EXAFS Study of Metal-Stabilized DNA Quadruplexes.

 

Richard H. Shafer1, Ivan V. Smirnov1, Ingrid J. Pickering2

1Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94143-0446 USA; 2Stanford Synchrotron Radiation Laboratory, SLAC, Stanford, CA 94309-0210 USA

 

Guanine quadruplexes are unique among unusual DNA structures in that they are stabilized by site-specific binding of metal ions. Structural details concerning the metal-DNA interaction, however, are difficult to obtain in NMR studies on quadruplex structures and there have been very few X-ray crystallography results on these systems. Thus in order to characterize this unusual metal-DNA interaction, and in particular to locate the metal binding site, we have initiated EXAFS (Extended X-ray Absorption Fine Structure) studies on several small DNA quadruplexes stabilized by Pb2+ or Sr2+. These divalent cations bind very tightly to the DNA sequence d(GTTGGTGTGGTTGG), resulting in a unimolecular folded quadruplex based on two guanine quartets, with one metal ion bound per structure. In each of the different metal-stabilized structures, we have obtained EXAFS data demonstrating a high degree of structural order surrounding the metal ion, with the Fourier transform of the EXAFS function showing one sharp maximum and one broad maximum. Analysis of the EXAFS function results in three shells of atoms at 2.65, 3,65 and 3.95 Å from the metal center. Similar results were obtained for a closely related DNA oligonucleotide as well as a model compound of known structure. These results locate the divalent cation in the region between the two quartets, coordinating to the four carbonyl oxygens in each quartet above and below the metal ion, in contrast to the proposed binding sites for monovalent cations.