[Intracomplex alkylation of octanucleotide pd(TGTTTGGC) by a 5'-(4-N-methyl-N-(2-chloroethyl)-amino)benzylphosphamide derivative of heptanucleotide pd(CCAAACA). Preparation of a covalent adduct and study of its spatial structure in an aqueous solution by two-dimensional (1)H-NMR spectroscopy].

Covalent adduct--the product of intracomplex alkylation at N-3-position of dC-8-nucleoside residue of target octanucleotide pd[TGTTTGGC] was completely synthesized by means of 4-[N-methyl-N-(2-chloroethyl)amino]benzyl-5'-phosphamido derivative of heptanucleotide pd[CCAAACA]. Its melting temperature was shown to be 70 degrees C. Tm did not depend on covalent adduct concentration and was by 40 degrees C higher than that for unmodified duplex pd[TGTTTGGC].pd[CCAAACA] at concentration of 0.5 x 10(-4) M. The spatial structure of the covalent adduct in aqueous solution was investigated by two-dimensional 3H-NMR spectroscopy. The assignment of oligonucleotide protons as well as protons of a modifying group was carried out using COSY, COSY-DQF and NOESY experiments. Conformational analysis of proton-proton coupling constants for H1', H2'a, H2'b and H3' protons showed the sugar residues to be in 2'-endo conformation. Analysis of NOE connectivities observed between the protons of the alkylating group and oligonucleotide protons yielded conclusion, regarding the 4-[N-methyl-N-(2-chloroethyl)amino]benzylamido 5'-residue being localized in the region of the lacked nucleoside residue of the heptanucleotide chain about 5 A apart from the dC-1 residue and from cytosine base of the alkylated dC-8 residue.