Reduction-cleavable desferrioxamine B pulldown system enriches Ni(ii)-superoxide dismutase from a Streptomyces proteome.

Two resins with the hydroxamic acid siderophore desferrioxamine B (DFOB) immobilised as a free ligand or its Fe(iii) complex were prepared to screen the Streptomyces pilosus proteome for proteins involved in siderophore-mediated Fe(iii) uptake. The resin design included a disulfide bond to enable the release of bound proteins under mild reducing conditions. Proteomics analysis of the bound fractions did not identify proteins associated with siderophore-mediated Fe(iii) uptake, but identified nickel superoxide dismutase (NiSOD), which was enriched on the apo-DFOB-resin but not the Fe(iii)-DFOB-resin or the control resin. While DFOB is unable to sequester Fe(iii) from sites deeply buried in metalloproteins, the coordinatively unsaturated Ni(ii) ion in NiSOD is present in a surface-exposed loop region at the N-terminus, which might enable partial chelation. The results were consistent with the notion that the apo-DFOB-resin formed a ternary complex with NiSOD, which was not possible for either the coordinatively saturated Fe(iii)-DFOB-resin or the non-coordinating control resin systems. In support, ESI-TOF-MS measurements from a solution of a model Ni(ii)-SOD peptide and DFOB showed signals that correlated with a ternary Ni(ii)-SOD peptide-DFOB complex. Although any biological implications of a DFOB-NiSOD complex are unclear, the work shows that the metal coordination properties of siderophores might influence an array of metal-dependent biological processes beyond those established in iron uptake.