Asymmetric structure of a three-arm DNA junction

Qiu Guo, Min Lu, M. E A Churchill, T. D. Tullius, N. R. Kallenbach

Research output: Contribution to journalArticle

Abstract

We present here experimental evidence that three-arm branched DNA molecules form an asymmetric structure in the presence of Mg2+. Electrophoretic mobility and chemical and enzymatic footprinting experiments on a three-arm branched DNA molecule formed from three 16-mer strands are described. The electrophoretic mobilities of three species of a three-arm junction in which pairs of arms are extended are found to differ in the presence of Mg2+: one combination of elongated arms migrates significantly faster than the other two. This effect is eliminated in the absence of Mg2+, leading us to suggest that the three-arm DNA junction forms an asymmetric structure due to preferential stacking of two of the arms at the junction in the presence of Mg2+. The pattern of self-protection of each 16-mer strand of the core complex exposed to Fe(II)·EDTA and DNase I scission is unique, consistent with formation of an asymmetric structure in the presence of Mg2+. We conclude that three-arm junctions resemble four-arm junctions in showing preferential stacking effects at the branch site. Comparison of the scission patterns of linear duplexes and the branched trimer by the reactive probes methidiumpropyl-EDTA·Fe(II) [MPE·Fe(II)] and Cu(I)-[o-phenanthroline]2 [(OP)2CuI] further indicates that the branch point represents a site of enhanced binding for drugs, as it does in the four-arm case. Reaction with diethyl pyrocarbonate (DEPC), a purine-specific probe sensitive to conformation, is enhanced at the branch site, consistent with loosening of base pairing or unpairing at this point. The resolving enzyme, bacteriophage T7 endonuclease I, cleaves each of the three stands of the three-arm junction one nucleotide away from the branch with roughly equal efficiency. Thus while the structure of the three-arm junction appears to reflect preferential stacking of particular arms in the presence of Mg2+, it differs in fundamental respects at the branch from that observed in four-arm structures.

Original languageEnglish (US)
Pages (from-to)10927-10934
Number of pages8
JournalBiochemistry
Volume29
Issue number49
StatePublished - 1990

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Electrophoretic mobility
Deoxyribonuclease I
DNA
Diethyl Pyrocarbonate
Bacteriophage T7
Molecules
Bacteriophages
Base Pairing
Conformations
Nucleotides
Binding Sites
Enzymes
Pharmaceutical Preparations
Experiments
purine
1,10-phenanthroline
Fe(II)-EDTA

ASJC Scopus subject areas

  • Biochemistry

Cite this

Guo, Q., Lu, M., Churchill, M. E. A., Tullius, T. D., & Kallenbach, N. R. (1990). Asymmetric structure of a three-arm DNA junction. Biochemistry, 29(49), 10927-10934.

Asymmetric structure of a three-arm DNA junction. / Guo, Qiu; Lu, Min; Churchill, M. E A; Tullius, T. D.; Kallenbach, N. R.

In: Biochemistry, Vol. 29, No. 49, 1990, p. 10927-10934.

Research output: Contribution to journalArticle

Guo, Q, Lu, M, Churchill, MEA, Tullius, TD & Kallenbach, NR 1990, 'Asymmetric structure of a three-arm DNA junction', Biochemistry, vol. 29, no. 49, pp. 10927-10934.
Guo Q, Lu M, Churchill MEA, Tullius TD, Kallenbach NR. Asymmetric structure of a three-arm DNA junction. Biochemistry. 1990;29(49):10927-10934.
Guo, Qiu ; Lu, Min ; Churchill, M. E A ; Tullius, T. D. ; Kallenbach, N. R. / Asymmetric structure of a three-arm DNA junction. In: Biochemistry. 1990 ; Vol. 29, No. 49. pp. 10927-10934.
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