CYTOCHROME C(Y) OF RHODOBACTER-CAPSULATUS IS ATTACHED TO THE CYTOPLASMIC MEMBRANE BY AN UNCLEAVED SIGNAL SEQUENCE-LIKE ANCHOR

During the photosynthetic growth of Rhodobacter capsulatus, electrons are conveyed from the cytochrome (cyt) bc(1) complex to the photochemi cal reaction center by either the periplasmic cyt c(2) or the membrane -bound cyt c(y). Cyt c(y) is a member of a recently established subcla ss of bipartite c-type cytochromes consisting of an amino (N)-terminal domain functioning as a membrane anchor and a carboxyl (C)-terminal d omain homologous to cyt c of various sources, Structural homologs of c yt c(y) have now been found in several bacterial species, including Rh odobacter sphaeroides. In this work, a C-terminally epitope-tagged and functional derivative of R. capsulatus cyt c(y) was purified from int racytoplasmic membranes to homogeneity. Analyses of isolated cyt c(y) indicated that its spectral and thermodynamic properties are very simi lar to those of other c-type cytochromes, in particular to those from bacterial and plant mitochondrial sources, Amino acid sequence determi nation for purified cyt c(y) revealed that its signal sequence-like N- terminal portion is uncleaved; hence, it is anchored to the membrane, To demonstrate that the N-terminal domain of cyt c(y) is indeed its me mbrane anchor, this sequence was fused to the N terminus of cyt c(2). The resulting hybrid cyt c (MA-c(2)) remained membrane bound and was a ble to support photosynthetic growth of R. capsulafin in the absence o f the cyt c(y) and c(2). Therefore, cyt c(2) can support cyclic electr on transfer during photosynthetic growth in either a freely difusible or a membraneanchored form, These findings should now allow for the fi rst time the comparison of electron transfer properties of a given ele ctron carrier when it is anchored to the membrane or is freely diffusi ble in the periplasm.