Stark spectroscopic studies of blue copper proteins: Azurin

The change in the dipole moments (\Delta mu\) and the average change in the polarizability ((Delta alpha)) upon excitation for the Cys(S)-->Cu(II) lig and-to-metal charge-transfer (LMCT) transitions in two species of azurin, a type I blue copper protein from Alcaligenes denitrificans (AD) and Pseudom onas aeruginosa (PA), were determined using Stark (electroabsorption) spect roscopy. Measurements at 77 K in a glycerol-water glassy matrix yield a val ue for \Delta mu\ of between 1.3 and 2.0 D. This value of \Delta mu\ is con sistent with the highly covalent nature of the S-Cu bond in the sound state and is in agreement with the predictions of previous electronic structure calculations. The polarizability of the excited state was smaller than the ground state by about 10-20 Angstrom (3). This negative value of (Delta alp ha), which is somewhat unusual, is interpreted in the context of the two-st ate model. Values for the electron-transfer matrix element (H-ab) and the e ffective charge-transfer distance (Rab) derived from our measurements are a lso reported.