LONG-RANGE DISTANCE CONSTRAINTS IN PLATINATED NUCLEOTIDES - STRUCTUREDETERMINATION OF THE 5'-ORIENTATIONAL ISOMER OF CIS-[PT(NH3)(4-AMINOTEMPO)(D(GPG))](+) FROM COMBINED PARAMAGNETIC AND DIAMAGNETIC NMR CONSTRAINTS WITH MOLECULAR MODELING

The compound cis-[Pt(NH3)(4-aminoTEMPO)ClI] (7) is a paramagnetic anal ogue of the anticancer drug cisplatin and of cis-[Pt(NH3)(C6H11NH2)Cl- 2] (1), a major metabolite of a recently developed, orally administere d derivative. The bifunctional mixed amine complex 7 and a monofunctio nal triamine complex, trans-[Pt(NH3)(2)(4-aminoTEMPO)Cl]NO3 (8), were synthesized to provide localized unpaired electron spin density for us e in NMR spectral studies of their polynucleotide adducts. Compounds 7 and 8 readily coordinate to the N(7) positions of guanosine nucleosid es, as revealed by H-1, P-31, and Pt-195 NMR spectroscopy. The NMR spe ctra were selectively broadened owing to distance-dependent relaxation from the unpaired electron localized on the nitroxyl radical of the 4 -aminoTEMPO ligand. Platination of d(GpG) by the mixed amine complex 7 afforded two orientational isomers which differed with respect to the positioning of the 4-aminoTEMPO group toward either the 3' or 5' side of the phosphodiester linkage. The purified orientational isomers wer e readily distinguished by selective broadening of the H-1 NMR resonan ces of the 3' and 5' deoxyribose rings. The minimum energy solution st ructure for the 5' orientational isomer of the platinated dinucleotide cis-[Pt(NH3)(4-aminoTEMPO)(d(GpG)}](+) (13) was determined by NMR met hods including combined diamagnetic (J coupling constants) and paramag netic (electron-H-1, P-31 distances) constraints. Moreover, with the p aramagnetic spin probe, we have been able to obtain the first observab le NMR distance constraints for determining the configuration of the z eta or alpha torsion angles in any oligonucleotide. Dynamics trajector ies (200 ps) for 13 demonstrated that only computations including para magnetic distance constraints could determine the zeta(-), alpha(-) co nformation of the phosphodiester linkage and the conformation of the 4 -aminoTEMPO ligand. These NMR data and computational methods demonstra te the utility of long-range paramagnetic distance constraints in eluc idating the NMR solution structures of DNA modified by cisplatin analo gues.