Abstract
Beef heart aconitase, isolated under aerobic conditions, has been studied with Mössbauer and EPR spectroscopy. In the oxidized state, the enzyme exhibits an EPR signal at g = 2.01. The Mössbauer data show that this signal is associated with a 3Fe cluster. In dithionite-reduced aconitase, the 3Fe cluster, probably of the [3Fe-3S] type, is in a paramagnetic state of interger electronic spin (S = 2); the Mössbauer spectra exhibit al the unique features reported for proteins with 3Fe clusters. On activation of aconitase with ferrous ion, the paramagnetic 3Fe cluster of dithionite-reduced enzyme is converted into a diamagnetic (S = 0) form. Activation studies with iron enriched in either 27 Fe or 56 Fe suggest that activation transforms the 3Fe cluster into a center that has a [4Fe-4S] core. This conclusion is supported by the observation that EPR signals characteristic of reduced [4Fe-4S] clusters can be elicited under appropriate conditions. It has frequently been assumed that the activation of aconitase with Fe2+ produces an active site containing a single ferrous ion. The data reported here suggest that a ferrous ion is used to rebuild a [4Fe-4S] cluster.
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