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Spectroscopic studies on fully oxidized and semi-reduced desulfoferrodoxin purified from Desulfovibrio desulfuricans ATCC 27774
478
Abstracts
6037
SPECTROSCOPIC STUDIES ON FULLY OXIDIZED AND SEMIREDUCED DESULFOFERRODOXlN PURIFIED FROM Desulfovibrio desulfuricans ATCC 27774
Pedro T-l!*, Jo& J. G. Moura2g4, NatarajanRavi3,Boi HanhHuynh3,Jean LeGalP and lsabelMowal y2g4 ’ Cenirode Teawlog~
Quimica e M&gica,
Apartado 127,2?80 O&as, Portugal.
Novade Lisboa, 2825 Monteda Cape&a, * Departamanto de Quimica, FCT, Univmidade
Portugal.
3 Department of Physics, Emory Univarsity, Atlanta, GA 30322, USA. 4 Department of Biodwmistry, University of Georgia, Athens, GA 30602, USA.
Desulfoferrodoxin is a new type of iron-sulfur protein that contains an unusual combination of metal centers. This protein was purified to homogeneity from extracts of Desulfovibrio dem’firicans ATCC 27774. The protein is a monomer of molecular mass 14 kDa. It contains two iron atoms per molecule corresponding to two different types of centers: an FeS4 site very similar to that found in desulforedoxin (distorted tetrahedral cysteine coordination) from Desulfovibrio gigus and an octahedral coordinated high-spin iron mainly with 0 or N ligation. The protein was first isolatedf’) in the semi-reduced form in which the FeS4 center is in the ferric state and the octahedral center is in the ferrous state. This is rather unusual since under aerobic conditions, mononuclear iron centers in proteins generally exists in the ferric state. However, this is most probably due to the fact that this center also has a unusual high redox potential (-240 mv). The visible spectrum has maxima at 495, 368 and 279 nm and the EPR spectrum shows resonances at g= 7.94, 5.75, 4.1 and 1.8 characteristic of a high-spin ferric ion (S=5/2) with E/D= 0.08. This protein was also isolated in the totally oxidized form. In this form both centers are in the ferric state. The visible spectrum has extra contributions at 335 and 640 nm, and the EPR spectrum presents new features at g= 9.74 and 4.3 which can be attributed to a high-spin ferric ion (S=5/2, E/D=0.33). Upon reduction with ascorbate this totally oxidized form yields the semi-reduced form and the opposite is accomplished by oxidation with ferricyanide. Only one NH2-terminal sequence was obtained for both forms and the same amino acid composition, sustaining that we are in the presence of two forms of the same protein. Based on the NH2-terminal amino acid sequence determined so far this protein appears to be a close analog to the Rbo gene product from Desulfovibrio vulgaris [*I which was suggested to be a rubredoxin oxido-reductase. Nevertheless, reduced pyridine nucleotides failed to reduce the desulforedoxin like center of desulfoferrodoxin. The nature of the centers and the function of this protein is currently under investigation. Work supported
by IN/C, NIH and JNICT.
(‘1 Moura, I., Tavares, P., Moura, J.J.G., Ravi, N., Huynh, B.H., Liu, M., LeGall, J. (1990) J. Biol. c&m. 265,21596-21602. [*I Brumlik, M.J., Voordow, 0. (1999) J. Bacferiol. 171, 4999660004.
Abstracts
6037
SPECTROSCOPIC STUDIES ON FULLY OXIDIZED AND SEMIREDUCED DESULFOFERRODOXlN PURIFIED FROM Desulfovibrio desulfuricans ATCC 27774
Pedro T-l!*, Jo& J. G. Moura2g4, NatarajanRavi3,Boi HanhHuynh3,Jean LeGalP and lsabelMowal y2g4 ’ Cenirode Teawlog~
Quimica e M&gica,
Apartado 127,2?80 O&as, Portugal.
Novade Lisboa, 2825 Monteda Cape&a, * Departamanto de Quimica, FCT, Univmidade
Portugal.
3 Department of Physics, Emory Univarsity, Atlanta, GA 30322, USA. 4 Department of Biodwmistry, University of Georgia, Athens, GA 30602, USA.
Desulfoferrodoxin is a new type of iron-sulfur protein that contains an unusual combination of metal centers. This protein was purified to homogeneity from extracts of Desulfovibrio dem’firicans ATCC 27774. The protein is a monomer of molecular mass 14 kDa. It contains two iron atoms per molecule corresponding to two different types of centers: an FeS4 site very similar to that found in desulforedoxin (distorted tetrahedral cysteine coordination) from Desulfovibrio gigus and an octahedral coordinated high-spin iron mainly with 0 or N ligation. The protein was first isolatedf’) in the semi-reduced form in which the FeS4 center is in the ferric state and the octahedral center is in the ferrous state. This is rather unusual since under aerobic conditions, mononuclear iron centers in proteins generally exists in the ferric state. However, this is most probably due to the fact that this center also has a unusual high redox potential (-240 mv). The visible spectrum has maxima at 495, 368 and 279 nm and the EPR spectrum shows resonances at g= 7.94, 5.75, 4.1 and 1.8 characteristic of a high-spin ferric ion (S=5/2) with E/D= 0.08. This protein was also isolated in the totally oxidized form. In this form both centers are in the ferric state. The visible spectrum has extra contributions at 335 and 640 nm, and the EPR spectrum presents new features at g= 9.74 and 4.3 which can be attributed to a high-spin ferric ion (S=5/2, E/D=0.33). Upon reduction with ascorbate this totally oxidized form yields the semi-reduced form and the opposite is accomplished by oxidation with ferricyanide. Only one NH2-terminal sequence was obtained for both forms and the same amino acid composition, sustaining that we are in the presence of two forms of the same protein. Based on the NH2-terminal amino acid sequence determined so far this protein appears to be a close analog to the Rbo gene product from Desulfovibrio vulgaris [*I which was suggested to be a rubredoxin oxido-reductase. Nevertheless, reduced pyridine nucleotides failed to reduce the desulforedoxin like center of desulfoferrodoxin. The nature of the centers and the function of this protein is currently under investigation. Work supported
by IN/C, NIH and JNICT.
(‘1 Moura, I., Tavares, P., Moura, J.J.G., Ravi, N., Huynh, B.H., Liu, M., LeGall, J. (1990) J. Biol. c&m. 265,21596-21602. [*I Brumlik, M.J., Voordow, 0. (1999) J. Bacferiol. 171, 4999660004.