THE [2FE-2S] CENTER OF THE CYTOCHROME BC COMPLEX IN BACILLUS-FIRMUS OF4 IN EPR - AN EXAMPLE OF A MENAQUINOL-OXIDIZING RIESKE CENTER

The EPR and electrochemical properties of the Rieske [2Fe-2S] centre f rom the facultatively alkalophilic eubacterium Bacillus firmus strain OF4 are described. The redox midpoint potential of the Rieske centre w as determined to be +105 +/- 10 mV at pH 7.0. This value was found to be independent of pH up to pH 8 and to decrease at higher pH values wi th a slope slightly steeper than - 60 mV/pH unit. With respect to its spectral characteristics the Rieske centre of B. firmus OF4 shows feat ures typical for the [2Fe-2S] centres of cytochrome be complexes. The alteration of the EPR lineshape between the ascorbate-and dithionite-r educed samples is indicative for an interaction of the Rieske centre w ith the menaquinone pool. The orientation of the g-tensor (the gz( )an d gy( )axes are parallel to the membrane plane, g(x) is oriented perpe ndicular) is similar to that from Rieske clusters from other cytochrom e bc(1) and b(6)f complexes. The Q(o)-site inhibitors UHDBT and DBMIB affect the EPR spectrum of the Rieske centre. The effect of DBMIB is d ifferent from what has been reported for UQ/PQ-oxidizing Rieske centre s. Furthermore, the binding of the Q(o)-site inhibitor stigmatellin in duces changes in the EPR lineshape, but has no influence on the redox midpoint potential of the cluster. The lack of a redox effect upon add ition of stigmatellin and the influence of DBMIB on the EPR spectrum i s proposed to be characteristic for Rieske centres from Gram-positive bacteria. The results obtained are compared to data sets from a variet y of Rieske centres from different organisms. Together with previous r esults, these data support the definition of a new subset of Rieske ce ntres with unique properties. Furthermore, the implications of the det ermined electrochemical parameters of the Rieske centre for the functi oning of the electron transport chain of B. firmus under alkalophilic conditions are discussed.