The Mycobacterium tuberculosis Ku C-terminus is a multi-purpose arm for binding DNA and LigD and stimulating ligation.

Autor:	Sowa DJ; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.; Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada., Warner MM; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.; Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada., Tetenych A; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.; Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada., Koechlin L; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.; Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada., Balari P; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada., Rascon Perez JP; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada., Caba C; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.; Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada., Andres SN; Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.; Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada.
Jazyk:	angličtina
Zdroj:	Nucleic acids research [Nucleic Acids Res] 2022 Oct 28; Vol. 50 (19), pp. 11040-11057.
DOI:	10.1093/nar/gkac906
Abstrakt:	Bacterial non-homologous end joining requires the ligase, LigD and Ku. Ku finds the break site, recruits LigD, and then assists LigD to seal the phosphodiester backbone. Bacterial Ku contains a core domain conserved with eukaryotes but has a unique C-terminus that can be divided into a minimal C-terminal region that is conserved and an extended C-terminal region that varies in sequence and length between species. Here, we examine the role of Mycobacterium tuberculosis Ku C-terminal variants, where we removed either the extended or entire C-terminus to investigate the effects on Ku-DNA binding, rates of Ku-stimulated ligation, and binding affinity of a direct Ku-LigD interaction. We find that the extended C-terminus limits DNA binding and identify key amino acids that contribute to this effect through alanine-scanning mutagenesis. The minimal C-terminus is sufficient to stimulate ligation of double-stranded DNA, but the Ku core domain also contributes to stimulating ligation. We further show that wildtype Ku and the Ku core domain alone directly bind both ligase and polymerase domains of LigD. Our results suggest that Ku-stimulated ligation involves direct interactions between the Ku core domain and the LigD ligase domain, in addition to the extended Ku C-terminus and the LigD polymerase domain. (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)
Databáze:	MEDLINE
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