Abstract
Solid-state super(13)C MAS NMR spectra were obtained for dark-adapted bacteriorhodopsin (bR) labeled with (4'- super(13)C)Tyr. Difference spectra (labeled minus natural abundance) taken at pH values between 2 and 12, and temperatures between 20 and -90 degree C, exhibit a single signal centered at 156 ppm, indicating that the 11 tyrosines are protonated over a wide pH range. However, at pH 13, a second line appears in the spectrum with an isotropic shift of 165 ppm. Comparisons with solution and solid-state spectra of model compounds suggest that this second line is due to the formation of tyrosinate. Integrated intensities indicate that about half of the tyrosines are deprotonated at pH 13.