The influence of conserved aromatic residues on the electron transfer reactivity of 2[4Fe-4S] ferredoxins

• Published in: Biochimica et biophysica acta 1996-07, Vol.1295 (2), p.201-208
• Main Authors: Quinkal, Isabelle, Kyritsis, Panayotis, Kohzuma, Takamitsu, Im, Sang-Choul, Sykes, A.Geoffrey, Moulis, Jean-Marc
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; C. pasteurianum; Clostridium - chemistry; Drug Stability; E. histolytica; Electrochemistry; Electron Transport; Ferredoxins - chemistry; Ferredoxins - metabolism; Iron-sulfur cluster; Kinetics; Molecular Sequence Data; Mutagenesis, Site-Directed; Oxidation-Reduction; Reduction potential; Site-directed mutagenesis; Spectrophotometry; Stopped-flow spectroscopy; Structure-Activity Relationship
• ISSN: 0167-4838; 0006-3002; 1879-2588
• DOI: 10.1016/0167-4838(96)00042-8

The detailed mechanism used by [4Fe-4S] ferredoxins to exchange electrons is not known. The importance of two highly conserved aromatic residues, each located close to one cluster of 2[4Fe-4S] ferredoxins has been probed by site-directed mutagenesis of Clostridium pasteurianum ferredoxin. All generated variants are less stable than the native protein and only hydrophobic residues can replace one of the two conserved aromatic residues. With leucine substituting both aromatics, Clostridium pasteurianum ferredoxin cannot even be completely purified because of its deleterious instability. The reduction potentials of Clostridium pasteurianum ferredoxin variants do not depend on the presence of aromatic residues near the clusters. However, the ferredoxin from Entamoeba histolytica which is naturally devoid of aromatic residues displays a reduction potential nearly 60 mV less negative than that of Clostridium pasteurianum ferredoxin. The rate constants for the oxidation of the reduced ferredoxins by the inorganic complexes hexaamine-cobalt(III) chloride and sodium ethylenediaminetetra-acetatecobaltate(III) are similar. This implies that electron transfer from the clusters of these molecules is not mediated by the conserved aromatic residues. These residues rather appear to be involved in maintaining the overall stability of ferredoxins.