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) inh ibited proton translocation in ubiquinol:cytochrome c oxidoreductase ( cytochrome bc(1) complex) from yeast mitochondria where it was bound t o aspartate-160 of cytochrome b, In the current study, we report that DCCD and its fluorescent analogue, N-cyclohexyl-N'-[4-(dimethylamino)n aphthyl] carbodiimide (NCD-4), inhibit 50-60% proton pumping in the cy tochrome bc(1) complex of the bacterium Rhodobacter sphaeroides with a 20% inhibition of electron transfer activity. Radioactive DCCD is bou nd exclusively to cytochrome b at aspartate-187, which is located at t he C-terminal region of the CD loop connecting membrane-spanning helic es C and D of cytochrome b, Fluorescent studies with NCD-4 revealed th at 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 membra ne. (C) 1998 Academic Press.