Characterization of Soluble Forms of the Membrane Bound Iso-Cytochrome c2 from Rhodobacter Capsulatus

Abstract

Soluble cytochrome c2 functions to couple the membrane bound cytochrome bc1 complex to photosynthetic reaction centers. When the cytochrome c2 gene is inactivated, Rb. capsulatusis still capable of photosynthetic growth due to the presence of a constitutively produced membrane bound isozyme of cytochrome c2 known as cytochrome cy (Jenney & Daldal, EMBO J. 12, 1283-1292 (1993)). We have now isolated small amounts of two forms of the membrane bound cytochrome c2 isozyme which are naturally soluble. Forms A and B separate on CM-cellulose chromatography. They have similar amino acid compositions, which are like the C-terminus of the translated isozyme gene and unlike that of cytochrome c2. The N-terminal sequence of form A is: EPAAPPPPAYVEVDPATITG… and of form B is: APPPPAYVEV…, indicating non-specific proteolytic cleavage at more than one site. The absorption spectra and redox potential of the two forms are like those of cytochrome c2. The kinetics of interaction of form B with the photosynthetic reaction center indicate that there is no stable complex formed and that the reaction is entirely second-order. The rate constant for iso-c2 is more than an order of magnitude smaller at low ionic strength and the electostatic effect is one half that of cytochrome c2. Thus, reaction centers show significant specificity for the natural electron donor.