MICROGONOTROPENS AND THEIR INTERACTIONS WITH DNA .3. STRUCTURAL-ANALYSIS OF THE 1/1 COMPLEX OF D(CGCAAATTTGCG)2 AND DIEN-MICROGONOTROPEN-C BY 2D NMR-SPECTROSCOPY AND RESTRAINED MOLECULAR MODELING

The solution structures of d(CGCAAATTTGCG)2 and the 1:1 complex of d(CGCAAATTTGCG)2 with dien-microgonotropen-c (5c) have been determined by 1D and 2D H-1 NMR spectroscopy and restrained molecular modeling. One hundred and two resonances for the free DNA and 196 for the DNA bound to 5c have been assigned. The 1 D (DNA imino protons) and 2D (NOESY) spectra of the 1:1 complex show that there is an asymmetric type of binding in the A+T-rich region involving five base pairs (5'-A6T7T8T9G10). The two most stable structures of the d(CGCAAATTTGCG)2:5c complex have (i) pyrrole rings A and B coplanar and in the minor groove with pyrrole ring C out of plane by approximately 70-degrees and (ii) pyrrole rings A, B, and C coplanar and in the minor groove. The amino terminal acetamide head is directed toward A6 while the carboxy terminal (dimethylamino)propyl tail is directed toward and above G10. The energy barrier between the two bound 5c conformations is 2.5 kcal/mol in favor of the structure with only two pyrrole rings in the minor groove. The dien polyamino substituent residing on the nitrogen of pyrrole ring C runs above and along the phosphate backbone, toward the major groove. The protonated terminal dimethylamine nitrogen of the (dimethylamino)propyl tail is adjacent to a negatively charged phosphodiester linkage (P11) on the minor groove side, while the protonated dien nitrogens reside on the edge of the major groove and pair with the phosphodiester linkages P8, P9, and P10. The off-rate of 5c from the 1:1 complex was found to be 1.3 +/- 0.2 s-1, corresponding to an activation energy of 17 kcal/mol. The relative positions of the DNA proton signals change as 5c binds to the DNA. This is due, in part, to the widening of the minor groove (up to 3 angstrom) in the binding site. Compound 5c binds 5-7 angstrom from the bottom of the groove and 5-6 and 4-6 angstrom distant from the (-) and (+) strands, respectively. Comparisons with the crystallographic data of the same DNA with and without distamycin were made. Molecular modeling of the free and 5c-bound DNA, based on NOE measurements, shows that there is a break in the C2v symmetry of the crystallized DNA at the A6T7 junction as it goes into aqueous solution. An increase in the helical bend of 10.6-degrees from that of the crystallized DNA was found to occur in the solution DNA while an increase of only 6.4-degrees was found for the solution DNA:5c complex relative to the crystallized dodecamer. Upon solvation, the length of the duplex increases by 0.1 angstrom/bp for both the dodecamer and the 5c-complexed dodecamer compared to the case of crystal structures of free DNA and distamycin-complexed DNA.