BIOTIN AND BIOTIN ANALOGS SPECIFICALLY MODIFY THE FLUORESCENCE DECAY OF AVIDIN

Avidin, a basic tetrameric glycoprotein, isolated from hen egg-white, binds up to four molecules of biotin with exceptionally high affinity. The presence of tryptophanyl residues in the active site pointed out the opportunity of correlating the protein fluorescence with biotin bi nding. We have performed both steady state and dynamic fluorescence ex periments using biotin or biotin-derived molecules (biotinamine, diami nobiotin and iminobiotin) as ligands. The fluorescence decay data can only be fitted by two continuous distributions of lifetimes which may reflect the presence of static or dynamic microheterogeneity in the en vironment of the tryptophan residues. We observed that the binding of biotin, biotinamine and iminobiotin reduces the widths of both distrib utions to discrete lifetimes thus indicating a more homogenous environ ment for the emitting tryptophan residues. Instead, the binding of dia minobiotin, which lacks the imidazolone ring, affects one lifetime dis tribution only. The binding of biotin also affects the rotational corr elation time of avidin, which becomes shorter, suggesting a more compa ct structure of the ligated protein. The utility of analyzing the fluo rescence in terms of distributions appears to be further warranted.