scholarly journals Peroxynitrite Mediates Active Site Tyrosine Nitration in Manganese Superoxide Dismutase. Evidence of a Role for the Carbonate Radical Anion

2010 ◽  
Vol 132 (48) ◽  
pp. 17174-17185 ◽  
Author(s):  
N. Basak Surmeli ◽  
Nadia K. Litterman ◽  
Anne-Frances Miller ◽  
John T. Groves
Keyword(s):  
Superoxide Dismutase ◽  
Active Site ◽  
Radical Anion ◽  
Manganese Superoxide Dismutase ◽  
Tyrosine Nitration ◽  
Carbonate Radical ◽  
Manganese Superoxide ◽  
Active Site Tyrosine ◽  
Carbonate Radical Anion

scholarly journals Cu,Zn-superoxide dismutase-driven free radical modifications: copper- and carbonate radical anion-initiated protein radical chemistry

2008 ◽  
Vol 417 (1) ◽  
pp. 341-353 ◽  
Author(s):  
Dario C. Ramirez ◽  
Sandra E. Gomez-Mejiba ◽  
Jean T. Corbett ◽  
Leesa J. Deterding ◽  
Kenneth B. Tomer ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Active Site ◽  
Radical Anion ◽  
Brain Homogenate ◽  
Experimental Conditions ◽  
Carbonate Radical ◽  
Carbonate Radical Anion ◽  
Protein Radicals ◽  
Mouse Brain Homogenate

The understanding of the mechanism, oxidant(s) involved and how and what protein radicals are produced during the reaction of wild-type SOD1 (Cu,Zn-superoxide dismutase) with H2O2 and their fate is incomplete, but a better understanding of the role of this reaction is needed. We have used immuno-spin trapping and MS analysis to study the protein oxidations driven by human (h) and bovine (b) SOD1 when reacting with H2O2 using HSA (human serum albumin) and mBH (mouse brain homogenate) as target models. In order to gain mechanistic information about this reaction, we considered both copper- and CO3•− (carbonate radical anion)-initiated protein oxidation. We chose experimental conditions that clearly separated SOD1-driven oxidation via CO3•− from that initiated by copper released from the SOD1 active site. In the absence of (bi)carbonate, site-specific radical-mediated fragmentation is produced by SOD1 active-site copper. In the presence of (bi)carbonate and DTPA (diethylenetriaminepenta-acetic acid) (to suppress copper chemistry), CO3•− produced distinct radical sites in both SOD1 and HSA, which caused protein aggregation without causing protein fragmentation. The CO3•− produced by the reaction of hSOD1 with H2O2 also produced distinctive DMPO (5,5-dimethylpyrroline-N-oxide) nitrone adduct-positive protein bands in the mBH. Finally, we propose a biochemical mechanism to explain CO3•− production from CO2, enhanced protein radical formation and protection by (bi)carbonate against H2O2-induced fragmentation of the SOD1 active site. Our present study is important for establishing experimental conditions for studying the molecular mechanism and targets of oxidation during the reverse reaction of SOD1 with H2O2; these results are the first step in analysing the critical targets of SOD1-driven oxidation during pathological processes such as neuroinflammation.

Single Crystal Polarized Spectroscopy of Manganese Superoxide Dismutase and Electronic Structure of the Active Site Metal Complex†

1998 ◽  
Vol 102 (23) ◽  
pp. 4668-4677 ◽  
Author(s):  
Mei M. Whittaker ◽  
Christopher A. Ekberg ◽  
Ross A. Edwards ◽  
Edward N. Baker ◽  
Geoffrey B. Jameson ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Electronic Structure ◽  
Metal Complex ◽  
Single Crystal ◽  
Active Site ◽  
Manganese Superoxide Dismutase ◽  
Polarized Spectroscopy ◽  
Manganese Superoxide

Quantitative assessment of tyrosine nitration of manganese superoxide dismutase in angiotensin II-infused rat kidney

2003 ◽  
Vol 285 (4) ◽  
pp. H1396-H1403 ◽  
Author(s):  
Wei Guo ◽  
Takeshi Adachi ◽  
Reiko Matsui ◽  
Shanqin Xu ◽  
Bingbing Jiang ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Angiotensin Ii ◽  
Western Blotting ◽  
Manganese Superoxide Dismutase ◽  
Rat Kidney ◽  
Oxidant Stress ◽  
Uv Detection ◽  
Tyrosine Nitration ◽  
Manganese Superoxide ◽  
Rat Kidneys

Hypertension caused by angiotensin II is characterized by an increase in tissue oxidant stress as evidenced by increased quantities of reactive oxygen and nitrogen species. Manganese superoxide dismutase (MnSOD) is a key mitochondrial antioxidant enzyme that is inactivated in conditions of oxidant stress by reacting with peroxynitrite to form 3-nitrotyrosine in its active site. The increase in 3-nitrotyrosine content in MnSOD in the kidney of angiotensin II-infused rats was assessed in this study by immunohistochemistry, Western blotting, immunoprecipitation, and HPLC with UV detection (HPLC-UV). MnSOD activity decreased ∼50% in angiotensin II-infused rat kidneys (24 ± 4.6 vs. 11 ± 5.2 U/mg) without a change in protein expression. Immunohistochemical staining showed 3-nitrotyrosine predominantly in distal tubules and collecting duct cells in the angiotensin II-infused rat kidneys. By two-photon microscopy, 3-nitrotyrosine colocalized with MnSOD. Total 3-nitrotyrosine content in kidney homogenates was increased in angiotensin II-infused rat kidney [3.2 ± 1.9 (sham treated) vs. 9.5 ± 2.3 ng/mg protein by HPLC-UV detection]. With tracer amounts of tyrosine-nitrated recombinant MnSOD, the most sensitive technique to detect tyrosine nitration of MnSOD was immunoprecipitation from tissue with anti-MnSOD antibody, followed by detection of 3-nitrotyrosine by Western blotting or HPLC. By HPLC, 3-nitrotyrosine content of kidney MnSOD increased 13-fold after angiotensin II infusion, representing an increase from approximately one-twentieth to one-fifth of the total 3-nitrotyrosine content in sham-treated and angiotensin II-infused rat kidney, respectively. Angiotensin II-induced hypertension is accompanied by increased tyrosine nitration of MnSOD, which, because it inactivates the enzyme, may contribute to increased oxidant stress in the kidney.

