ELECTRON-PARAMAGNETIC-RESONANCE STUDIES OF SUCCINATE-UBIQUINONE OXIDOREDUCTASE FROM PARACOCCUS-DENITRIFICANS - EVIDENCE FOR A MAGNETIC INTERACTION BETWEEN THE 3FE-4S CLUSTER AND CYTOCHROME-B

Electron paramagnetic resonance (EPR) studies of succinate:ubiquinone oxidoreductase (SQR) from Paracoccus denitrificans have been undertake n in the purified and membrane-bound states, Spectroscopic ''signature s'' accounting for the three iron-sulfur clusters (2Fe-2S, 3Fe-4S, and 4Fe-4S), cytochrome b, flavin, and protein-bound ubisemiquinone radic als have been obtained in air-oxidized, succinate-reduced, and dithion ite-reduced preparations at 4-10 K. Spectra obtained at 170 K in the p resence of excess succinate showed a signal typical of that of a flavi n radical, but superimposed with another signal. The superimposed sign al originated from two bound ubisemiquinones, as shown by spectral sim ulations, Power saturation measurements performed on the air-oxidized enzyme provided evidence for a weak magnetic dipolar interaction opera ting between the oxidized 3Fe-4S cluster and the oxidized cytochrome b . Power saturation experiments performed on the succinate- and dithion ite-reduced forms of the enzyme demonstrated that the 4Fe-4S cluster i s coupled weakly to both the 2Fe-2S and the 3Fe-4S clusters, Quantitat ive interpretation of these power saturation experiments has been achi eved through redox calculations. They revealed that a spin-spin intera ction between the reduced 3Fe-4S cluster and the cytochrome b (oxidize d) may also exist. These findings form the first direct EPR evidence f or a close proximity (less than or equal to 2 nm) of the high potentia l 3Fe-4S cluster, situated in the succinate dehydrogenase part of the enzyme, and the low potential, low spin b-heme in the membrane anchor of the enzyme.