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Wilkinson, Adam, Smith, Andrew, Bullard, Desmond, Lavesa-Curto, Manuel, Sayer, Heather, Bonner, Alexandra, Hemmings, Andrew 
ORCID: https://orcid.org/0000-0003-3053-3134 and Bowater, Richard ORCID: https://orcid.org/0000-0002-2745-7807
(2005)
Analysis of ligation and DNA binding by Escherichia coli DNA ligase (LigA).
Biochimica et Biophysica Acta - Proteins and Proteomics, 1749 (1).
pp. 113-122.
ISSN 1570-9639
Abstract
NAD+-dependent DNA ligases are essential enzymes in bacteria, with the most widely studied of this class of enzymes being LigA from Escherichia coli. NAD+-dependent DNA ligases comprise several discrete structural domains, including a BRCT domain at the C-terminus that is highly-conserved in this group of proteins. The over-expression and purification of various fragments of E. coli LigA allowed the investigation of the different domains in DNA-binding and ligation by this enzyme. Compared to the full-length protein, the deletion of the BRCT domain from LigA reduced in vitro ligation activity by 3-fold and also reduced DNA binding. Using an E. coli strain harbouring a temperature-sensitive mutation of ligA, the over-expression of protein with its BRCT domain deleted enabled growth at the non-permissive temperature. In gel-mobility shift experiments, the isolated BRCT domain bound DNA in a stable manner and to a wider range of DNA molecules compared to full LigA. Thus, the BRCT domain of E. coli LigA can bind DNA, but it is not essential for DNA nick-joining activity in vitro or in vivo.
| Item Type: | Article |
|---|---|
| Faculty \ School: | Faculty of Science > School of Biological Sciences |
| UEA Research Groups: | Faculty of Science > Research Groups > Plant Sciences Faculty of Science > Research Groups > Molecular Microbiology Faculty of Science > Research Groups > Chemistry of Life Processes Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry Faculty of Science > Research Groups > Biophysical Chemistry (former - to 2017) |
| Related URLs: | |
| Depositing User: | EPrints Services |
| Date Deposited: | 01 Oct 2010 13:37 |
| Last Modified: | 24 Sep 2024 09:54 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/1014 |
| DOI: | 10.1016/j.bbapap.2005.03.003 |
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