Abstract
Caenorhabditis elegans expresses two manganese superoxide dismutase enzymes (MnSOD-2 and MnSOD-3) that are targeted to the mitochondrion. MnSOD-2 is constitutively expressed, while synthesis of MnSOD-3 is inducible. The structures of these two mononuclear metalloenzymes have been determined to 1.8 and 1.7 A resolution, respectively. Pink crystals formed in space group P4(1)2(1)2 for each, with unit-cell parameters a = b = 81.0, c = 137.4 A for MnSOD-2 and a = b = 81.8, c = 136.0 A for MnSOD-3. The final structure of MnSOD-3 was refined to R = 21.6% and R(free) = 26.2% at 293 K, and R = 18.9% and R(free) = 22.6% at 100 K, while that of MnSOD-2 was refined to R = 16.9% and R(free) = 20.1% at 100 K. The asymmetric unit cell is comprised of two subunits. The resulting structures are very similar to that of human MnSOD and form a tetramer corresponding to a dimer of dimers. The subunit interface between dimers is comprised of two four-helix bundles that stabilize the biologically significant homotetramer.
Highlights
The superoxide dismutases (SODs; EC 1.15.11) are ubiquitous metalloproteins whose purpose is to detoxify the highly reactive superoxide anion (OÀ2 ) by its dismutation into oxygen and hydrogen peroxide (McCord & Fridovich, 1969)
It serves as an interesting model to investigate SOD since it encodes five sod genes producing two cytosolic Cu/ZnSODs (SOD-1 and SOD-5), one extracellular Cu/ZnSOD (SOD-4) and two MnSODs (SOD-2 and SOD-3; designated MnSOD-2 and MnSOD-3, respectively) (C. elegans Genome Consortium, 1998)
MnSOD-2 and MnSOD-3 were overexpressed in E. coli without an N-terminal histidine tag, they each purified reasonably well using metal-chelation affinity chromatography (MCAC)
Summary
The superoxide dismutases (SODs; EC 1.15.11) are ubiquitous metalloproteins whose purpose is to detoxify the highly reactive superoxide anion (OÀ2 ) by its dismutation into oxygen and hydrogen peroxide (McCord & Fridovich, 1969). Many organisms contain more than one type of SOD, distinguishable by their metal cofactor and subcellular location. Most eukaryotes, such as Caenorhabditis elegans, express both Cu/ZnSOD and MnSOD (Fridovich, 1975). Dissection of the insulin/IGF signalling pathway has identified sod-3 as a significant target for the DAF-16/FOXO transcription factor. Inhibition of the insulin/IGF signalling pathway results in elevated expression of sod-3 and an extension in lifespan (Murphy et al, 2003; Lee et al, 2003; Dong et al, 2007). In a complementary approach to the study of differences in expression and cellular roles of the SODs of C. elegans, we are investigating the differences between the proteins themselves. We present the structures of the two MnSODs which are strikingly similar to that from human (Hearn et al, 2003)
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