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Footprinting studies of DNA‐sequence recognition by nogalamycin

Footprinting studies of DNA‐sequence recognition by nogalamycin
Footprinting studies of DNA‐sequence recognition by nogalamycin

We have studied the DNA sequence binding preference of the antitumour antibiotic nogalamycin by DNase‐I footprinting using a variety of DNA fragments. The DNA fragments were obtained by cloning synthetic oligonucleotides into longer DNA fragments and were designed to contain isolated ligand‐binding sites surrounded by repetitive sequences such as (A)n· (T)n and (AT)n. Within regions of (A)n· (T)n, clear footprints are observed with low concentrations of nogalamycin (< 5 μM), with apparent binding affinities for tetranucleotide sequences which decrease in the order TGCA > AGCT = ACGT > TCGA. In contrast, within regions of (AT)n, the ligand binds best to AGCT; binding to TCGA and TGCA is no stronger than to alternating AT. Within (ATT)n, the preference is for ACGT > TCGA. Although each of these binding sites contains all four base pairs, there is no apparent consensus sequence, suggesting that the selectivity is affected by local DNA dynamic and structural effects. At higher drug concentrations (> 25 μM), nogalamycin prevents DNAse‐I cleavage of (AT)n but shows no interaction with regions of (AC)n· (GT)n. Regions of (A)n· (T)n, which are poorly cut by DNase I, show enhanced rates of cleavage in the presence of low concentrations of nogalamycin, but are protected from cleavage at higher concentrations. We suggest that this arises because drug binding to adjacent regions distorts the DNA to a structure which is more readily cut by the enzyme and which is better able to bind further ligand molecules.

0014-2956
31-36
Fox, Keith R.
9da5debc-4e45-473e-ab8c-550d1104659f
Alam, Zafir
2ca062b2-23fc-4088-8281-6b6e21059700
Fox, Keith R.
9da5debc-4e45-473e-ab8c-550d1104659f
Alam, Zafir
2ca062b2-23fc-4088-8281-6b6e21059700

Fox, Keith R. and Alam, Zafir (1992) Footprinting studies of DNA‐sequence recognition by nogalamycin. European Journal of Biochemistry, 209 (1), 31-36. (doi:10.1111/j.1432-1033.1992.tb17257.x).

Record type: Article

Abstract

We have studied the DNA sequence binding preference of the antitumour antibiotic nogalamycin by DNase‐I footprinting using a variety of DNA fragments. The DNA fragments were obtained by cloning synthetic oligonucleotides into longer DNA fragments and were designed to contain isolated ligand‐binding sites surrounded by repetitive sequences such as (A)n· (T)n and (AT)n. Within regions of (A)n· (T)n, clear footprints are observed with low concentrations of nogalamycin (< 5 μM), with apparent binding affinities for tetranucleotide sequences which decrease in the order TGCA > AGCT = ACGT > TCGA. In contrast, within regions of (AT)n, the ligand binds best to AGCT; binding to TCGA and TGCA is no stronger than to alternating AT. Within (ATT)n, the preference is for ACGT > TCGA. Although each of these binding sites contains all four base pairs, there is no apparent consensus sequence, suggesting that the selectivity is affected by local DNA dynamic and structural effects. At higher drug concentrations (> 25 μM), nogalamycin prevents DNAse‐I cleavage of (AT)n but shows no interaction with regions of (AC)n· (GT)n. Regions of (A)n· (T)n, which are poorly cut by DNase I, show enhanced rates of cleavage in the presence of low concentrations of nogalamycin, but are protected from cleavage at higher concentrations. We suggest that this arises because drug binding to adjacent regions distorts the DNA to a structure which is more readily cut by the enzyme and which is better able to bind further ligand molecules.

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Published date: October 1992

Identifiers

Local EPrints ID: 475586
URI: http://eprints.soton.ac.uk/id/eprint/475586
ISSN: 0014-2956
PURE UUID: 1f436f53-f3c6-4562-8734-d0d2cd7d06ec
ORCID for Keith R. Fox: ORCID iD orcid.org/0000-0002-2925-7315

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Date deposited: 22 Mar 2023 17:30
Last modified: 17 Mar 2024 02:34

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Contributors

Author: Keith R. Fox ORCID iD
Author: Zafir Alam

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