NMR SOLUTION STRUCTURE OF A DNA DECAMER CONTAINING AN INTERSTRAND CROSS-LINK OF THE ANTITUMOR DRUG CIS-DIAMMINEDICHLOROPLATINUM(II)

A 10 base pairs double-stranded oligonucleotide with the sequence d(CC TCGCTCTC). d(GAGAG*CGAGG) containing a single interstrand cross-link resulting from chelation of the N7 position of two guanine residues on the opposite strands of DNA at the d(GC/G*C) site by a cis-diamminep latinum(II) residue was analyzed by H-1 NMR spectroscopy. All the exch angeable and nonexchangeable protons resonance lines (except some H5'- H5 '') were assigned. NOESY spectra and chemical shifts indicated that the cross-linkage of the guanines of G5 and G*6 induced extrahelicit y of C5 and C6. Moreover, several unusual proximities were observed su ch as: (i) NOE cross-peaks between the H2'-H2 '' of G5 or G*6 and the aromatic proton of their 5' neighbor C4 or A7 (ii) the absence of cro ss-peak for the steps G5-C6, C6-T7 and C5-G4 (iii) a strong NOESY con nectivity between H8(G5) and H2(A7). All these data allowed us to des cribe the head to tail arrangement of the two crosslinked guanines as well as their stacking with flanking neighbor nucleotides (G5 with T7 .A7 base pair and G6 with C4.G4 base pair). Using all the NOESY and T OCSY data (208 constraints), we have obtained a solution structure of the cross-linked duplex by using the NMR-constrained molecular mechani cs program JUMNA. The reversal position of the two cross-linked guanin es placed the cis-diammineplatinum(II) residue in the minor groove. Th e stacking of the two cross-linked guanines with the surrounding bases induced a bend of 40 degrees toward the minor groove. The locally lef t-helix formation, the extrusion of the cytosines and the stacking of the platinated guanines led to an unwinding of 76 degrees. This value is in good agreement with the values deduced from gel electrophoresis experiments.