Mutational analysis of Chlorella virus DNA ligase: catalytic roles of domain I and motif VI

Sriskanda, Verl; Shuman, Stewart

doi:10.1093/nar/26.20.4618

Cited by 82 publications

(115 citation statements)

References 17 publications

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“…The k st of the truncated human DNA ligase is ~ 40 min −1 15, the measured value for full‐length EhDNAligI is k st = 2.1 min −1 at 25 °C (Table 1). This value is 20 times lower than the value observed for human DNA ligase I; however, the rate constant is in the range of the observed values for other DNA ligases 34. From the single‐turnover kinetic analysis, we have two observations: first, the deletion of the first 39 amino acids (ΔPIP) does not have a significant effect in rate constant for single‐turnover ligation (full‐length k st = 2.1 min −1 and ΔPIP k st = 2 min −1 ) and second, EhPCNA is not able to significantly alter the rate constant for full‐length and ΔPIP DNA ligases (full‐length k st = 2.2 min −1 , ΔPIP k st = 2.3 min −1 ).…”

Section: Resultscontrasting

confidence: 59%

Proliferating cell nuclear antigen restores the enzymatic activity of a DNA ligase I deficient in DNA binding

Trasviña-Arenas¹,

Cardona-Félix²,

Azuara-Licéaga

et al. 2017

FEBS Open Bio

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Proliferating cell nuclear antigen (PCNA) coordinates multienzymatic reactions by interacting with a variety of protein partners. Family I DNA ligases are multidomain proteins involved in sealing of DNA nicks during Okazaki fragment maturation and DNA repair. The interaction of DNA ligases with the interdomain connector loop (IDCL) of PCNA through its PCNA‐interacting peptide (PIP box) is well studied but the role of the interacting surface between both proteins is not well characterized. In this work, we used a minimal DNA ligase I and two N‐terminal deletions to establish that DNA binding and nick‐sealing stimulation of DNA ligase I by PCNA are not solely dependent on the PIP box–IDCL interaction. We found that a truncated DNA ligase I with a deleted PIP box is stimulated by PCNA. Furthermore, the activity of a DNA ligase defective in DNA binding is rescued upon PCNA addition. As the rate constants for single‐turnover ligation for the full‐length and truncated DNA ligases are not affected by PCNA, our data suggest that PCNA stimulation is achieved by increasing the affinity for nicked DNA substrate and not by increasing catalytic efficiency. Surprisingly C‐terminal mutants of PCNA are not able to stimulate nick‐sealing activity of Entamoeba histolytica DNA ligase I. Our data support the notion that the C‐terminal region of PCNA may be involved in promoting an allosteric transition in E. histolytica DNA ligase I from a spread‐shaped to a ring‐shaped structure. This study suggests that the ring‐shaped PCNA is a binding platform able to stabilize coevolved protein–protein interactions, in this case an interaction with DNA ligase I.

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Section: Resultscontrasting

confidence: 59%

Proliferating cell nuclear antigen restores the enzymatic activity of a DNA ligase I deficient in DNA binding

Trasviña-Arenas¹,

Cardona-Félix²,

Azuara-Licéaga

et al. 2017

FEBS Open Bio

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“…The likely explanation for the ATP effect is that Rnl2 is prone to dissociate from the newly formed RNA-adenylate product of step 2, and that an immediate reaction with ATP to form ligase-adenylate precludes it from rebinding to the RNA-adenylate for subsequent catalysis of strand joining. Similar ATP trapping effects leading to the accumulation of high levels of the adenylated nucleic acid intermediate have been observed for DNA ligases when they react with a substrate containing a 1-nt gap (26,28). Sugino et al (10) showed that catalysis of step 3 by T4 Rnl1 was inhibited by ATP and suggested that the RNA intermediates dissociate from Rnl1 after RNA-adenylate is formed.…”

Section: Resultsmentioning

confidence: 63%

Bacteriophage T4 RNA ligase 2 (gp24.1) exemplifies a family of RNA ligases found in all phylogenetic domains

