Studies of the Cytochrome Subunits of Menaquinone:Cytochromec Reductase (bc Complex) of Bacillus subtilis

Abstract

The menaquinone:cytochrome c reductase, or bc complex , of Bacillus subtilis belongs to a third class of bc-type complex, distinct from the bc 1 and b 6 f classes. Using a mutagenesis approach, we demonstrate that the cyto-chrome b (QcrB) and c (QcrC) subunits of the complex give rise to bands at 22 and 29 kDa, respectively, after denaturing electrophoresis; that both subunits are required for proper complex assembly and/or stability; and that both subunits retain one heme molecule under denaturing conditions. This unusual property of a b-type cytochrome was investigated further. We present evidence for the existence of a covalent linkage between the polypeptide and heme b H and of an important role for Cys 43 in binding of heme b H. It is proposed that heme is also covalently attached to the cytochrome b subunit of b 6 f complexes of chloroplasts and cyanobacteria. The cytochrome bc 1 complex (quinol:cytochrome c oxi-doreductase) is an integral membrane protein complex that functions as part of an electron transfer chain by passing electrons from quinol in the membrane to a c-type cytochrome. Coupled to electron transfer is the transport of protons across the membrane, and consequently, the enzyme contributes to the proton motive force. The complex has been isolated from mitochondria and several bacteria, and a similar complex, called the b 6 f complex, has been isolated from plant chloro-plasts and from cyanobacteria. Recently, crystal structures of the soluble part of the Rieske protein (1) and of the intact complex (2) from bovine heart mitochondria have been solved. There is considerable variation, depending on the source, in the number of subunits making up the complex, but three subunits are always present: an iron-sulfur (Rieske) protein containing a high potential [2Fe-2S] cluster with cysteine and histidine ligation; a cytochrome b containing two low spin b-type hemes, b L and b H , both with bis-histidine ligation but with different electrochemical properties; and a cytochrome c