Redox Properties of Human Manganese Superoxide Dismutase and Active-Site Mutants†

Biochemistry ◽  
2001 ◽  
Vol 40 (35) ◽  
pp. 10586-10591 ◽  
Author(s):  
Vincent J.-P. Lévêque ◽  
Carrie K. Vance ◽  
Harry S. Nick ◽  
David N. Silverman
Keyword(s):  
Superoxide Dismutase ◽  
Active Site ◽  
Manganese Superoxide Dismutase ◽  
Redox Properties ◽  
Manganese Superoxide ◽  
Active Site Mutants

scholarly journals Decreases in manganese superoxide dismutase expression and activity contribute to oxidative stress in persistent pulmonary hypertension of the newborn

2012 ◽  
Vol 303 (10) ◽  
pp. L870-L879 ◽  
Author(s):  
Adeleye J. Afolayan ◽  
Annie Eis ◽  
Ru-Jeng Teng ◽  
Ivane Bakhutashvili ◽  
Sushma Kaul ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Nitric Oxide ◽  
Pulmonary Hypertension ◽  
Superoxide Dismutase ◽  
Manganese Superoxide Dismutase ◽  
Pulmonary Arteries ◽  
Tyrosine Nitration ◽  
Enos Expression ◽  
Manganese Superoxide ◽  
Expression And Activity

A rapid increase in the synthesis and release of nitric oxide (NO) facilitates the pulmonary vasodilation that occurs during birth-related transition. Alteration of this transition in persistent pulmonary hypertension of the newborn (PPHN) is associated with impaired function of endothelial nitric oxide synthase (eNOS) and an increase in oxidative stress. We investigated the hypothesis that a decrease in expression and activity of mitochondrial localized manganese superoxide dismutase (MnSOD) in pulmonary artery endothelial cells (PAEC) increases oxidative stress and impairs eNOS function in PPHN. We isolated PAEC and pulmonary arteries from fetal lambs with PPHN induced by prenatal ductus arteriosus ligation or sham ligation (control). We investigated MnSOD expression and activity, tyrosine nitration of MnSOD, and mitochondrial O2− levels in PAEC from control and PPHN lambs. We introduced exogenous MnSOD via an adenoviral vector (ad-MnSOD) transduction into PAEC and pulmonary arteries of PPHN lambs. The effect of ad-MnSOD was investigated on: mitochondrial O2− levels, MnSOD and eNOS expression and activity, intracellular hydrogen peroxide (H2O2) levels, and catalase expression in PAEC. MnSOD mRNA and protein levels and activity were decreased and MnSOD tyrosine nitration was increased in PPHN-PAEC. ad-MnSOD transduction of PPHN-PAEC increased its activity two- to threefold, decreased mitochondrial O2− levels, and increased H2O2 levels and catalase expression. ad-MnSOD transduction improved eNOS expression and function and the relaxation response of PPHN pulmonary arteries. Our observations suggest that decreased MnSOD expression and activity contribute to the endothelial dysfunction observed in PPHN.

scholarly journals Cryotrapping peroxide in the active site of human mitochondrial manganese superoxide dismutase crystals for neutron diffraction

2022 ◽  
Vol 78 (1) ◽  
Author(s):  
Jahaun Azadmanesh ◽  
William E. Lutz ◽  
Leighton Coates ◽  
Kevin L. Weiss ◽  
Gloria E. O. Borgstahl
Keyword(s):  
Superoxide Dismutase ◽  
Data Collection ◽  
Neutron Diffraction ◽  
Active Site ◽  
Active Sites ◽  
Manganese Superoxide Dismutase ◽  
Crystal Form ◽  
X Rays ◽  
Manganese Superoxide ◽  
Neutron Crystallography

Structurally identifying the enzymatic intermediates of redox proteins has been elusive due to difficulty in resolving the H atoms involved in catalysis and the susceptibility of ligand complexes to photoreduction from X-rays. Cryotrapping ligands for neutron protein crystallography combines two powerful tools that offer the advantage of directly identifying hydrogen positions in redox-enzyme intermediates without radiolytic perturbation of metal-containing active sites. However, translating cryogenic techniques from X-ray to neutron crystallography is not straightforward due to the large crystal volumes and long data-collection times. Here, methods have been developed to visualize the evasive peroxo complex of manganese superoxide dismutase (MnSOD) so that all atoms, including H atoms, could be visualized. The subsequent cryocooling and ligand-trapping methods resulted in neutron data collection to 2.30 Å resolution. The P6122 crystal form of MnSOD is challenging because it has some of the largest unit-cell dimensions (a = b = 77.8, c = 236.8 Å) ever studied using high-resolution cryo-neutron crystallography. The resulting neutron diffraction data permitted the visualization of a dioxygen species bound to the MnSOD active-site metal that was indicative of successful cryotrapping.

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