Development of Novel Spin-labeled Nucleosides which detect base-pairing

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We designed novel spin-labeled nucleosides, in which a spin source, an N-tert-butylaminoxyl radical, was directly attached to nucleobase. 2-N-tert-Butylaminoxyladenosine, 2-N-tert-butylaminoxylinosine, and 8-N-tert-butylaminoxylinosine, and 5-N-tert-butylaminoxylcytidine derivative were synthesized and thier stabilities and properties were studied by using electron peramagnetic resonance(EPR). It was of interest to investigate if EPR spectra of N-tert-butylaminoxylnucleosides changed upon formation of hydrogen bonding with complementary base. EPR spectra of O-silylated-5-N-tert-butylaminoxylcytidine in CH_2Cl_2 changed upon addition of O-silylated guanosine, which might form hydrogen bonds with the aminoxyl cytidine. 2-N-tert-Butylaminoxyladenosine was introduced into oligoncleotide and its EPR spectra were measured. The EPR spectrum of the obtained oligonucleotide showed three lines in phosphate buffer. The high line disappeared upon duplex formation with the complementary strand to show that the EPR spectra might give useful information on the structure of oligonucleotide. EPR spectra of 2-N-tert-butylaminoxylinosine showed pH dependency due to structural change of purine moiety