Desulfovibrio africanus ferredoxin III (Da FdIII) readily interconverts between a 7Fe and an 8Fe form with Asp-14 believed to provide a cluster ligand in the latter form. To investigate the factors important for cluster interconversion in Fe/S cluster-containing proteins we have studied two variants of Da FdIII produced by site-directed mutagenesis, Asp14Glu and Asp14His, with cluster incorporation performed in vitro. Characterisation of these proteins by UV/visible, EPR and 1H NMR spectroscopies revealed that the formation of the stable 7Fe form of these proteins takes some time to occur. Evidence is presented which indicates the [4Fe–4S]2+ cluster is incorporated prior to the [3Fe–4S]1+ cluster.
