The recent expression of an azurin mutant where the blue type 1 copper site
is replaced by the purple Cu-A site of Paracoccus denitrificans cytochrome
c oxidase has yielded an optimal system for examining the unique electron
mediation properties of the binuclear Cu-A center, because both type 1 and
Cu-A centers are placed in the same location in the protein while all other
structural elements remain the same, Long-range electron transfer is induc
ed between the disulfide radical anion, produced pulse radiolytically, and
the oxidized binuclear Cu-A center in the purple azurin mutant. The rate co
nstant of this intramolecular process, k(ET) = 650 +/- 60 s(-1) at 298 K an
d pH 5.1, is almost 3-fold faster than for the same process in the wild-typ
e single blue copper azurin from Pseudomonas aeruginosa (250 +/- 20 s(-1)),
in spite of a smaller driving force (0.69 eV for purple Cu-A azurin vs, 0.
76 eV for blue copper azurin), The reorganization energy of the Cu-A center
is calculated to be 0.4 eV, which is only 50% of that found for the wild-t
ype azurin, These results represent a direct comparison of electron transfe
r properties of the blue and purple Cu-A sites in the same protein framewor
k and provide support for the notion that the binuclear purple Cu-A center
is a more efficient electron transfer agent than the blue single copper cen
ter because reactivity of the former involves a lower reorganization energy
.