Effect of Mg(II) binding on the structure and activity of Escherichia coli DNA topoisomerase I

Chang Xi Zhu, Camille J. Roche, Yuk Ching Tse-Dinh

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Escherichia coli DNA topoisomerase I requires Mg(II) as a cofactor for the relaxation of negatively supercoiled DNA. Mg(II) binding to the enzyme was shown by fluorescence spectroscopy to affect the tertiary structure of the enzyme. Addition of 2 mM MgCl2 resulted in a 30% decrease in the maximum emission of tryptophan fluorescence of the enzyme. These Mg(II)-induced changes in fluorescence properties were reversible by the addition of EDTA and not obtained with other divalent cations. After incubation with Mg(II) and dialysis, inductively coupled plasma (ICP) analysis showed that each enzyme molecule could form a complex with 1-2 Mg(II) bound to each enzyme molecule. Such Mg(II)·enzyme complexes were found to be active in the relaxation of negatively supercoiled DNA in the absence of additional Mg(II). Results from ICP analysis after equilibrium dialysis and relaxation assays with limiting Mg(II) concentrations indicated that both Mg(II) binding sites had to be occupied for the enzyme to catalyze relaxation of negatively supercoiled DNA.

Original languageEnglish (US)
Pages (from-to)16206-16210
Number of pages5
JournalJournal of Biological Chemistry
Volume272
Issue number26
DOIs
StatePublished - Jun 27 1997
Externally publishedYes

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Type I DNA Topoisomerase
Escherichia coli
Superhelical DNA
Enzymes
Dialysis
Inductively coupled plasma
Fluorescence
Magnesium Chloride
Molecules
Fluorescence Spectrometry
Divalent Cations
Fluorescence spectroscopy
Edetic Acid
Tryptophan
Assays
Binding Sites

ASJC Scopus subject areas

  • Biochemistry

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Effect of Mg(II) binding on the structure and activity of Escherichia coli DNA topoisomerase I. / Zhu, Chang Xi; Roche, Camille J.; Tse-Dinh, Yuk Ching.

In: Journal of Biological Chemistry, Vol. 272, No. 26, 27.06.1997, p. 16206-16210.

Research output: Contribution to journalArticle

Zhu, Chang Xi ; Roche, Camille J. ; Tse-Dinh, Yuk Ching. / Effect of Mg(II) binding on the structure and activity of Escherichia coli DNA topoisomerase I. In: Journal of Biological Chemistry. 1997 ; Vol. 272, No. 26. pp. 16206-16210.
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