Department of Biochemistry

Michael R. Gunther, Ph.D.

Ph.D. - Colorado State, 1992

Associate Professor
Department of Biochemistry
West Virginia University
Robert C. Byrd Health Sciences Center
P.O. Box 9142
Morgantown, WV 26506
Phone: (304) 293-0714
Fax: (304) 293-6846
Email: mgunther@hsc.wvu.edu

Research:

The oxidation of proteins, DNA, and lipids to form damaged and frequently toxic intermediates has been implicated in a wide variety of diseases, including cancer, coronary artery disease, aging, neurodegeneration, and arthritis. While the majority of intentional biological oxidations, particularly those involved in energy production, are two-electron processes, many damaging oxidations occur by one-electron mechanisms, resulting in the formation of free radicals, which are by definition molecules or atoms with unpaired electrons. Free radicals are typically very reactive, particularly with molecular oxygen, and their reactions frequently result in the formation of additional free radicals, resulting in chain reactions that rapidly propagate damage from an initial oxidized site. Uncontrolled radical chain reactions in cells or tissues frequently result in the death of the affected system.

The research being conducted in our laboratory is directed toward understanding the mechanism by which mutant forms of the enzyme superoxide dismutase (SOD) cause amyotrophic lateral sclerosis (ALS, also known as Lou Gehrig’s disease). Copper, zinc-SOD is a ubiquitous enzyme that detoxifies superoxide, the one-electron reduction product of molecular oxygen, through disproportionation, to form water and hydrogen peroxide. About ¼ of the inherited cases of ALS (and therefore about 5% of all cases) are caused by inheritance of a mutant gene for SOD, and the ALS-associated enzymes are the only known mutants of SOD. Transgenic mice that express any of the mutant human SODs but not the wild-type protein develop ALS-like disease without a decrease in overall SOD activity, indicating that the mutant enzymes gain a toxic function. Because of its role as a free radical detoxifying enzyme, many hypotheses about the mechanism by which the mutant SODs cause ALS focus on free radical reactions that are catalyzed (or not prevented) by the mutant enzymes. Initial experiments indicated that the mutant enzymes react with hydrogen peroxide to form the very toxic and reactive hydroxyl radical to a greatly increased extent, but recent work has questioned those results. Since the inheritance of a mutant SOD is the only cause of ALS that has been definitively identified, it is hoped that understanding the mechanism through which the mutations cause the disease will provide more general information about the induction of the disease in general.

My general scientific interests are in the area of the formation and reactions of protein-centered free radicals, particularly with respect to disease induction. Since transition metals frequently contain unpaired electrons, they have a tendency to initiate free radical reactions, particularly when reacting with materials such as hydrogen peroxide, which is a byproduct of oxidative metabolism. A number of xenobiotics also cause peroxide formation in vivo. The radical-forming reactions between metalloproteins and peroxides are therefore both physiologically feasible and also potentially quite relevant to disease induction.

The hypothesis being examined in our lab is that the reaction SOD with peroxides results in the formation of SOD-centered free radicals at different, more toxic sites on the mutant enzymes than on the wild-type protein. This work is being done using the purified proteins, produced by expression in yeast cells. The primary tool used to characterize free radicals is electron paramagnetic resonance (EPR) spectroscopy, also known as electron spin resonance (ESR). Using the spin-trapping technique, transient free radicals can be stabilized by their reactions with chemicals known as "spin traps" which react with free radicals to form more stable radicals, the characterization of which allows the structural determination of the original radical.

Selected References:

