Comparison of the crystal structures of genetically engineered human manganese superoxide dismutase and manganese superoxide dismutase from thermus thermophilus: Differences in dimer–dimer interaction

Abstract

The three‐dimensional X‐ray structure of a recombinant human mitochondrial manganese superoxide dismutase (MnSOD) (chain length 198 residues) was determined by the method of molecular replacement using the related structure of MnSOD from Thermus thermophilus as a search model. This tetrameric human MnSOD crystallizes in space group P21212 with a dimer in the asymmetric unit (Wagner, U.G., Werber, M.M., Beck, Y., Hartman, J.R., Frolow, F., & Sussman, J.L., 1989, J. Mol. Biol. 206, 787–788). Refinement of the protein structure (3, 148 atoms with Mn and no solvents), with restraints maintaining noncrystallographic symmetry, converged at an R‐factor of 0.207 using all data from 8.0 to 3.2 Å resolution and group thermal parameters. The monomer–monomer interactions typical of bacterial Fe‐ and Mn‐containing SODs are retained in the human enzyme, but the dimer–dimer interactions that form the tetramer are very different from those found in the structure of MnSOD from T. thermophilus. In human MnSOD one of the dimers is rotated by 84° relative to its equivalent in the thermophile enzyme. As a result the monomers are arranged in an approximately tetrahedral array, the dimer–dimer packing is more intimate than observed in the bacterial MnSOD from T. thermophilus, and the dimers interdigitate. The metal—ligand interactions, determined by refinement and verified by computation of omit maps, are identical to those observed in T. thermophilus MnSOD.

Biophysical and biochemical characterization of a liposarcoma‐derived recombinant MnSOD protein acting as an anticancer agent

“…CD was calibrated using (1S)- (1) [k] obs is the ellipticity (deg), mrw is the mean residue molecular weight, 111 Da, C is the protein concentration (g mL 21 ) and l is the optical path length of the cell (cm). A 0.1-cm path length cell and a protein concentration of about 0.05 mg mL 21 in 20-mM ammonium trifluoracetate, pH 7.8, were used.…”

“…A 0.1-cm path length cell and a protein concentration of about 0.05 mg mL 21 in 20-mM ammonium trifluoracetate, pH 7.8, were used. CD spectra were recorded with a time constant of 4 s, a 2-nm bandwidth and a scan rate of 5 nm min 21 . Four spectra were averaged after baseline correction by subtracting a buffer spectrum.…”

Synthesis, X-Ray Crystallographic Characterization, and Electronic Structure Studies of a Di-Azide Iron(III) Complex: Implications for the Azide Adducts of Iron(III) Superoxide Dismutase

“…6 In the resting states of both proteins, the metal ions are in five-coordinate, distorted trigonal bipyramidal ligand environments with a histidine (His) and a solvent molecule in the axial positions and two His residues and an aspartate (Asp) in the equatorial plane (Figure 1, left). [6][7][8][9][10][11] The substrate is proposed to bind trans to the Asp ligand (i.e., between the two equatorial His residues) to yield a six-coordinate, roughly octahedral complex. The rate constants for the reaction of Mn-and FeSODs with superoxide approach the diffusion-controlled limit, thus largely preventing direct studies of catalytic intermediates.…”

X‐ray structure analysis of an engineered Fe‐superoxide dismutase Gly‐Ala mutant with significantly reduced stability to denaturant

“…1 where the two regions which dominate the dimer-dimer quaternary interactions are indicated. Comparison of the dimer-dimer contacts within the tetrameric members of this family shows that they vary greatly from enzyme to enzyme [7,8]. In T. thermophilus SOD the dimers associate quite loosely by interactions involving residues of the helical hairpin and residues in the loop (144-152 in M. tuberculosis numbering) which links the two outer strands of the ~-sheet in the adjacent dimer [6].…”

“…In T. thermophilus SOD the dimers associate quite loosely by interactions involving residues of the helical hairpin and residues in the loop (144-152 in M. tuberculosis numbering) which links the two outer strands of the ~-sheet in the adjacent dimer [6]. In contrast, the dimers of the human mitochondrial SOD associate more tightly by virtue of extensive interactions between the helical hairpins of adjacent subunits [2,8] which give rise to four-helix bundle structures. In the M. tuberculosis SOD, the dimers associate such that the 144-152 loops of adjacent subunits interact extensively at the dimerdimer interface where they are disposed around one of the tetramer two-fold axes.…”