CHARACTERIZATION OF NONCOVALENT COMPLEXES FORMED BETWEEN MINOR-GROOVEBINDING-MOLECULES AND DUPLEX DNA BY ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY

The noncovalent complex formed in solution between minor groove bindin g molecules and an oligonucleotide duplex was investigated by electros pray ionization-mass spectrometry (ESI-MS). The oligonucleotide duplex formed between two sequence-specific 14-base pair oligonucleotides wa s observed intact by ESI-MS and in relatively high abundance compared to the individual single-stranded components. Only sequence-specific A :B duplexes were observed, with no evidence of random nonspecific aggr egation (i.e., A:A or B:B) occurring under the conditions utilized. Du e to the different molecular weights of the two 14-base Fair oligonucl eotides, unambiguous determination of each oligonucleotide and the seq uence-specific duplex was confirmed through their detection at unique mass-to-charge ratios. The noncovalent complexes formed between the se lf-complementary 5'-dCGCAAA7TTTGCG-3' oligonucleotide and three minor groove binding molecules (distamycin A, pentamidine, and Hoechst 33258 ) were also observed. Variation of several electrospray ionization int erface parameters as well as collision-induced dissociation methods we re utilized to characterize the nature and stability of the noncovalen t complexes. The noncovalent complexes upon collisional activation dis sociated into single-stranded oligonucleotides and single-stranded oli gonucleotides associated with a minor groove binding molecule. ESI-MS shows potential for the study of small molecule-oligonucleotide duplex interactions and determination of small molecule binding stoichiometr y.