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Characterization of an ATP-dependent type I DNA ligase from Arabidopsis thaliana

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Abstract

Here we report the purification and biochemical characterization of recombinant Arabidopsis thaliana DNA ligase I. We show that this ligase requires ATP as a source for adenylation. The calculated K m [ATP] for ligation is 3 μM. This enzyme is able to ligate nicks in oligo(dT)/poly(dA) and oligo(rA)/poly(dT) substrates, but not in oligo(dT)/poly(rA) substrates. Double-stranded DNAs with cohesive or blunt ends are also good substrates for the ligase. These biochemical features of the purified enzyme show the characteristics typical of a type I DNA ligase. Furthermore, this DNA ligase is able to perform the reverse reaction (relaxation of supercoiled DNA) in an AMP-dependent and PPi-stimulated manner.

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Author notes

  1. Alicja Ziemienowicz

    Present address: Plant Protection and Biotechnology Laboratory, Dept. of Biotechnology, Intercollegiate Faculty of Biotechnology, University of Gdansk, ul. Kladki 24, 80-822, Gdansk, Poland

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  1. Friedrich Miescher-Institute, P.O. Box 2543, 4002, Basel, Switzerland

    You-Qiang Wu & Barbara Hohn

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Wu, YQ., Hohn, B. & Ziemienowicz, A. Characterization of an ATP-dependent type I DNA ligase from Arabidopsis thaliana. Plant Mol Biol 46, 161–170 (2001). https://doi.org/10.1023/A:1010679901911

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