Kinetics of lipid-membrane binding and conformational change of L-BABP

GALASSI, V. (PRIMERA AUTORIA COMPARTIDA); NOLAN, V (PRIMERA AUTORIA COMPARTIDA); VILLARREAL, M.A.; PERDUCCA, M.; MONACO, H.; MONTICH, G.G.

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

ACADEMIC PRESS INC ELSEVIER SCIENCE

Año: 2009 vol. 382 p. 771 - 775

0006-291X

We designed an experimental approach to differentiate the kinetics of protein binding to a lipid membranefrom the kinetics of the associated conformational change in the protein. We measured the fluorescenceintensity of the single Trp6 in chicken liver bile acid-binding protein (L-BABP) as a functionof time after mixing the protein with lipid membranes. We mixed the protein with pure lipid membranes,with lipid membranes in the presence of a soluble quencher, and with lipid membranes containing a fluorescencequencher attached to the lipid polar head group. We fitted simultaneously the experimentalcurves to a three-state kinetic model. We conclude that in a first step, the binding of L-BABP to the interfacialregion of the anionic lipid polar head groups occurred simultaneously with a conformationalchange to the partly unfolded state. In a second slower step, Trp6 buried within the polar head groupregion, releasing contacts with the aqueous phase