Mössbauer studies of beef heart aconitase: evidence for facile interconversions of iron-sulfur clusters.

Abstract

Beef heart aconitase, isolated under aerobic conditions, was studied with Mossbauer and EPR spectroscopy. In the oxidized state, the ezyme exhibits an EPR signal at g = 2.01. The Mossbauer data show that his 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 integer electronic spin (S = 2); the Mossbauer spectra exhibit all the unique features reported for proteins with 3Fe clusters. On activation of aconitase with Fe2+, the paramagnetic 3Fe cluster of dithionite-reduced enzyme is converted into a diamagnetic (S = 0) form. Activation studies with Fe enriched in either 57Fe or 56Fe suggested 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 Fe2+. The data reported here suggest that a Fe2+ is used to rebuild a [4Fe-4S] cluster.