This project will explore the synthesis and use of multiply labeled DNA fragments that include both 15N and 13C labels. By incorporation of multiple labels we should be able to increase dramatically the amount of information that can be obtained from an NMR experiment. This multi- labeling approach will be accomplished both by incorporation of multiple- labeled monomers, and also by incorporation of more than one labeled monomer. The routes that we have already developed for preparation of the singly-labeled 15N deoxynucleosides will serve as a starting point for preparation of the multiply labeled compounds. The new labeled compounds targeted at this time are listed below: 1. [7-15N]-Hypoxanthine 2. [6,7-15N]-Adenine 3. [6,7-15N]-Deoxyadenosine 4. [3,6-15N]-Adenine 5. [3,6-15N]-Deoxyadenosine 6. [1,6,7-15N]-Deoxyadenosine 7. [2-13C]-[2,3-15N]-Deoxyguanosine 8. [2-13C]-[1,2-15N]-Deoxyguanosine 9. [2-13C]-[1,2,7-15N]-Deoxyguanosine 10. [2-13C]-[2,3-15N]-O6-Methyldeoxyguanosine 11. [2-13C]-[1,2-15N]-O6-Methyldeoxyguanosine 12. [2-13C]-[1,2,7-15N]-O6-Methyldeoxyguanosine These compounds, along with [4-15N]-deoxycytidine, will be incorporated into DNA fragments for NMR studies. The [1-15N]-, [2-15N]-, and [6-15N]- labels monitor interactions of the Watson-Crick face, while the [7-15N]- and [3-15N]-labels monitor interactions in the major and minor grooves. The [2-13C] atom will provide information about ring current effects, and will be used to detect the 15N chemical shift in molecules where there is non-exchangeable proton coupled to the 15N label. Specifically, we will define the base pairing present in O6MeG.A and O6MeG.G mismatches, and the base pairing in parallel-DNA. We will probe drug.DNA and peptide.DNA interactions, including both DNA.DNA and ligand.DNA H-bonding, as well as the exclusion of water that occurs upon drug on peptide binding. Further, in each of these systems, we will attempt to correlate the identity of the H-bond acceptor (O or N) with the chemical shift change of the donor NH group. In addition, with these multi-labeled compounds, we will also be able to monitor changes in hydration which occur at the site of the lesion, the site of ligand binding, or elsewhere in the complex.
Showing the most recent 10 out of 22 publications
