Phototriggering Electron Flow through Re^I-modified Pseudomonas aeruginosa Azurins

The [Re^I(CO)_3(4,7-dimethyl-1,10-phenanthroline)(histidine-124)(tryptophan-122)] complex, denoted [Re^I(dmp)(W122)], of Pseudomonas aeruginosa azurin behaves as a single photoactive unit that triggers very fast electron transfer (ET) from a distant (2 nm) Cu^I center in the protein. Analysis of time-resolved (ps–μs) IR spectroscopic and kinetics data collected on [Re^I(dmp)(W122)AzM] (in which M=Zn^(II), Cu^(II), Cu^I; Az=azurin) and position-122 tyrosine (Y), phenylalanine (F), and lysine (K) mutants, together with excited-state DFT/time-dependent (TD)DFT calculations and X-ray structural characterization, reveal the character, energetics, and dynamics of the relevant electronic states of the [Re^I(dmp)(W122)] unit and a cascade of photoinduced ET and relaxation steps in the corresponding Re–azurins. Optical population of [Re^I(imidazole-H124)(CO)_3]→dmp ^1CT states (CT=charge transfer) is followed by around 110 fs intersystem crossing and about 600 ps structural relaxation to a ^3CT state. The IR spectrum indicates a mixed Re^I(CO)_3,A→dmp/π→π^*(dmp) character for aromatic amino acids A122 (A=W, Y, F) and Re^I(CO)_3→dmp metal–ligand charge transfer (MLCT) for [Re^I(dmp)(K122)AzCu^(II)]. In a few ns, the ^3CT state of [Re^I(dmp)(W122)AzM] establishes an equilibrium with the [Re^I(dmp.^−)(W122.^+)AzM] charge-separated state, ^3CS, whereas the ^3CT state of the other Y, F, and K122 proteins decays to the ground state. In addition to this main pathway, ^3CS is populated by fs- and ps-W(indole)→Re^(II) ET from ^1CT and the initially "hot" ^3CT states, respectively. The ^3CS state undergoes a tens-of-ns dmp.^−→W122.^+ ET recombination leading to the ground state or, in the case of the Cu^I azurin, a competitively fast (≈30 ns over 1.12 nm) Cu^I→W.^+ ET, to give [Re^I(dmp.^−)(W122)AzCu^(II)]. The overall photoinduced CuI→Re(dmp) ET through [Re^I(dmp)(W122)AzCu^I] occurs over a 2 nm distance in <50 ns after excitation, with the intervening fast ^3CT–^3CS equilibrium being the principal accelerating factor. No reaction was observed for the three Y, F, and K122 analogues. Although the presence of [Re(dmp)(W122)AzCu^(II)] oligomers in solution was documented by mass spectrometry and phosphorescence anisotropy, the kinetics data do not indicate any significant interference from the intermolecular ET steps. The ground-state dmp–indole π–π interaction together with well-matched W/W.^+ and excited-state [Re^II(CO)_3(dmp.^−)]/[Re^I(CO)_3(dmp.^−)] potentials that result in very rapid electron interchange and ^3CT–^3CS energetic proximity, are the main factors responsible for the unique ET behavior of [Re^I(dmp)(W122)]-containing azurins.