A pure S = 3/2 [Fe4S4]+ cluster in the A33Y variant of Pyrococcus furiosus ferredoxin.
The properties of the [4Fe-4S]2+/+ cluster in wild-type and the A33Y variant of Pyrococcus furiosus ferredoxin have been investigated by the combination of EPR, variable-temperature magnetic circular dichroism (VTMCD) and resonance Raman (RR) spectroscopies. The A33Y variant involves the replacement of an alanine whose alpha-C is less than 4 A from one of the cluster iron atoms by a tyrosine residue. Although the spectroscopic results give no indication of tyrosyl cluster ligation, the presence of a tyrosine residue in close proximity to the cluster results in a 38-mV decrease in the midpoint potential of the [4Fe-4S]2+/+ couple and has a marked effect on the ground state properties of the reduced cluster. The mixed spin [4Fe-4S]+ cluster in the wild-type protein, 80% S = 3/2 (E/D = 0.22, D = +3.3 cm(-1)) and 20% S = 1/2 (g = 2.10, 1.87, 1.80), is converted into a homogeneous S = 3/2 (E/D = 0.30, D = -0.7 cm(-1)) form in the A33Y variant. As the first example of a pure S = 3/2 [4Fe-4S]+ cluster in a ferredoxin, this variant affords the opportunity for detailed characterization of the excited electronic properties via VTMCD studies and demonstrates that the protein environment can play a crucial role in determining the ground state properties of [4Fe-4S]+ clusters.
Department of Chemistry, Center for Metalloenzyme Studies, University of Georgia, Athens 30602, USA.
R E
RE
Duderstadt
P S
PS
Brereton
M W
MW
Adams
M K
MK
Johnson
D002942:Circular Dichroism; D005288:Ferredoxins / Q000737:chemistry*; D014644:Genetic Variation; D016296:Mutagenesis; D020027:Pyrococcus furiosus / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D013059:Spectrum Analysis, Raman; D013696:Temperature
D005288:Ferredoxins; D011994:Recombinant Proteins