Ultraviolet Resonance Raman Study on Protein Structure Dynamics of Cytochrome c Oxidase

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A high performance ultraviolet resonance Raman spectrophotometer for biological applications has been constructed. A solar-blind intensified charge-coupled device was attached to a 1260 mm single spectrograph with a 200 nm-blazed, 3600 grooves/mm grating. The 488 nm output of a mode-lacked argon ion laser was frequency doubled to obtain 244nm UV light for resonance Raman excitation. This Raman spectrophotometer has been applied to cytochrome c oxidase to probe structural changes of tyrosine and tryptophan residues upon redox and ligation-state changes. Resting oxidized, fully-reduced and mixed-valence forms of the enzyme in the presence and absence of carbonmonoxide or cyanide have been measured. It became evident that a redox change of heme a and / or Cu_A induced most prominent changes on tyrosine and tryptophan residues while ligation to heme a_3 and redox change of heme a_3 and / or Cu_B less affected the protein structures. Basic data to analyze time-resolved UV resonance Raman spectra of cytochrome c oxidase have been established.