• Gunther, MR, Hanna, PM, Mason, RP, and Cohen, MS.  Hydroxyl radical formation from copper (I) and hydrogen peroxide: A spin trapping study.  Arch. Biochem. Biophys. 316, 515-522, 1995
• Gunther, MR, Kelman, DJ, Corbett, JT, and Mason, RP. Self-peroxidation of metmyoglobin results in formation of an oxygen-reactive tryptophan-centered radical.  J. Biol. Chem. 270, 16075-16081, 1995
• Barr, DP, Gunther, MR, Deterding, LJ, Tomer, KB, and Mason, RP.  ESR Spin-trapping of a protein-derived tyrosyl radical from the reaction of cytochrome c with hydrogen peroxide.  J. Biol. Chem. 271, 15498-15503, 1996
• DeGray, JA, Gunther, MR, Tschirret-Guth, R, Ortiz de Montellano, PR, and Mason, RP.   Peroxidation of a specific tryptophan of metmyoglobin by hydrogen peroxide. J. Biol. Chem. 272, 2359-2362, 1997
• Gunther, MR, Hsi, LC, Curtis, JF, Gierse, JK, Marnett, LJ, Eling, TE, and Mason, RP.   Nitric oxide trapping of the tyrosyl radical of Prostaglandin H Synthase-2 leads to tyrosine iminoxyl radical and nitrotyrosine formation.  J. Biol. Chem. 272, 17086-17090, 1997
• Jiang, JJ, Jordan, SJ, Barr, DP, Gunther, MR, Maeda, H, and Mason, RP.  In vivo production of nitric oxide in rats following administration of hydroxyurea.  Mol. Pharm 52, 1081-1086, 1997
• Gunther, MR, Tschirret-Guth, RA, Witkowska, HE, Fann, YC, Barr, DP, Ortiz de Montellano, PR, and Mason, RP.  Site-specific spin trapping of tyrosine radicals in the oxidation of metmyoglobin by hydrogen peroxide.  Biochem. J., 330, 1293-1299, 1998
• Goodwin, DC, Gunther, MR, Hsi, LC, Crews, BC, Eling, TE, Mason, RP, and Marnett, LJ.  Nitric oxide trapping of tyrosyl radicals generated during prostaglandin endoperoxide synthase turnover:  detection of the radical derivative of tyrosine 385.  J. Biol. Chem., 273, 8903-8909, 1998
• Singh, RJ, Karoui, H, Gunther, MR, Beckman, JS, Mason, RP, and Kalyanaraman, B.  Reexamination of the mechanism of hydroxyl radical adducts formed from the reaction between familial amyotrophic lateral sclerosis (FALS)-associated CuZnSOD mutants and H2O2.  Proc. Natl. Acad. Sci. U.S.A., 95, 6675-6680, 1998
• Chen, YR, Gunther, MR, and Mason, RP.  An ESR spin-trapping investigation of the reaction of mitochondrial cytochrome c oxidase with H2O2.  J. Biol. Chem., 275, 3308-3314, 1999
• Gunther, MR, Sampath, V, and Caughey, WS.  Potential roles of myoglobin autoxidation in myocardial ischemia-reperfusion injury.  Free Radic. Biol. Med., 26, 1388-1395, 1999
• Chen, YR, Sturgeon, BE, Gunther, MR, and Mason, RP.  Electron spin resonance investigation of the cyanyl and azidyl radical formation by cytochrome c oxidase.  J. Biol. Chem. 274, 24611-24616, 1999
• Gunther, MR, Sturgeon, BE, and Mason, RP.  A long-lived tyrosyl radical from the reaction between horse metmyoglobin and hydrogen peroxide.  Free Radic. Biol. Med. 28, 709-719, 2000
• Sahlin, M., Cho, K-B, Potsch, S., Lytton, S., Huque, Y., Gunther, M.R., Sjoberg, B-M., Mason, R.P., and Graslund, A.  Peroxide adduct radicals formed in the iron/oxygen reconstitution reaction of mutant ribonucleotide reductase R2 proteins from Escherichia coli.  J. Biol. Inorg. Chem. 7, 74-82, 2002
• Gunther, M.R., Peters, J.A., and Sivaneri, M.K.  Histidinyl radical formation in the self-peroxidation reaction of bovine copper-zinc superoxide dismutase.  J. Biol. Chem. 277, 9160-9166, 2002. 
• Shvedova, A. A., Kisin, E.R., Murray, A.R., Commineni, C., Castranova, V., Mason, R.P., Kadiska, M.B., and Gunther, M.R.  Antioxidant balance and free radical generation in vitamin E deficient mice after dermal exposure to cumene hydroperoxide.  Chem. Res. Toxicol, 15:1451-1459, 2002. 
• Gunther, M.R., Lardinois, O., Tschirret-Guth, R.A., and Ortiz de Montellano, P.R.  Tryptophan-14 is the preferred site of DBNBS spin trapping in the self-peroxidation reaction of sperm whale metmyoglobin.  Chem. Res. Toxicol. 16:652-660, 2003.
• Gunther, M.R., VanGilder, R., Fang, J., and Beattie, D.S..  Expression of a familial amyotrophic lateral sclerosis-associated mutant human superoxide dismutase in yeast leads to decreased mitochondrial electron transport.  Arch. Biochem. Biophys. 431:207-214, 2004. 
• Gunther, M.R. and Donahue, J.A.  Bicarbonate and active site zinc modulate the self-peroxidation of bovine copper-zinc superoxide dismutase.  Free Radic. Res. 41:1005-1016, 2007.
• Murray, E. Kisin, V. Castranova, C. Kommineni, M.R. Gunther, and A.A. Shvedova.  Phenol induced in vivo oxidative stress in skin:  Evidence for enhanced free radical generation, thiol oxidation, and antioxidant depletion.  Chem. Res. Toxicol. (2007) in press.