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Issue 11, 2011
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Minor groove recognition is important for the transcription factor PhoB: a surface plasmon resonance study

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Abstract

The two-component regulatory system PhoR/PhoB induces the expression of several genes in response to phosphate starvation in Escherichia coli. In order to quantify these proteinDNA interactions and to study the time-resolved dynamics of the binding mechanism, the specific recognition of different oligonucleotide duplexes by the DNA-binding domain of PhoB (PhoBDBD) was analyzed using surface plasmon resonance. In addition the two point mutants PhoBDBDD196A and PhoBDBDR219A were obtained and the DNA recognition in comparison to the wildtype PhoBDBD was investigated. Aspartic acid 196 and arginine 219 mediate specific minor groove interactions. All results reveal that at high PhoBDBD-concentrations all recognition sequences of the pho box are occupied. Decreasing the protein amount results in a mixture of free oligonucleotides and DNA molecules occupied by two WT-PhoBDBD. Moreover, the SPR results indicate that both binding site segments, the TGTCA-motif and the A/T-rich minor groove, are essential for the binding process. A comparison of different regulons additionally proved the dependency of the recognition process on the base composition of the minor groove.

Graphical abstract: Minor groove recognition is important for the transcription factor PhoB: a surface plasmon resonance study

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Publication details

The article was received on 05 Jul 2011, accepted on 19 Aug 2011 and first published on 12 Sep 2011


Article type: Paper
DOI: 10.1039/C1MB05281K
Citation: Mol. BioSyst., 2011,7, 3132-3142
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    Minor groove recognition is important for the transcription factor PhoB: a surface plasmon resonance study

    M. Ritzefeld, K. Wollschläger, G. Niemann, D. Anselmetti and N. Sewald, Mol. BioSyst., 2011, 7, 3132
    DOI: 10.1039/C1MB05281K

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