PROTEIN TUNING OF EXCITED-STATE CHARGE-TRANSFER DYNAMICS IN AZURIN

Resonance Raman spectra of azurin from Alcaligenes denitrificans have been measured at wavelengths throughout the S(Cys) --> Cu charge-trans fer absorption band to examine the effects of structure and environmen t on the excited-state charge-transfer dynamics. Measurement of the de polarization ratios suggest that the Raman scattering is enhanced by a single electronic transition. Self-consistent analysis of the absorpt ion spectrum and the resonance Raman fundamental, overtone, and combin ation bands yields a total inner-sphere reorganization energy of 0.26 +/- 0.02 eV, similar to that for Pseudomonas aeruginosa. The reorganiz ation energy is distributed along similar vibrational modes in the two azurins, although the contribution from each mode is different. To un derstand these differences, molecular comparisons of the known structu res and molecular modeling of the electrostatic field within the prote in were performed. The results show that the two proteins have very si milar structures, although the copper site itself has a slightly more tetrahedral geometry in A. denitrificans. The resonance Raman spectral differences between these two azurins are interpreted to arise from t his copper site structural difference and a greater distribution of ch arges and/or dipoles near the coordinating histidines in A. denitrific ans.