Abstract
The role of tyrosines in the bacteriorhodopsin (bR) photocycle has been investigated by using Fourier transform infrared (FTIR) and UV difference spectroscopies. Tyrosine contributions to the BR/sub 570/ ..-->.. M/sub 412/ FTIR difference spectra recorded at several temperatures and pH's were identified by isotopically labeling tyrosine residues in bacteriorhodopsin. The frequencies and deuterium/hydrogen exchange sensitivities of these peaks and of peaks in spectra of model compounds in several environments suggest that at least two different tyrosine groups participate in the bR photocycle during the formation of M/sub 412/. One group undergoes a tyrosinate ..-->.. tyrosine conversion during the BR/sub 570/ ..-->.. K/sub 630/ transition. A second tyrosine group deprotonates between L/sub 550/ and M/sub 412/. Low-temperature UV difference spectra in the 220-350-nm region of both purple membrane suspensions and rehydrated films support these conclusions. The UV spectra also indicate perturbations(s) of one or more tryptophan group(s). Several carboxyl groups appear to undergo a series of protonation changes between BR/sub 570/ and M/sub 412/, as indicated by infrared absorption changes in the 1770-1720-cm/sup -1/ region. These results are consistent with the existence of a proton wire in bacteriorhodopsin that involves both tyrosine and carboxyl groups.