DETECTION OF A YB3-SITE IN REGENERATED BACTERIORHODOPSIN THAT IS COORDINATED WITH THE PROTEIN AND PHOSPHOLIPID HEAD GROUPS( BINDING)

Near infrared Yb3+ vibronic sideband spectroscopy was used to characte rize specific lanthanide binding sites in bacteriorhodopsin (bR) and r etinal free bacteriorhodopsin (bO). The VSB spectra for deionized bO r egenerated with a ratio of 1:1 and 2:1 ion to bO are identical. Applic ation of a two-dimensional anti-correlation technique suggests that on ly a single Yb3+ site is observed. The Yb3+ binding site in bO is obse rved to consist of PO2- groups and carboxylic acid groups, both of whi ch are bound in a bidentate manner. An additional contribution most li kely arising from a phenolic group is also observed. This implies that the ligands for the observed single binding site are the lipid head g roups and amino acid residues. The vibronic sidebands of Yb3+ in deion ized bR regenerated at a ratio of 2:1 ion to bR are essentially identi cal to those in bO. The other high-affinity binding site is thus eithe r not evident or its fluorescence is quenched, A discussion is given o n the difference in binding of Ca2+ (or Mg2+) and lanthanides in phosp holipid membrane proteins.