Structural basis for nick recognition by a minimal pluripotent DNA ligase

Nair, Pravin A.; Nandakumar, Jayakrishnan; Smith, Paul; Odell, Mark; Lima, Christopher D.; Shuman, Stewart

doi:10.1038/nsmb1266

Cited by 79 publications

(182 citation statements)

References 50 publications

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“…Consistently, the orientations of the OBD were strikingly different among the 3 crystals, suggesting the high mobility of this domain (4)(5)(6). This unfixed location of the OBD was also highlighted from the crystal structures of smaller ATPdependent DNA ligases (23,24), which exhibited OBD orientations, not only different from our structure, but also from each other. Furthermore, in the EM map, this domain alone lacks contact with PCNA, whereas the DBD and AdD interact with PCNA.…”

Section: Resultssupporting

confidence: 61%

“…The domain configuration in the hLigI-DNA crystal is completely free from the restrictions of contacting PCNA, thus allowing the DNA ligase domains to encircle the DNA more tightly. A similar closed ligase structure, encircling the substrate DNA, was also observed in the chlorella virus DNA ligase-DNA complex, which was crystallized in the absence of PCNA (23). The DBD of PfuLig and hLigI consists of 2 triangular, prism-shaped subdomains (4, 6), which are connected through 2 loops (Fig.…”

Section: Resultsmentioning

confidence: 89%

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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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Section: Resultssupporting

confidence: 61%

Section: Resultsmentioning

confidence: 89%

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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“…Although a detailed understanding of the structural basis for the 3Ј-ribonucleotide effect on sealing by NHEJ ligases awaits their crystallization in complex with nucleic acid substrates, we can draw some inferences from the crystal structures of human LIG1 (27), Chlorella virus ligase (30), and E. coli LigA (35) bound to nicked duplexes with 5Ј-PO 4 or 5Ј-AppN ends. All DNA ligases have a core catalytic domain composed of two structural modules: (i) a nucleotidyltransferase domain that binds the adenylate moieties of ATP (or NAD ϩ ) and AppDNA and (ii) an OB domain that forms part of a C-shaped protein clamp around the DNA duplex.…”

Section: Discussionmentioning

confidence: 99%

“…To verify the specificity of the ribonucleotide effect on NHEJ ligases and to exclude the possibility that the D11R1 and D12 substrates are inherently differentially sealable, we tested the substrates in parallel with Chlorella virus DNA ligase, which is the smallest eukaryotic DNA ligase known and which has been extensively characterized, biochemically and structurally (26,27). Chlorella virus ligase was equally adept at sealing the D11R1 and D12 nicks (Fig.…”

Section: Pseudomonas Ligd Preferentially Seals a 3ј-monoribonucleotidmentioning

confidence: 99%

Bacterial Nonhomologous End Joining Ligases Preferentially Seal Breaks with a 3′-OH Monoribonucleotide

Zhu

Shuman

2008

Journal of Biological Chemistry

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Many bacterial species have a nonhomologous end joining system of DNA repair driven by dedicated DNA ligases (LigD and LigC). LigD is a multifunctional enzyme composed of a ligase domain fused to two other catalytic modules: a polymerase that preferentially adds ribonucleotides to double-strand break ends and a phosphoesterase that trims 3 -oligoribonucleotide tracts until only a single 3 -ribonucleotide remains. LigD and LigC have a feeble capacity to seal 3 -OH/5 -PO 4 DNA nicks. Here, we report that nick sealing by LigD and LigC enzymes is stimulated by the presence of a single ribonucleotide at the broken 3 -OH end. The ribonucleotide effect on LigD and LigC is specific for the 3 -terminal nucleotide and is either diminished or abolished when additional vicinal ribonucleotides are present. No such 3 -ribonucleotide effect is observed for bacterial LigA or Chlorella virus ligase. We found that in vitro repair of a double-strand break by Pseudomonas LigD requires the polymerase module and results in incorporation of an alkalilabile ribonucleotide at the repair junction. These results illuminate an underlying logic for the domain organization of LigD, whereby the polymerase and phosphoesterase domains can heal the broken 3 -end to generate the monoribonucleotide terminus favored by the nonhomologous end joining ligases.Direct evidence that bacteria catalyze DNA double-strand break (DSB) 2 repair via nonhomologous end joining (NHEJ) emerged from studies of the repair of linear plasmid DNAs transfected into mycobacteria (1). NHEJ entails approximation of the broken DNA ends, aided by the bacterial end-binding protein Ku, followed by sealing of at least one of the broken strands by a specialized bacterial ATP-dependent DNA ligase, either LigD or LigC (1-9, 36). Unlike homologous recombination, which is generally error-free, NHEJ can be either faithful or mutagenic. The signature feature of NHEJ in mycobacteria is that half of the repair events at blunt or complementary 5Ј-overhang DSBs are unfaithful because the DSB ends are extended by polymerases or resected by nucleases prior to sealing by ligase (1,8,9,36). Because NHEJ does not rely on a homologous DNA template, it can operate when only one chromosomal copy is available, e.g. during late stationary phase or after sporulation (10 -13). Whereas mutagenic DSB repair might seem counterproductive, it can be advantageous, especially if the alternative is death of the quiescent bacterium or spore.Biochemical, structural, and genetic studies of the NHEJ ligases and Ku proteins of Mycobacterium, Pseudomonas, Bacillus, and Agrobacterium are beginning to define a pathway with unique features and distinctive enzymatic components (1, 4 -9, 14 -17, 36). Ku and LigD are critical agents of the pathway. The efficiency of plasmid-based NHEJ of blunt and 5Ј-overhang DSBs in mycobacteria is reduced several hundredfold by deletion of Ku and by ϳ100-fold by deletion of LigD (1, 36). LigC provides an efficient backup sealing function in mycobacteria when LigD sealing activity is a...

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“…Escherichia coli LigA and related NAD-dependent ligases have a C-terminal helix-hairpin-helix domain that functions similarly to the N-terminal DBD of LigI (20,21). Even Chlorella virus ligase, which is the smallest known eukaryotic DNA ligase and lacks a DBD, has a small latch structure that allows the enzyme to fully encircle the DNA (22).…”

mentioning

confidence: 99%