The chemistry central to the function of the DNA repair proteins apurinic/apyrimidic endonuclease 1 (Ape1) and polymerase beta (Pol beta) is the chemistry of water activated by a magnesium ion. These proteins are key constituents of the base excision repair (BER) pathway, a process that plays a critical role in preventing the cytotoxic and mutagenic effects of most spontaneous, alkylation and oxidative DNA damage. The proposed research represents a novel application of low temperature (10K) solid-state 25Mg NMR applied to DNA repair proteins and their model systems. The principal aim for the proposed research is to establish a relationship between 25Mg magnetic resonance parameters (quadrupole coupling constants, shielding tensors and their relative orientations) and the structure/function relationships for known Mg-DNA repair proteins and extrapolating those relationships to other Mg-dependent DNA repair proteins where the role of the metal is less defined. These experiments are performed in concert with ab initio electronic structure calculations. The combination of these two methods provides a firm basis for the """"""""chemical"""""""" understanding of the role these metals play in their respective proteins. Additionally, triple resonance experiments (1H, [15N or 31P], 25Mg) are proposed to facilitate the identity of neighboring ligands and the determination of selective distances between the magnesium and the substrates. The results of the entirety of these experiments are critical for the mechanistic understanding of the role the metal plays in these critical DNA repair proteins.

Agency
National Institute of Health (NIH)
Institute
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Type
Research Project (R01)
Project #
5R01EB003893-04
Application #
7645052
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Mclaughlin, Alan Charles
Project Start
2006-08-17
Project End
2012-06-30
Budget Start
2009-07-01
Budget End
2012-06-30
Support Year
4
Fiscal Year
2009
Total Cost
$585,810
Indirect Cost
Name
Battelle Pacific Northwest Laboratories
Department
Type
DUNS #
032987476
City
Richland
State
WA
Country
United States
Zip Code
99352
Lipton, Andrew S; Heck, Robert W; Hernick, Marcy et al. (2008) Residue ionization in LpxC directly observed by 67Zn NMR spectroscopy. J Am Chem Soc 130:12671-9
Lipton, A S; Heck, R W; Primak, S et al. (2008) Characterization of Mg2+ binding to the DNA repair protein apurinic/apyrimidic endonuclease 1 via solid-state 25Mg NMR spectroscopy. J Am Chem Soc 130:9332-41
Lipton, Andrew S; Heck, Robert W; Staeheli, Greg R et al. (2008) A QM/MM approach to interpreting 67Zn solid-state NMR data in zinc proteins. J Am Chem Soc 130:6224-30