scholarly journals Two DNA-binding and Nick Recognition Modules in Human DNA Ligase III

2008 ◽  
Vol 283 (16) ◽  
pp. 10764-10772 ◽  
Author(s):  
Elizabeth Cotner-Gohara ◽  
In-Kwon Kim ◽  
Alan E. Tomkinson ◽  
Tom Ellenberger
Keyword(s):  
Dna Binding ◽  
Dna Ligase ◽  
Human Dna ◽  
Dna Ligase Iii

scholarly journals Human DNA Ligase III Recognizes DNA Ends by Dynamic Switching between Two DNA-Bound States

Biochemistry ◽  
2010 ◽  
Vol 49 (29) ◽  
pp. 6165-6176 ◽  
Author(s):  
Elizabeth Cotner-Gohara ◽  
In-Kwon Kim ◽  
Michal Hammel ◽  
John A. Tainer ◽  
Alan E. Tomkinson ◽  
...  
Keyword(s):  
Bound States ◽  
Dna Ligase ◽  
Dynamic Switching ◽  
Human Dna ◽  
Dna Ends ◽  
Dna Ligase Iii

scholarly journals The Human DNA Ligase III Gene Encodes Nuclear and Mitochondrial Proteins

1999 ◽  
Vol 19 (5) ◽  
pp. 3869-3876 ◽  
Author(s):  
Uma Lakshmipathy ◽  
Colin Campbell
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Mitochondrial Protein ◽  
Nuclear Accumulation ◽  
Dna Ligase ◽  
Mitochondrial Enzymes ◽  
Antisense Mrna ◽  
Human Dna ◽  
Green Fluorescent ◽  
Dna Ligase Iii

ABSTRACT We provide evidence that the human DNA ligase III gene encodes a mitochondrial form of this enzyme. First, the DNA ligase III cDNA contains an in-frame ATG located upstream from the putative translation initiation start site. The DNA sequence between these two ATG sites encodes an amphipathic helix similar to previously identified mitochondrial targeting peptides. Second, recombinant green fluorescent protein harboring this sequence at its amino terminus was efficiently targeted to the mitochondria of Cos-1 monkey kidney cells. In contrast, native green fluorescent protein distributed to the cytosol. Third, a series of hemagglutinin-DNA ligase III minigene constructs were introduced into Cos-1 cells, and immunocytochemistry was used to determine subcellular localization of the epitope-tagged DNA ligase III protein. These experiments revealed that inactivation of the upstream ATG resulted in nuclear accumulation of the DNA ligase III protein, whereas inactivation of the downstream ATG abolished nuclear localization and led to accumulation within the mitochondrial compartment. Fourth, mitochondrial protein extracts prepared from human cells overexpressing antisense DNA ligase III mRNA possessed substantially less DNA ligase activity than did mitochondrial extracts prepared from control cells. DNA end-joining activity was also substantially reduced in extracts prepared from antisense mRNA-expressing cells. From these results, we conclude that the human DNA ligase III gene encodes both nuclear and mitochondrial enzymes. DNA ligase plays a central role in DNA replication, recombination, and DNA repair. Thus, identification of a mitochondrial form of this enzyme provides a tool with which to dissect mammalian mitochondrial genome dynamics.

scholarly journals Mammalian DNA ligase III: molecular cloning, chromosomal localization, and expression in spermatocytes undergoing meiotic recombination.

1995 ◽  
Vol 15 (10) ◽  
pp. 5412-5422 ◽  
Author(s):  
J Chen ◽  
A E Tomkinson ◽  
W Ramos ◽  
Z B Mackey ◽  
S Danehower ◽  
...  
Keyword(s):  
Meiotic Recombination ◽  
Chromosome 17 ◽  
Dna Ligase ◽  
Cdna Clones ◽  
Dna Ligases ◽  
Cell Extracts ◽  
Human Dna ◽  
Dna Ligase I ◽  
Dna Ligase Iii

