OPTICAL-ABSORPTION SPECTRA OF AZURIN AND STELLACYANIN IN GLYCEROL WATER AND ETHYLENE-GLYCOL WATER SOLUTIONS IN THE TEMPERATURE-RANGE 290-20-K

We have measured the optical absorption spectra of azurin and stellacyanin in the wavelength range 1100-350 nm and in the temperature interval 290-20 K. Samples are protein aqueous solutions containing 65% (v/v) glycerol or ethylene glycol as cryoprotectants and remain homogeneous and transparent throughout the whole temperature range investigated. Spectra are deconvoluted into Gaussian components and the temperature dependence of the zeroth, first and second moments of the various bands is analyzed, within the harmonic Franck-Condon approximation, to obtain information on the stereodynamic properties of the active sites of these proteins. Sizable differences of the integrated intensities of all the bands with temperature are observed and are attributed to variations of the metal-ligand relative positions (i.e., deformations of the active site) that occur as the temperature is lowered. The mean effective frequency of the nuclear vibrations coupled to all the observed bands is about 150 cm-1 for both proteins in both solvents used; this indicates that the electronic transitions from which the optical spectrum originates are substantially coupled with low-frequency vibrational modes, likely ligand-metal-ligand deformations. The relevance of the stereodynamic properties of azurin and stellacyanin, investigated in this work, to their functional behavior is also suggested. © 1990.