Purification, crystallization and X-ray structures of the two manganese superoxide dismutases from<i>Caenorhabditis elegans</i>

Trinh, Chi H.; Hunter, Thérèse; Stewart, Emma E.; Phillips, Simon E. V.; Hunter, Gary J.

doi:10.1107/s1744309108037056

Cited by 18 publications

(22 citation statements)

References 34 publications

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“…We generated a Drosophila SOD2 homology model using the program MUSTER (Wu and Zhang ) and the structure of the C. elegans manganese superoxide dismutase (3DC6) (Trinh et al. ) as the structural template (64% identical: Fig. a).…”

Section: Resultsmentioning

confidence: 99%

A novel DrosophilaSOD2 mutant demonstrates a role for mitochondrial ROS in neurodevelopment and disease

et al. 2012

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Reactive oxygen species (ROS) play essential roles in cell signaling, survival, and homeostasis. Aberrant ROS lead to disease and contribute to the aging process. Numerous enzymes and vigilant antioxidant pathways are required to regulate ROS for normal cellular health. Mitochondria are a major source of ROS, and mechanisms to prevent elevated ROS during oxidative phosphorylation require super oxide dismutase (SOD) activity. SOD2, also known as MnSOD, is targeted to mitochondria and is instrumental in regulating ROS by conversion of superoxides to hydrogen peroxide, which is further broken down into H2O and oxygen. Here, we describe the identification of a novel mutation within the mitochondrial SOD2 enzyme in Drosophila that results in adults with an extremely shortened life span, sensitivity to hyperoxia, and neuropathology. Additional studies demonstrate that this novel mutant, SOD2bewildered, exhibits abnormal brain morphology, suggesting a critical role for this protein in neurodevelopment. We investigated the basis of this neurodevelopmental defect and discovered an increase in aberrant axonal that could underlie the aberrant neurodevelopment and brain morphology defects. This novel allele, SOD2bewildered, provides a unique opportunity to study the effects of increased mitochondrial ROS on neural development, axonal targeting, and neural cell degeneration in vivo.

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Section: Resultsmentioning

confidence: 99%

A novel DrosophilaSOD2 mutant demonstrates a role for mitochondrial ROS in neurodevelopment and disease

et al. 2012

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“…Previously, the crystal structure studies revealed that human MnSOD is a homotetramer whicheach monomer consists of 198 amino acids and is divided into N-terminal helices and a mixed C-terminal a/b domain (Borgstahl et al 1992;Wagner et al 1993). Moreover, many site-specific mutagenesis studies indicated that several residues in C-terminal domain associate with the interactions at the active site as well as the interactions at dimer interface that shown to be important for SOD activity (Borgstahl et al 1992;Sheng et al 2013;Trinh et al 2008;Wagner et al 1993). Therefore, decrease of enzymatic activity in our case could be probably due to the disturbance of the C-terminal fused peptides at active site or dimer interface.…”

Section: Discussionmentioning

confidence: 97%

Angiopep-2-Mediated Delivery of Human Manganese Superoxide Dismutase in Brain Endothelial Cells and its Protective Effect Against Oxidative Stress

Eiamphungporn

Yainoy

Prachayasittikul

2014

Int J Pept Res Ther

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Oxidative stress has been considered as the leading cause of blood-brain barrier disruption which implicates many neurological disorders. Manganese superoxide dismutase (MnSOD or SOD2) is one of the crucial antioxidant enzymes that can provide substantial protection against oxidative damage. However, the therapeutic effect of the enzyme in such neurological diseases is limited due to poor transduction into brain microvascular endothelial cells. In the present study, a fusion protein of human SOD2 and a brain targeting peptide, Angiopep-2 (AP-2), was generated by genetic engineering. The SOD2-AP-2 and control SOD2 were successfully expressed in Escherichia coli and purified using immobilized metal affinity chromatography. Purified SOD2-AP-2 exhibited 1,090 l/mg of specific SOD activity, which retained a significant activity in the same order of magnitude as that of native SOD2. The in vitro transduction demonstrated that 1 lM of SOD2-AP-2 delivered efficiently to immortalized mouse brain endothelial cell line within 30 min whereas, control SOD2 did not. Moreover, pretreatment with 50 units of SOD2-AP-2 for 1 h could significantly protect cells against paraquat up to 2 mM but control SOD2 pretreatment did not show a protective effect. Taken together, our findings pave the way for SOD2-AP-2 to be a potential therapeutic candidate for neurological diseases.

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“…The crystal structures of Mn-SOD from Caenorhabditis elegans [50], and E. coli [49] suggest that the dimer interface is located near the active site of each monomer and contributes to the integrity of the active site of the enzyme. In human SOD tetrameric complex that is formed by dimerization of two dimeric subunits, the dimer interface between the subunits also play important role in thermostability and activity [18,51].…”

Section: Discussionmentioning

confidence: 98%

Unique Characteristics of Recombinant Hybrid Manganese Superoxide Dismutase from Staphylococcus equorum and S. saprophyticus

et al. 2016

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A recombinant hybrid of manganese dependent-superoxide dismutase of Staphylococcus equorum and S. saprophyticus has successfully been overexpressed in Escherichia coli BL21(DE3), purified, and characterized. The recombinant enzyme suffered from degradation and aggregation upon storage at -20 °C, but not at room temperature nor in cold. Chromatographic analysis in a size exclusion column suggested the occurrence of dimeric form, which has been reported to contribute in maintaining the stability of the enzyme. Effect of monovalent (Na(+), K(+)), divalent (Ca(2+), Mg(2+)), multivalent (Mn(2+/4+), Zn(2+/4+)) cations and anions (Cl(-), SO4 (2-)) to the enzyme stability or dimeric state depended on type of cation or anion, its concentration, and pH. However, tremendous effect was observed with 50 mM ZnSO4, in which thermostability of both the dimer and monomer was increased. Similar situation was not observed with MnSO4, and its presence was detrimental at 200 mM. Finally, chelating agent appeared to destabilize the dimer around neutral pH and dissociate it at basic pH. The monomer remained stable upon addition of ethylene diamine tetraacetic acid. Here we reported unique characteristics and stability of manganese dependent-superoxide dismutase from S. equorum/saprophyticus.

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Purification, crystallization and X-ray structures of the two manganese superoxide dismutases fromCaenorhabditis elegans

Cited by 18 publications

References 34 publications

A novel DrosophilaSOD2 mutant demonstrates a role for mitochondrial ROS in neurodevelopment and disease

A novel DrosophilaSOD2 mutant demonstrates a role for mitochondrial ROS in neurodevelopment and disease

Angiopep-2-Mediated Delivery of Human Manganese Superoxide Dismutase in Brain Endothelial Cells and its Protective Effect Against Oxidative Stress

Unique Characteristics of Recombinant Hybrid Manganese Superoxide Dismutase from Staphylococcus equorum and S. saprophyticus

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