Shuman²

2002

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

147

174

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RNA ligases participate in repair, splicing, and editing pathways that either reseal broken RNAs or alter their primary structure. Bacteriophage T4 RNA ligase (gp63) is the best-studied member of this class of enzymes, which includes yeast tRNA ligase and trypanosome RNA-editing ligases. Here, we identified another RNA ligase from the bacterial domain-a second RNA ligase (Rnl2) encoded by phage T4. Purified Rnl2 (gp24.1) catalyzes intramolecular and intermolecular RNA strand joining through ligase-adenylate and RNA-adenylate intermediates. Mutational analysis identifies amino acids required for the ligase-adenylation or phosphodiester synthesis steps of the ligation reaction. The catalytic residues of Rnl2 are located within nucleotidyl transferase motifs I, IV, and V that are conserved in DNA ligases and RNA capping enzymes. Rnl2 has scant amino acid similarity to T4 gp63. Rather, Rnl2 exemplifies a distinct ligase family, defined by variant motifs, that includes the trypanosome-editing ligases and a group of putative RNA ligases encoded by eukaryotic viruses (baculoviruses and an entomopoxvirus) and many species of archaea. These findings have implications for the evolution of covalent nucleotidyl transferases and virus-host dynamics based on RNA restriction and repair.T 4 RNA ligase (1) is the founding member of a family of RNA end-joining enzymes involved in RNA repair, splicing, and editing pathways (2-8). T4 RNA ligase joins 3Ј OH and 5Ј PO 4 RNA termini by means of three nucleotidyl transfer steps similar to those of DNA ligases (9-11).Step 1 is the reaction of RNA ligase with ATP to form a covalent ligase-(lysyl-N)-AMP intermediate. In step 2, the AMP is transferred to a 5Ј PO 4 RNA end to form an RNA-adenylate intermediate (AppRNA). In step 3, attack by an RNA 3Ј OH on RNA-adenylate seals the ends and releases AMP. T4 RNA ligase can join two single-stranded RNA molecules without a complementary bridging polynucleotide. T4 RNA ligase can also catalyze intramolecular ligation to form a covalently closed RNA circle. RNA ligase has been a powerful tool in the synthesis of RNAs of defined sequence, RNA 3Ј end modification, RNA 3Ј end-labeling, RNA sequencing, and structural analysis (11).During T4 infection in vivo, the RNA ligase, together with T4 polynucleotide kinase (Pnk), performs an RNA repair function that remodels and then seals broken tRNA ends. In Escherichia coli strains containing the prr locus, the host cell tRNA Lys is cleaved 5Ј to the wobble position by a T4-induced anticodon nuclease. If Pnk and RNA ligase are not present, the synthesis of viral proteins is blocked by depletion of tRNA Lys , and the phage cannot replicate (2,(12)(13)(14). This pathway represents an RNA-based restriction system of host defense against a foreign invader. tRNA anticodon nuclease systems are present in other pathogenic bacteria (14). The bacterial toxins colicins D and E5 are also anticodon nucleases that attack specific tRNAs (15). Thus, RNA repair enzymology has broad significance as a means to combat, or reco...

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“…8D). This ATP-dependent trapping phenomenon has been described previously for eukaryotic viral DNA ligases (17,29) and shown to result from dissociation of the enzyme from the DNA-adenylate and immediate reaction of the free ligase with ATP to yield ligase-AMP, which cannot rebind to AppDNA and thus cannot catalyze the phosphodiester formation step (there being only one adenylatebinding pocket on the enzyme). The inference from the ATPtrapping effect during reaction of LigD with a nicked substrate (which is specific for LigD and not seen for LigB) is that LigD by itself is an inherently inefficient nick-joining enzyme, because it does not remain bound to the step 2 reaction product.…”

