Crystal structure and nonhomologous end-joining function of the ligase component of Mycobacterium DNA ligase D Journal Article
| Authors: | Akey, D.; Martins, A.; Aniukwu, J.; Glickman, M. S.; Shuman, S.; Berger, J. M. |
| Article Title: | Crystal structure and nonhomologous end-joining function of the ligase component of Mycobacterium DNA ligase D |
| Abstract: | DNA ligase D (LigD) is a large polyfunctional enzyme involved in nonhomologous end-joining (NHEJ) in mycobacteria. LigD consists of a C-terminal ATP-dependent ligase domain fused to upstream polymerase and phosphoesterase modules. Here we report the 2.4Å crystal structure of the ligase domain of Mycobacterium LigD, captured as the covalent ligase-AMP intermediate with a divalent metal in the active site. A chloride anion on the protein surface coordinated by the ribose 3′-OH and caged by arginine and lysine side chains is a putative mimetic of the 5′-phosphate at a DNA nick. Structure-guided mutational analysis revealed distinct requirements for the adenylylation and end-sealing reactions catalyzed by LigD. We found that amutation of Mycobacterium LigD that ablates only ligase activity results in decreased fidelity of NHEJ in vivo and a strong bias of mutagenic events toward deletions instead of insertions at the sealed DNA ends. This phenotype contrasts with the increased fidelity of double-strand break repair in ΔligD cells or in a strain in which only the polymerase function of LigD is defective. We surmise that the signature error-prone quality of bacterial NHEJ in vivo arises from a dynamic balance between the end-remodeling and end-sealing steps. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc. |
| Keywords: | controlled study; unclassified drug; gene mutation; gene deletion; mutation; nonhuman; protein conformation; phenotype; dna repair; enzyme activity; bacteria (microorganisms); dna strand breakage; mycobacterium tuberculosis; dna; double stranded dna; amino acid sequence; molecular sequence data; base sequence; crystal structure; models, molecular; protein structure, tertiary; crystallization; structure analysis; polydeoxyribonucleotide synthase; nick end labeling; enzyme structure; biochemistry; mycobacterium; corynebacterineae; phosphate; enzymes; lysine; arginine; dna ligases; adenosine phosphate; adenylation; cells; bacteria; bacterial enzyme; dna ligase d; ribose; anion; divalent metal; ligase domain; polyfunctional enzyme; 5' phospate |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 281 |
| Issue: | 19 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2006-05-12 |
| Start Page: | 13412 |
| End Page: | 13423 |
| Language: | English |
| DOI: | 10.1074/jbc.M513550200 |
| PUBMED: | 16476729 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | --- - "Cited By (since 1996): 25" - "Export Date: 4 June 2012" - "CODEN: JBCHA" - "Source: Scopus" |
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