Abstract
Proton transfer involving internal water molecules that provide hydrogen bonds and facilitate proton diffusion has been identified in some membrane proteins. Arg-94 in cytochrome b of the Rhodobacter sphaeroides bc(1) complex is fully conserved and is hydrogen-bonded to the heme propionate and a chain of water molecules. To further elucidate the role of Arg-94, we generated the mutations R94A, R94D, and R94N. The wild-type and mutant bc(1) complexes were purified and then characterized. The results show that substitution of Arg-94 decreased electron transfer activity and proton pumping capability and increased O(2)(.) production, suggesting the importance of Arg-94 in the catalytic mechanism of the bc(1) complex in R. sphaeroides. This also suggests that the transport of H(+), O(2), and O(2)(.) in the bc(1) complex may occur by the same pathway.