INSIGHT INTO THE ENVIRONMENT OF TRYPTOPHAN IN A HYDROPHOBIC MODEL PEPTIDE UPON AGGREGATION AND INTERACTION WITH LIPID VESICLES - A STEADY-STATE AND TIME-RESOLVED FLUORESCENCE STUDY

Fluorescence spectroscopy is extensively used to monitor binding of pe ptides to lipid vesicles as well as orientation in the lipid bilayer. in steady-state fluorescence, the emission characteristics of intrinsi c and extrinsic fluorophores, which are sensitive to environment are m onitored. Life time measurements should yield useful information about the location and flexibility of fluorophores, as these factors have a significant effect on the life times. However, studies on protein str ucture and dynamics indicate that interpretation of life-time data is complicated (Beechem. J.M. and Brand, L. (1985) Annu. Rev. Biochem. 54 , 43-71). Hence, simple well-defined systems should help in interpreta tion of life time data, especially in lipid-peptide interactions. In o rder to examine how fluorescence characteristics of tryptophan and ant hroyl group would reflect molecular details of peptide aggregation and lipid-peptide interaction, studies have been carried out on a model h ydrophobic peptide and its fatty acylated derivative. Steady-state flu orescence measurements suggest that: (1) The fatty acyl chain attached to an amino acid associates with the peptide chain in aqueous environ ment. (2) In the lipid bilayer, the acyl chain is oriented perpendicul ar to the lipid bilayer surface with the peptide chain at an angle to it. Analysis of the fluorescence decay of tryptophan indicates the pre dominance of a very short life-time component (<1ns) in aqueous enviro nment and lipid-vesicles. Since the preexponentials were not negative, it is unlikely that this is due to extensive deactivation process. We attribute the observation of the low life time component to predomina nce of one rotamer around (C-alpha-C-beta) bond of tryptophan in aqueo us and lipid environments. Our investigations suggest that fluorescenc e life time data need to be complemented with steady state measurement s to get an insight into details of lipid-peptide interaction.