SITE-DIRECTED ISOTOPE LABELING AND FTIR SPECTROSCOPY OF BACTERIORHODOPSIN

Insight into Integral membrane proteins function is presently limited by the difficulty of producing three-dimensional crystals. In addition , X-ray structures Of proteins normally do not provide information abo ut the protonation state and structural changes of individual residues , We report here the first use of Site-directed isotope labelling and Fourier transform infrared (FTIR) difference spectroscopy to detect st ructural changes at the level of single residues in an Integral membra ne protein. Two site-directed Isotope labeled (SDIL) tyrosine analogue s of bacteriorhodopsin were produced which exhibit normal activity. FT IR spectroscopy shows that out of 11 tyrosines, only Tyr 185 is struct urally active during the early photocycle and may be part of a proton wire.