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The anaerobic, but aerotolerant Propionibacterium freudenreichii sp. shermanii contains a single superoxide dismutase [EC 1.15.1.1.] exhibiting comparable activity with iron or manganese as metal cofactor. The formation of superoxide dismutase is not depending on the supplementation of iron or manganese to the culture medium. Even in the absence of these metals the protein is built in comparable amounts. Bacteria grown in the absence of iron and manganese synthesize a superoxide dismutase with very low activity which had incorporated copper. If the medium was also depleted of copper, cobalt was incorporated, leading to an enzymically inactive form. In the absence of cobalt an enzymically inactive superoxide dismutase was built with unknown metal contents. Upon aeration the amount of superoxide dismutase activity increased continuously up to 9 h, due to a de novo synthesis of the protein. This superoxide dismutase had incorporated iron into the active centre. The superoxide dismutase of Propionibacterium shermanii is able to form a much wider variety of complexes with trace metal ions in vivo than previously recognized, leading to the hypothesis that the original function of these proteins was the binding of cytoplasmic trace metals present in excess.

Propionibacterium shermanii contains a single constitutive superoxide dismutase (SOD) which is active with either iron or manganese incorporated in the same protein moiety. Copper and cobalt can also be incorporated by the bacteria in the active center of the SOD under conditions of metal deficiency, but in this case the enzyme is enzymatically inactive. In contrast to other bacterial SODs, the Fe-SOD of P. shermanii remains highly resistant to inactivation by hydrogen peroxide, as does Mn-SOD. Both SOD types cannot be distinguished by their inactivation patterns. Incubation with hydrogen peroxide results in a concentration- and time-dependent decrease in tryptophan fluorescence, independent of the metal present in the active center. Moreover, the Fe-SOD shows a time-dependent decrease in spin concentration after addition of hydrogen peroxide, which reflects alterations in the environment of the metal rather than a reduction of Fe3+ to Fe2+. No obvious correlations exist, however, between these effects and the enzymatic activity of the enzyme. The resistance of the SODs from P. shermanii to inactivation by hydrogen peroxide seems to be caused by the fact that a tryptophan residue near the metal-chelating histidine-75--which is present in all Fe-SODs being rapidly inactivated by this agent--is exchanged for valine.

Propionibacterium shermanii, an aerotolerant anaerobic bacterium, has already been shown to incorporate, depending on the metal supplementation to the medium, either iron or manganese or copper into the same superoxide dismutase protein. The in vivo incorporation of cobalt in the same superoxide dismutase was obtained in an iron-, manganese- and copper-depleted medium. The protein was isolated and characterized by NMR which offers the possibility to identify the amino acid residues at the active site exploiting isotropically shifted proton resonance.

Propionibacterium shermanii, an aerotolerant anaerobe, produces an iron-containing or a manganese-containing superoxide dismutase, depending on the metal supplied in the culture medium [Meier, B., Barra, D., Bossa, F., Calabrese, L. & Rotilio, G. (1982) J. Biol. Chem. 257, 13977-13980]. In this study, we demonstrate in vivo incorporation of copper into an active superoxide-dismutase protein when iron and manganese are absent from the growth medium. Superoxide dismutases containing either iron, manganese or copper were isolated from P. shermanii, their complete amino acid sequences were determined and the identity of their protein moieties was established. The polypeptide chain is made up of 201 amino acid residues, corresponding to a molecular mass of 22.6 kDa. From sedimentation equilibrium experiments, the native protein shows a molecular mass of approximately 86 kDa and therefore consists of four identical subunits. The primary structure was compared with the structure of other Fe-superoxide dismutases and Mn-superoxide dismutases, in particular those possessing a strict metal cofactor specificity.