Reversible fast-dimerization of bovine serum albumin detected by

VALERIA LEVI, F. LUIS GONZA´LEZ FLECHA

BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS

Año: 2002 vol. 1599 p. 141 - 148

0304-4165

Self-association of bovine serum albumin (BSA) was explored using fluorescence resonance energy transfer (FRET) between twopopulations of the protein labeled separately with either fluorescein-5V-isothiocyanate (FITC) or eosin-5V-isothiocyanate (EITC). The energytransfer reached the steady state after 5 s at 25 jC, indicating a fast exchange between oligomer subunits. The dependence of the energytransfer efficiency on the protein concentration and its reversion by unlabeled BSA demonstrate that association between BSA monomersoccurs through a reversible path that involves specific interactions between the protein molecules. Because energy transfer took place evenafter blocking Cys 34 with iodoacetamide, this residue might not be involved in the reversible self-association process. The number ofsubunits forming the oligomer and its dissociation constant were determined from measurements of energy transfer as a function of thedonor–acceptor ratio and of the total protein concentration. Analysis of these data indicated that BSA is in a monomer–dimer equilibriumwith a dissociation constant of 10F2 AM at 25 jC in 10 mM MOPS-K (pH 5.8).D 2002 Published by Elsevier Science B.