Twin hydroxymethyluracil-A base pair steps define the binding site for the DNA-bending protein TF1

Anne Grove, Marxa L. Figueiredo, Aldo Galeone, Luciano Mayol, E. Peter Geiduschek

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

The DNA-bending protein TF1 is the Bacillus subtilis bacteriophage SPO1- encoded homolog of the bacterial HU proteins and the Escherichia coli integration host factor. We recently proposed that TF1, which binds with high affinity (K(d) was ~3 nM) to preferred sites within the hydroxymethyluracil (hmU)-containing phage genome, identifies its binding sites based on sequence-dependent DNA flexibility. Here, we show that two hmU-A base pair steps coinciding with two previously proposed sites of DNA distortion are critical for complex formation. The affinity of TF1 is reduced 10-fold when both of these hmU-A base pair steps are replaced with A-hmU, G-C, or C-G steps; only modest changes in affinity result when substitutions are made at other base pairs of the TF1 binding site. Replacement of all hmU residues with thymine decreases the affinity of TF1 greatly; remarkably, the high affinity is restored when the two hmU-A base pair steps corresponding to previously suggested sites of distortion are reintroduced into otherwise T- containing DNA. T-DNA constructs with 3-base bulges spaced apart by 9 base pairs of duplex also generate nM affinity of TF1. We suggest that twin hmU-A base pair steps located at the proposed sites of distortion are key to target site selection by TF1 and that recognition is based largely, if not entirely, on sequence-dependent DNA flexibility.

Original languageEnglish (US)
Pages (from-to)13084-13087
Number of pages4
JournalJournal of Biological Chemistry
Volume272
Issue number20
DOIs
StatePublished - May 16 1997
Externally publishedYes

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Base Pairing
Binding Sites
DNA
Proteins
Bacteriophages
Integration Host Factors
Bacillus Phages
Site selection
Thymine
Bacterial Proteins
Escherichia coli Proteins
Bacilli
5-hydroxymethyluracil
Bacillus subtilis
Substitution reactions
Genes
Genome
Escherichia coli

ASJC Scopus subject areas

  • Biochemistry

Cite this

Twin hydroxymethyluracil-A base pair steps define the binding site for the DNA-bending protein TF1. / Grove, Anne; Figueiredo, Marxa L.; Galeone, Aldo; Mayol, Luciano; Geiduschek, E. Peter.

In: Journal of Biological Chemistry, Vol. 272, No. 20, 16.05.1997, p. 13084-13087.

Research output: Contribution to journalArticle

Grove, Anne ; Figueiredo, Marxa L. ; Galeone, Aldo ; Mayol, Luciano ; Geiduschek, E. Peter. / Twin hydroxymethyluracil-A base pair steps define the binding site for the DNA-bending protein TF1. In: Journal of Biological Chemistry. 1997 ; Vol. 272, No. 20. pp. 13084-13087.
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