Chemical Biology of Endogenous DNA Damage by Lipid and DNA Peroxidation Products
Life has evolved on this planet from the reducing environment of the initial world to the oxidizing one of today. Molecular oxygen (O2) comprises 20% of the air we breathe and we use it to generate the bulk of the ATP we synthesize via mitochondrial respiration. However, this high O2 content makes us sensitive to spontaneous oxidation of cellular constituents. This is particularly important for the polyunsaturated fatty acid components of phospholipids and the sugar or base components of DNA. Lipid or DNA peroxidation products, malondialdehyde or base propenals, exert pathophysiological effects that can lead to mutation or cell death. Our laboratory has been particularly interested in the ability of these products to react with DNA to form adducts that induce mutations because it is likely that these mutations contribute to the induction of cancer. We have identified the major adducts produced by malondialdehyde and base propenal and used a combination of organic synthesis and molecular biology to build viral genomes containing the adducts at defined positions. By transfecting these vectors into mammalian cells and monitoring their fate during DNA replication, we are able to determine the types of mutations they induce and the pathways by which they are repaired. This has provided us with a comprehensive picture of the consequences of DNA damage by lipid and DNA peroxidation products.
We also are investigating the molecular basis for the induction of these mutations using a variety of kinetic and structural studies of complexes of adduct-containing template-primers complexed to purified DNA polymerases. This has given us a high resolution picture of the etiology of mammalian mutations. In addition to studying the mechanisms of mutagenesis and repair of specific DNA lesions, we are developing high sensitivity, mass spectrometry-based methods for their quantification in genomic DNA and biological fluids. Our goal is to measure the basal levels of these adducts in people and to determine the factors that increase or decrease them. By applying these methods in population-based studies, we hope to be able to understand the factors that contribute to cancer etiology by increasing or decreasing the levels of endogenous DNA damage.
L.A. VanderVeen, T.M. Harris, L. Jen-Jacobson, and L.J. Marnett “Formation of DNA-Protein Cross-Links Between g-Hydroxypropanodeoxyguanosine and EcoRI,” Chem.Res.Toxicol., 21, 1733-1738 (2008)
J. Szekely, C.J. Rizzo, and L.J. Marnett “The Chemical Properties of Oxopropenyl Adducts of Purine and Pyrimidine Nucleosides and Their Reactivity Toward Amino Acid Crosslink Formation,” J.Amer.Chem.Soc., 130, 2195-2201 (2008)
C.G. Knutson, H. Wang, C. J. Rizzo, and L.J. Marnett “Metabolism and Elimination of the Endogenous DNA Adduct, M1dG, in the Rat,” J.Biol.Chem., 282, 36257-36264 (2007)