Determination of affinity, stoichiometry and sequence selectivity of minor groove binder complexes with double-stranded oligodeoxynucleotides by electrospray ionization mass spectrometry
mass spectrometry; DNA; minor groove; noncovalent interaction; binding constant; dissociation constant; stoichiometry
Abstract :
[en] Electrospray mass spectrometry was evaluated regarding the reliability of the determination of the stoichiometries and equilibrium association constants from single spectra. Complexes between minor groove binders (Hoechst 33258, Hoechst 33342, DAPI, netropsin and berenil) and 12mer oligonucleotide duplexes with a central sequence (A/T)(4) flanked by G/C base pairs were chosen as model systems. To validate the electrospray ionization mass spectrometry (ESI-MS) method, comparisons were made with circular dichroism and fluorescence spectroscopy measurements. ESI-MS allowed the detection of minor (2 drug + DNA) species for Hoechst 33258, Hoechst 33342, DAPI and berenil with duplex d(GGGG(A/T)(4)GGGG). d(CCCC(A/T)(4)CCCC), which were undetectable with the other techniques. Assuming that the duplexes and the complexes have the same electrospray response factors, the equilbrium association constants of the 1:1 and 2:1 complexes were determined by ESI-MS, and the values show a good quantitative agreement with fluorescence determined constants for Hoechst 33258 and Hoechst 33342. It is also shown that ESI-MS can quickly give reliable information on the A/T sequence selectivity of a drug: the signal of a complex is directly related to the affinity of the drug for that particular duplex. The potential of ESI-MS as a qualitative and quantitative affinity screening method is emphasized.
Rosu, Frédéric ; Université de Liège - ULiège > Chimie physique, spectrométrie de masse
Gabelica, Valérie ; Université de Liège - ULiège > Chimie physique, spectrométrie de masse
Houssier, Claude ; Université de Liège - ULiège > Services généraux (Faculté des sciences) > Relations académiques et scientifiques (Sciences)
De Pauw, Edwin ; Université de Liège - ULiège > Chimie physique, spectrométrie de masse
Language :
English
Title :
Determination of affinity, stoichiometry and sequence selectivity of minor groove binder complexes with double-stranded oligodeoxynucleotides by electrospray ionization mass spectrometry
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