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Electrochemical and spectroscopic characterization of 7Fe, 8Fe and 3Fe forms of Azotobacter vinelandii ferredoxin I
REDOX THERMODYNAMICS
A008
15
ELECTROCHEMICAL AND SPECTROSCOPIC CHARACTERIZATION OF 7FE, 8FE AND 3FE FORMS OF AZOTOBACTER VINELANDII FERREDOXIN I
B. Revntiens (a), J. N. Butt (b), G. M. Jensen (a), F. A. Armstrong (b), B. K. Burgess (c), and P. J. Stephens (a) (a) Department of Chemistry, University of Southern California, Los Angeles, CA 90089-0482; (b) Department of Chemistry, University of California at Irvine; (c) Department of Molecular Biology and Biochemistry, University of California at Irvine, Irvine, CA 92717.
Azotobacter vinelandii ferredoxin I (AvFdI) contains one [3Fe-4S] cluster and one [4Fe-4S] cluster (1,2). As purified aerobically these clusters exist in the +l and +2 oxidation states, respectively. Dithionite reduces the [3Fe4S] cluster to the 0 oxidation level. Recently, voltammetry (using modified graphite electrodes and aminoglycoside promoters) has allowed study of the reduction of the [4Fe-4S] cluster and further reduction of the [3Fe-4S] cluster (3). We report an extension of these studies to two chemically modified forms of AvFdI. In one, (8Fe)FdI, the [3Fe-4S] cluster has been converted to a [4Fe-4S] cluster (4). In the second, (3Fe)FdI, the [4Fe-4S] cluster has been removed by ferricyanide oxidation (5). The redox potentials of the [3Fe-4S] and [4Fe-4S] clusters of (8Fe)FdI and (3Fe)FdI permit the effects of cluster-cluster interaction and of protein conformational change to be examined. EPR and NMR spectra of chemically and electrochemically reduced (7Fe)FdI, @Fe)FdI, and (3Fe)FdI are reported and analyzed. (1) C. D. Stout, J. Mol Biol., 205,545 (1989) (2) P. J. Stephens et al., Biochemistry, 30,320O (1991) (3) S. E. Iismaa et al., J. Biol. Chem., 266,21563 (1991) (4) G. M. Jensen, A. E. Vazquez, B. K. Burgess, and P. J. Stephens, submitted. (5) T. V. Morgan et al., FEBS Lett, 183,206 (1985)
A008
15
ELECTROCHEMICAL AND SPECTROSCOPIC CHARACTERIZATION OF 7FE, 8FE AND 3FE FORMS OF AZOTOBACTER VINELANDII FERREDOXIN I
B. Revntiens (a), J. N. Butt (b), G. M. Jensen (a), F. A. Armstrong (b), B. K. Burgess (c), and P. J. Stephens (a) (a) Department of Chemistry, University of Southern California, Los Angeles, CA 90089-0482; (b) Department of Chemistry, University of California at Irvine; (c) Department of Molecular Biology and Biochemistry, University of California at Irvine, Irvine, CA 92717.
Azotobacter vinelandii ferredoxin I (AvFdI) contains one [3Fe-4S] cluster and one [4Fe-4S] cluster (1,2). As purified aerobically these clusters exist in the +l and +2 oxidation states, respectively. Dithionite reduces the [3Fe4S] cluster to the 0 oxidation level. Recently, voltammetry (using modified graphite electrodes and aminoglycoside promoters) has allowed study of the reduction of the [4Fe-4S] cluster and further reduction of the [3Fe-4S] cluster (3). We report an extension of these studies to two chemically modified forms of AvFdI. In one, (8Fe)FdI, the [3Fe-4S] cluster has been converted to a [4Fe-4S] cluster (4). In the second, (3Fe)FdI, the [4Fe-4S] cluster has been removed by ferricyanide oxidation (5). The redox potentials of the [3Fe-4S] and [4Fe-4S] clusters of (8Fe)FdI and (3Fe)FdI permit the effects of cluster-cluster interaction and of protein conformational change to be examined. EPR and NMR spectra of chemically and electrochemically reduced (7Fe)FdI, @Fe)FdI, and (3Fe)FdI are reported and analyzed. (1) C. D. Stout, J. Mol Biol., 205,545 (1989) (2) P. J. Stephens et al., Biochemistry, 30,320O (1991) (3) S. E. Iismaa et al., J. Biol. Chem., 266,21563 (1991) (4) G. M. Jensen, A. E. Vazquez, B. K. Burgess, and P. J. Stephens, submitted. (5) T. V. Morgan et al., FEBS Lett, 183,206 (1985)