'Footprinting' proteins on DNA with peroxonitrous acid

Peter A. King, Elizabeth Jamison, Daniel Strahs, Vernon E. Anderson, Michael D. Brenowitz

Research output: Contribution to journalArticle

90 Citations (Scopus)

Abstract

The peroxonitrite anion (ONOO-) is a stable species in alkaline solution that quickly generates a strong oxidant at neutral pH. A convenient procedure for the preparation of ONOOK has been developed based on the procedure of Keith & Powell [(1969) J. Chem. Soc. A, 90], which when added to a sample of duplex DNA buffered at neutral pH rapidly generates a strong oxidant capable of nonspecifically cleaving the DNA present. We show that this solution containing ONOOK can be used to hydroxyl radical footprint the binding the cl-repressor (el) of phage λ with the right operator, Op. In addition, we show that the individual-site binding isotherms determined by quantitative DNase I, Fe-EDTA and ONOOK footprinting are identical within experimental error. The identical isotherms obtained with the three different reagents with greatly differing sampling times indicates that the sampling time of the footprinting probe need not be short relative to the kinetic dissociation constants that govern protein-DNA interactions.

Original languageEnglish (US)
Pages (from-to)2473-2478
Number of pages6
JournalNucleic Acids Research
Volume21
Issue number10
StatePublished - May 25 1993

Fingerprint

Protein Footprinting
DNA
Proteins
Protein
Oxidants
Acids
Isotherms
Sampling
Preparation
Bacteriophages
Peroxynitrous Acid
Probe
Kinetics
Deoxyribonuclease I
Ethylenediaminetetraacetic acid
Binding sites
Edetic Acid
Hydroxyl Radical
Anions
Negative ions

ASJC Scopus subject areas

  • Genetics
  • Statistics, Probability and Uncertainty
  • Applied Mathematics
  • Health, Toxicology and Mutagenesis
  • Toxicology
  • Genetics(clinical)

Cite this

King, P. A., Jamison, E., Strahs, D., Anderson, V. E., & Brenowitz, M. D. (1993). 'Footprinting' proteins on DNA with peroxonitrous acid. Nucleic Acids Research, 21(10), 2473-2478.

'Footprinting' proteins on DNA with peroxonitrous acid. / King, Peter A.; Jamison, Elizabeth; Strahs, Daniel; Anderson, Vernon E.; Brenowitz, Michael D.

In: Nucleic Acids Research, Vol. 21, No. 10, 25.05.1993, p. 2473-2478.

Research output: Contribution to journalArticle

King, PA, Jamison, E, Strahs, D, Anderson, VE & Brenowitz, MD 1993, ''Footprinting' proteins on DNA with peroxonitrous acid', Nucleic Acids Research, vol. 21, no. 10, pp. 2473-2478.
King PA, Jamison E, Strahs D, Anderson VE, Brenowitz MD. 'Footprinting' proteins on DNA with peroxonitrous acid. Nucleic Acids Research. 1993 May 25;21(10):2473-2478.
King, Peter A. ; Jamison, Elizabeth ; Strahs, Daniel ; Anderson, Vernon E. ; Brenowitz, Michael D. / 'Footprinting' proteins on DNA with peroxonitrous acid. In: Nucleic Acids Research. 1993 ; Vol. 21, No. 10. pp. 2473-2478.
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