Electrochemical study of the redox properties of [2Fe‐2S] ferredoxins Evidence for superreduction of the Rieske [2Fe‐2S] cluster

Abstract

what determines the reduction potential(s) of these proteins, and what determines whether their functioning is associated with the transfer of no, one, one pair, or multiple pairs of electrons.By a fortunate coincidence of circumstances we have found it possible to study the complete redox behaviour of a biological [2Fe-2S] system: (i) the Rieske protein can be purified as a water-soluble fragment with no observable change in its ironsulfur cluster [5]; (ii) the fragment exhibits a direct, unmediated electrochemical response on glassy carbon [6]; (iii) the first reduction potential of the Rieske cluster is unusually high, namely Era,7 = + 0.3 V [7]; (iv) carbon electrodes have a high overpotential for H 2 evolution, therefore, their use allows for aqueous-solution bio-electrochemistry down to E = -1 V [8]. The voltammetric characterization of the Rieske cluster and the comparison of its properties with other (i.e. low-potential) biological [2Fe-2S] clusters are the subject of this study.

Direct electrochemistry and EPR spectroscopy of spinach ferredoxin mutants with modified electron transfer properties

“…The response of this homogeneous preparation on glassy carbon is indistinguishable from that previously observed with the natural mixture (see Fig. 1, Table 1, [10]). Therefore, below we refer to the homogeneous FdI as wild-type.…”

“…All reported potentials have been recalculated with respect to the NHE. The glassy carbon surface was activated by oxidation in 65% HNO3, neutralization, mild polishing with 0.6 pm particles [9] followed by heating to redness over a methane flame to obtain a hydrophilic surface [10]. The final solution contained ca.…”

“…The direct electrochemistry of spinach ferredoxin has been previously described as a quasi-reversible response on glassy carbon [10]. Wild-type spinach ferredoxin is a mixture of two proteins with strong sequence homology.…”

Voltammetric Studies of Spinach Ferredoxin on a Glassy Carbon Electrode Modified with Cobalt(II) Tetrasulfophthalocyanine

“…Electrochemistry has evolved as a valuable technique for the study of redox properties of small redox proteins. Several studies on the dependence of the redox potentials of the proteins on parameters such as temperature and pH have been conducted [2][3][4]. Kinetic data in the form of rate constants have also been obtained from electrochemical studies of proteins [4][5][6].…”

Formation and characterization of an all-ferrous Rieske cluster and stabilization of the [2Fe-2S] 0 core by protonation

“…A [2Fe-2S] 0 cluster was generated artificially in spinach ferredoxin by irreversible complexation of the protein to a chromium reductant, increasing the reduction potential significantly and adding extra positive charge (12). Voltammetric signals from the soluble Rieske domain from the bovine heart cytochrome bc 1 complex (Ϫ0.84 V at pH 7, potential reported to be pH independent) were attributed to formation of the all-ferrous cluster, but no characterization was attempted (13). Here, we describe the reversible formation of a stable, unmodified [2Fe-2S] 0 clustercontaining protein and its extensive characterization.…”