Abstract
Cytochrome c1 of Rhodobacter (Rba.) species provides a series of mutants which change barriers for electron transfer through the cofactor chains of cytochrome bc1 by modifying heme c1 redox midpoint potential. Analysis of post-flash electron distribution in such systems can provide useful information about the contribution of individual reactions to the overall electron flow. In Rba. capsulatus, the non-functional low-potential forms of cytochrome c1 which are devoid of the disulfide bond naturally present in this protein revert spontaneously by introducing a second-site suppression (mutation A181T) that brings the potential of heme c1 back to the functionally high levels, yet maintains it some 100 mV lower from the native value. Here we report that the disulfide and the mutation A181T can coexist in one protein but the mutation exerts a dominant effect on the redox properties of heme c1 and the potential remains at the same lower value as in the disulfide-free form. This establishes effective means to modify a barrier for electron transfer between the FeS cluster and heme c1 without breaking disulfide. A comparison of the flash-induced electron transfers in native and mutated cytochrome bc1 revealed significant differences in the post-flash equilibrium distribution of electrons only when the connection of the chains with the quinone pool was interrupted at the level of either of the catalytic sites by the use of specific inhibitors, antimycin or myxothiazol. In the non-inhibited system no such differences were observed. We explain the results using a kinetic model in which a shift in the equilibrium of one reaction influences the equilibrium of all remaining reactions in the cofactor chains. It follows a rather simple description in which the direction of electron flow through the coupled chains of cytochrome bc1 exclusively depends on the rates of all reversible partial reactions, including the Q/QH2 exchange rate to/from the catalytic sites.
Highlights
Bioenergetic enzymes use chains of cofactors to transfer electrons over long distances and connect catalytic sites with substrate redox pools
We observe that the disulfide and A181T can coexist, but the mutation exerts a dominant effect on the redox properties of heme c1 and the potential remains at the same lower value as in the disulfide-free form
Earlier work has established two independent structural motifs that contribute to controlling the integrity of the heme binding pocket to raise the redox potential in cytochrome c1 of Rba. capsulatus [26]
Summary
Bioenergetic enzymes use chains of cofactors to transfer electrons over long distances and connect catalytic sites with substrate redox pools. One of the prominent examples of this sort of arrangement is a twochain assembly in cytochrome bc (complex III), a common component of respiratory and photosynthetic electron transfer systems In this enzyme a c-chain comprising high-potential cytochromes c1 and c and the 2 iron–2 sulfur cluster (FeS) and the b-chain comprising two low-potential hemes b (bL and bH) and quinone of the Qi site are linked together by the action of the centrally located quinone binding Qo site. The other group of mutants includes the lowpotential forms of closely related Rba. capsulatus cytochrome c1 which are devoid of the disulfide bond naturally present in this species [26,28] (see Fig. 1) and in Rba. sphaeroides [29] These nonfunctional forms revert spontaneously by introducing a second-site suppression (mutation A181T) that brings the potential of heme c1 back to the functionally high level yet maintains it some 100 mV lower from the native value. To explain the results we developed a kinetic model which in our view improves the general understanding of electron flow through the coupled chains of cytochrome bc
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
