THE INFLUENCE OF CONSERVED AROMATIC RESIDUES ON THE ELECTRON-TRANSFERREACTIVITY OF 2[4FE-4S] FERREDOXINS

The detailed mechanism used by [4Fe-4S] ferredoxins to exchange electr ons is not known. The importance of two highly conserved aromatic resi dues, each located close to one cluster of 2[4Fe-4S] ferredoxins has b een probed by site-directed mutagenesis of Clostridium pasteurianum fe rredoxin. All generated variants are less stable than the native prote in and only hydrophobic residues can replace one of the two conserved aromatic residues. With leucine substituting both aromatics, Clostridi um pasteurianum ferredoxin cannot even be completely purified because of its deleterious instability. The reduction potentials of Clostridiu m pasteurianum ferredoxin variants do not depend on the presence of ar omatic residues near the clusters. However, the ferredoxin from Entamo eba histolytica which is naturally devoid of aromatic residues display s a reduction potential nearly 60 mV less negative than that of Clostr idium pasteurianum ferredoxin. The rate constants for the oxidation of the reduced ferredoxins by the inorganic complexes hexaamine-cobalt(I II) chloride and sodium ethylenediaminetetra-acetatecobaltate(III) are similar. This implies that electron transfer from the clusters of the se molecules is not mediated by the conserved aromatic residues. These residues rather appear to be involved in maintaining the overall stab ility of ferredoxins.