Interaction between photoactivated rhodopsin and the C-terminal peptide of transducin alpha-subunit studied by FTIR spectroscopy.

Structural changes in the complex formed between photolyzed bovine rhodopsin and a synthetic 11-mer peptide corresponding to the C-terminal region of the transducin alpha-subunit (Gtalpha) were analyzed by means of Fourier transform infrared spectroscopy. A complex with a protonated Schiff base appears at the beginning, accompanying the formation of an alpha-helix. This complex evolves into another which abolishes the original structure but retains the protonated Schiff base. This complex exhibits the same spectral shape as that of the final stable complex with an unprotonated Schiff base. The Fourier transform infrared spectrum for the formation of this final complex was compared to that with transducin [Nishimura, S., Sasaki, J., Kandori, H., Matsuda, T., Fukada, Y., and Maeda, A. (1996) Biochemistry 35, 13267-13271]. A large part of the frequency shifts of the peptide carbonyl vibrations which form upon complex formation with transducin but are absent with the synthetic 11-mer peptide must be structural changes in other sites, such as the nucleotide binding site in Gtalpha. The peptide, like transducin, shows the perturbation of a carboxylic acid in an extremely apolar environment. Some of the changes in the peptide backbone remain in the complex formed with the peptide. These are due to sites where rhodopsin interacts with the C-terminal region of Gtalpha. Specifically, the labeling of the peptide amide corresponding to Leu349 of transducin by 15N reveals weakening of the hydrogen bond of the peptide N-H of Leu349 and/or distortion of a peptide bond between Gly348 and Leu349 upon complex formation.