylase and Proliferating Cell Nuclear Antigen from the Euryarchaeon Pyrococcus furiosus

Physical and Functional Interactions between Uracil-DNA Glycosylase and Proliferating Cell Nuclear Antigen from the Euryarchaeon Pyrococcus furiosus

Mechanism of replication machinery assembly as revealed by the DNA ligase–PCNA–DNA complex architecture

Multiple Interactions of the Intrinsically Disordered Region between the Helicase and Nuclease Domains of the Archaeal Hef Protein

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