Section: Characterization Of M Tuberculosis Ligd-m Tuberculosis Ligmentioning

confidence: 60%

Biochemical and Genetic Analysis of the Four DNA Ligases of Mycobacteria

Gong

Martins

Bongiorno

et al. 2004

Journal of Biological Chemistry

129

142

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Mycobacterium tuberculosis encodes an NAD ؉ -dependent DNA ligase (LigA) plus three distinct ATP-dependent ligase homologs (LigB, LigC, and LigD). Here we purify and characterize the multiple DNA ligase enzymes of mycobacteria and probe genetically whether the ATP-dependent ligases are required for growth of M. tuberculosis. We find significant differences in the reactivity of mycobacterial ligases with a nicked DNA substrate, whereby LigA and LigB display vigorous nick sealing activity in the presence of NAD ؉ and ATP, respectively, whereas LigC and LigD, which have ATPspecific adenylyltransferase activity, display weak nick joining activity and generate high levels of the DNA- DNA ligases are grouped into two families, ATP-dependent ligases and NAD ϩ -dependent ligases, according to their nucleotide substrate requirement (1, 2). The ligase reaction entails three nucleotidyl transfer steps (1). In the first step, attack by ligase on the ␣ phosphorus of ATP or NAD ϩ results in release of PP i or NMN and formation of a covalent ligase-adenylate intermediate. In the second step, the AMP is transferred to the 5Ј end of the 5Ј phosphate-terminated DNA strand to form DNA-adenylate (AppDNA). In the third step, ligase catalyzes attack by a DNA 3Ј-OH on DNA-adenylate to join the two polynucleotides and liberate AMP. ATP-dependent DNA ligases are found in all three domains of life (Bacteria, Archaea, and Eukarya), whereas the NAD ϩ -dependent enzymes are present only in bacteria and entomopoxviruses (1-7).All known bacteria encode a highly conserved NAD ϩ -dependent DNA ligase (LigA). A conditional mutant of Escherichia coli LigA results in growth arrest at the restrictive temperature; thus LigA is essential in E. coli (8,9). LigA is also essential in Salmonella typhimurium, Bacillus subtilis, and Staphylococcus aureus (10 -12). Some bacteria, including E. coli, S. typhimurium, Shigella flexneri, Yersinia pestis, and Pseudomonas putida, have a second NAD ϩ -dependent ligase (13), the function of which is not known.The presumption that bacteria encode only NAD ϩ -dependent DNA ligases was overturned by the demonstration in 1997 of an ATP-dependent ligase in the respiratory pathogen Haemophilus influenzae (5). The 268-amino acid (aa) 1 H. influenzae ligase consists of a minimized catalytic domain. ATP-dependent ligase homologues coexist with NAD ϩ -dependent enzymes in several other bacterial species, including major human pathogens such as Neisseria meningitidis, Y. pestis, Vibrio cholerae, Pseudomonas aeruginosa, and Mycobacterium tuberculosis (3,14,15).Remarkably, M. tuberculosis encodes three distinct ATP-dependent ligase homologues (LigB, LigC, and LigD) plus an NAD ϩ -dependent ligase (LigA) (Fig. 1) (16). To begin to understand the rationale for this plethora of ligases in a single bacterium, we have produced and characterized recombinant versions of the mycobacterial DNA ligase enzymes and probed genetically the essentiality or dispensability of the ATP-dependent ligases for growth of M. tuberculosis. EXPERIMENTAL ...

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Mutational analysis of Chlorella virus DNA ligase: catalytic roles of domain I and motif VI

Cited by 82 publications

References 17 publications

Proliferating cell nuclear antigen restores the enzymatic activity of a DNA ligase I deficient in DNA binding

Proliferating cell nuclear antigen restores the enzymatic activity of a DNA ligase I deficient in DNA binding

Bacteriophage T4 RNA ligase 2 (gp24.1) exemplifies a family of RNA ligases found in all phylogenetic domains

Biochemical and Genetic Analysis of the Four DNA Ligases of Mycobacteria

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