The ubiquitous superoxide dismutases (SODs) are critical on protecting the cell against a toxic radical anion, superoxide. SODs catalyse the disproportionation of superoxide to molecular oxygen and hydrogen peroxide in a mechanism that involves proton-coupled electron-transfer (PCET). SODs are known as the fastest enzyme with the largest k_cat/K_m of any known enzyme. Manganese SOD (MnSOD) from Escherichia coli is known to be fully fully active only as a homodimer. To gain insights into the mechanism of the PCET reaction mechanism, a potentially key residue Ser126 at the conserved but asymmetric dimer interface of the MnSOD from E. coli was mutated to Asp with the initial intent to generate a monomeric species. The expression was carried out in E. coli QC781, a SOD-deficient strain, transformed by pDT1-5 containing a sodA locus. 

Mutation of Ser126 significantly reduces the superoxide dismutase activities to 6 % and 5% of those of the wild-type enzyme at pH 7.8 and 6.0, respectively. Sedimentation velocity analysis indicated a dimeric species at pH 7.8, which becomes partially dissociated at pH 6.0. X-ray diffraction studies on crystals prepared at pH 8.7 (1.65 Ǻ) and pH 6.7 (1.56 Ǻ) show different space groups. The tightly packed symmetrical dimer interface is preserved but exhibited a considerable disorder at pH 6.7. The structural analyses of MnSOD S126D by exploring possible changes in water structure provide new information to examine the hypothesis of the Glu170 bridge as a proton shuttle in an outer-sphere mechanism for proton delivery to the nascent peroxo species.