Dicyclohexylcarbodiimide inhibits proton pumping in Ubiquinol:: Cytochrome c oxidoreductase of Rhodobacter sphaeroides and binds to aspartate-187 of cytochrome b

In recent studies we reported that dicyclohexylcarbodiimide (DCCD) inhibited proton translocation in ubiquinol:cytochrome c oxidoreductase (cytochrome bc(1) complex) from yeast mitochondria where it was bound to aspartate-160 of cytochrome b, In the current study, we report that DCCD and its fluorescent analogue, N-cyclohexyl-N'-[4-(dimethylamino)naphthyl] carbodiimide (NCD-4), inhibit 50-60% proton pumping in the cytochrome bc(1) complex of the bacterium Rhodobacter sphaeroides with a 20% inhibition of electron transfer activity. Radioactive DCCD is bound exclusively to cytochrome b at aspartate-187, which is located at the C-terminal region of the CD loop connecting membrane-spanning helices C and D of cytochrome b, Fluorescent studies with NCD-4 revealed that aspartate-187 is located in a mildly hydrophobic pocket in the bc(1) complex at a distance of 2-3 Angstrom from the surface of the membrane. (C) 1998 Academic Press.