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Tetramerization reinforces the dimer interface of MnSOD.

Sheng Y, Durazo A, Schumacher M, Gralla EB, Cascio D, Cabelli DE, Valentine JS - PLoS ONE (2013)

Bottom Line: Although CaMnSODc was found to crystallize as a tetramer, there is no indication from the solution properties that the functionality of CaMnSODc in vivo depends upon the formation of the tetrameric structure.Dimeric CaMnSODc was found to be significantly more subject to thermal or denaturant-induced unfolding than tetrameric ScMnSOD.We conclude that the tetrameric assembly strongly reinforces the dimer interface, which is critical for MnSOD activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of California Los Angeles, Los Angeles, California, United States of America.

ABSTRACT
Two yeast manganese superoxide dismutases (MnSOD), one from Saccharomyces cerevisiae mitochondria (ScMnSOD) and the other from Candida albicans cytosol (CaMnSODc), have most biochemical and biophysical properties in common, yet ScMnSOD is a tetramer and CaMnSODc is a dimer or "loose tetramer" in solution. Although CaMnSODc was found to crystallize as a tetramer, there is no indication from the solution properties that the functionality of CaMnSODc in vivo depends upon the formation of the tetrameric structure. To elucidate further the functional significance of MnSOD quaternary structure, wild-type and mutant forms of ScMnSOD (K182R, A183P mutant) and CaMnSODc (K184R, L185P mutant) with the substitutions at dimer interfaces were analyzed with respect to their oligomeric states and resistance to pH, heat, and denaturant. Dimeric CaMnSODc was found to be significantly more subject to thermal or denaturant-induced unfolding than tetrameric ScMnSOD. The residue substitutions at dimer interfaces caused dimeric CaMnSODc but not tetrameric ScMnSOD to dissociate into monomers. We conclude that the tetrameric assembly strongly reinforces the dimer interface, which is critical for MnSOD activity.

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The tetramer interfaces are highly disordered, when CaMnSODc is in the tetramer form.The ribbon diagram of ScMnSOD (PDB code: 3LSU) is shown in Panel A. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. The ribbon diagram of tetrameric CaMnSODc (PDB code: 3QVN) and the N-terminal helical region (residues 1–91) of a CaMnSODc monomer are shown in Panel B. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. Manganese ions are indicated as purple spheres.
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pone-0062446-g002: The tetramer interfaces are highly disordered, when CaMnSODc is in the tetramer form.The ribbon diagram of ScMnSOD (PDB code: 3LSU) is shown in Panel A. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. The ribbon diagram of tetrameric CaMnSODc (PDB code: 3QVN) and the N-terminal helical region (residues 1–91) of a CaMnSODc monomer are shown in Panel B. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. Manganese ions are indicated as purple spheres.

Mentions: WT ScMnSOD and WT CaMnSODc and their RP-mutant proteins were overexpressed and purified from S. cerevisiae. Seeking clues as to why ScMnSOD was a tetramer and CaMnSODc a dimer or “loose tetramer” in solution, despite sharing a high sequence identity, we determined their crystal structures [9]. The crystal structure further confirmed that ScMnSOD was a homotetramer (Figure 2A). By contrast, CaMnSODc appeared as a homotetramer (Figure 2B) in crystal structures.


Tetramerization reinforces the dimer interface of MnSOD.

Sheng Y, Durazo A, Schumacher M, Gralla EB, Cascio D, Cabelli DE, Valentine JS - PLoS ONE (2013)

The tetramer interfaces are highly disordered, when CaMnSODc is in the tetramer form.The ribbon diagram of ScMnSOD (PDB code: 3LSU) is shown in Panel A. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. The ribbon diagram of tetrameric CaMnSODc (PDB code: 3QVN) and the N-terminal helical region (residues 1–91) of a CaMnSODc monomer are shown in Panel B. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. Manganese ions are indicated as purple spheres.
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Related In: Results  -  Collection

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pone-0062446-g002: The tetramer interfaces are highly disordered, when CaMnSODc is in the tetramer form.The ribbon diagram of ScMnSOD (PDB code: 3LSU) is shown in Panel A. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. The ribbon diagram of tetrameric CaMnSODc (PDB code: 3QVN) and the N-terminal helical region (residues 1–91) of a CaMnSODc monomer are shown in Panel B. The four subunits are colored in: A, yellow; B, orange; C, green; D, cyan. Manganese ions are indicated as purple spheres.
Mentions: WT ScMnSOD and WT CaMnSODc and their RP-mutant proteins were overexpressed and purified from S. cerevisiae. Seeking clues as to why ScMnSOD was a tetramer and CaMnSODc a dimer or “loose tetramer” in solution, despite sharing a high sequence identity, we determined their crystal structures [9]. The crystal structure further confirmed that ScMnSOD was a homotetramer (Figure 2A). By contrast, CaMnSODc appeared as a homotetramer (Figure 2B) in crystal structures.

Bottom Line: Although CaMnSODc was found to crystallize as a tetramer, there is no indication from the solution properties that the functionality of CaMnSODc in vivo depends upon the formation of the tetrameric structure.Dimeric CaMnSODc was found to be significantly more subject to thermal or denaturant-induced unfolding than tetrameric ScMnSOD.We conclude that the tetrameric assembly strongly reinforces the dimer interface, which is critical for MnSOD activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of California Los Angeles, Los Angeles, California, United States of America.

ABSTRACT
Two yeast manganese superoxide dismutases (MnSOD), one from Saccharomyces cerevisiae mitochondria (ScMnSOD) and the other from Candida albicans cytosol (CaMnSODc), have most biochemical and biophysical properties in common, yet ScMnSOD is a tetramer and CaMnSODc is a dimer or "loose tetramer" in solution. Although CaMnSODc was found to crystallize as a tetramer, there is no indication from the solution properties that the functionality of CaMnSODc in vivo depends upon the formation of the tetrameric structure. To elucidate further the functional significance of MnSOD quaternary structure, wild-type and mutant forms of ScMnSOD (K182R, A183P mutant) and CaMnSODc (K184R, L185P mutant) with the substitutions at dimer interfaces were analyzed with respect to their oligomeric states and resistance to pH, heat, and denaturant. Dimeric CaMnSODc was found to be significantly more subject to thermal or denaturant-induced unfolding than tetrameric ScMnSOD. The residue substitutions at dimer interfaces caused dimeric CaMnSODc but not tetrameric ScMnSOD to dissociate into monomers. We conclude that the tetrameric assembly strongly reinforces the dimer interface, which is critical for MnSOD activity.

Show MeSH
Related in: MedlinePlus