Three biochemically distinct DNA ligase activities have been identified in mammalian cell extracts. We have recently purified DNA ligase II and DNA ligase III to near homogeneity from bovine liver and testis tissue, respectively. Amino acid sequencing studies indicated that these enzymes are encoded by the same gene. In the present study, human and murine cDNA clones encoding DNA ligase III were isolated with probes based on the peptide sequences. The human DNA ligase III cDNA encodes a polypeptide of 862 amino acids, whose sequence is more closely related to those of the DNA ligases encoded by poxviruses than to replicative DNA ligases, such as human DNA ligase I. In vitro transcription and translation of the cDNA produced a catalytically active DNA ligase similar in size and substrate specificity to the purified bovine enzyme. The DNA ligase III gene was localized to human chromosome 17, which eliminated this gene as a candidate for the cancer-prone disease Bloom syndrome that is associated with DNA joining abnormalities. DNA ligase III is ubiquitously expressed at low levels, except in the testes, in which the steady-state levels of DNA ligase III mRNA are at least 10-fold higher than those detected in other tissues and cells. Since DNA ligase I mRNA is also present at high levels in the testes, we examined the expression of the DNA ligase genes during spermatogenesis. DNA ligase I mRNA expression correlated with the contribution of proliferating spermatogonia cells to the testes, in agreement with the previously defined role of this enzyme in DNA replication. In contrast, elevated levels of DNA ligase III mRNA were observed in primary spermatocytes undergoing recombination prior to the first meiotic division. Therefore, we suggest that DNA ligase III seals DNA strand breaks that arise during the process of meiotic recombination in germ cells and as a consequence of DNA damage in somatic cells.

scholarly journals 1HN, 13C, and 15N backbone resonance assignments of the human DNA ligase 3 DNA-binding domain (residues 257-477)

2019 ◽  
Vol 13 (2) ◽  
pp. 305-308
Author(s):  
Braden M. Roth ◽  
Kristen M. Varney ◽  
Hui Yang ◽  
David J. Weber ◽  
Alan E. Tomkinson
Keyword(s):  
Dna Binding ◽  
Resonance Assignments ◽  
Binding Domain ◽  
Dna Ligase ◽  
Dna Binding Domain ◽  
Backbone Resonance Assignments ◽  
Backbone Resonance ◽  
Human Dna

scholarly journals Human DNA ligase III bridges two DNA ends to promote specific intermolecular DNA end joining

2015 ◽  
Vol 43 (14) ◽  
pp. 7021-7031 ◽  
Author(s):  
Vandna Kukshal ◽  
In-Kwon Kim ◽  
Gregory L. Hura ◽  
Alan E. Tomkinson ◽  
John A. Tainer ◽  
...  
Keyword(s):  
Dna Ligase ◽  
End Joining ◽  
Human Dna ◽  
Dna Ends ◽  
Dna Ligase Iii

scholarly journals The DNA binding domain of human DNA ligase I interacts with both nicked DNA and the DNA sliding clamps, PCNA and hRad9-hRad1-hHus1

DNA Repair ◽  
2009 ◽  
Vol 8 (8) ◽  
pp. 912-919 ◽  
Author(s):  
Wei Song ◽  
John M. Pascal ◽  
Tom Ellenberger ◽  
Alan E. Tomkinson
Keyword(s):  
Dna Binding ◽  
Binding Domain ◽  
Dna Ligase ◽  
Dna Binding Domain ◽  
Human Dna ◽  
Sliding Clamps ◽  
Dna Ligase I

scholarly journals An interaction between the mammalian DNA repair protein XRCC1 and DNA ligase III.

1994 ◽  
Vol 14 (1) ◽  
pp. 68-76 ◽  
Author(s):  
K W Caldecott ◽  
C K McKeown ◽  
J D Tucker ◽  
S Ljungquist ◽  
L H Thompson
Keyword(s):  
Dna Repair ◽  
Affinity Chromatography ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Affinity Purification ◽  
Alkylating Agents ◽  
Chinese Hamster ◽  
Dna Ligase ◽  
Base Excision ◽  
Dna Ligase Iii

XRCC1, the human gene that fully corrects the Chinese hamster ovary DNA repair mutant EM9, encodes a protein involved in the rejoining of DNA single-strand breaks that arise following treatment with alkylating agents or ionizing radiation. In this study, a cDNA minigene encoding oligohistidine-tagged XRCC1 was constructed to facilitate affinity purification of the recombinant protein. This construct, designated pcD2EHX, fully corrected the EM9 phenotype of high sister chromatid exchange, indicating that the histidine tag was not detrimental to XRCC1 activity. Affinity chromatography of extract from EM9 cells transfected with pcD2EHX resulted in the copurification of histidine-tagged XRCC1 and DNA ligase III activity. Neither XRCC1 or DNA ligase III activity was purified during affinity chromatography of extract from EM9 cells transfected with pcD2EX, a cDNA minigene that encodes untagged XRCC1, or extract from wild-type AA8 or untransfected EM9 cells. The copurification of DNA ligase III activity with histidine-tagged XRCC1 suggests that the two proteins are present in the cell as a complex. Furthermore, DNA ligase III activity was present at lower levels in EM9 cells than in AA8 cells and was returned to normal levels in EM9 cells transfected with pcD2EHX or pcD2EX. These findings indicate that XRCC1 is required for normal levels of DNA ligase III activity, and they implicate a major role for this DNA ligase in DNA base excision repair in mammalian cells.

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