THE BINDING OF NATURAL AND FLUORESCENT LYSOPHOSPHOLIPIDS TO WILD-TYPE AND MUTANT RAT LIVER FATTY ACID-BINDING PROTEIN AND ALBUMIN

摘要

Rat liver fatty acid-binding protein (FABP) is able to bind a wide range of non-polar anionic ligands, including lysophospholipids. In order to understand the nature of lysophospholipid interactions with liver FABP, the binding of natural lysophospholipids and two fluorescent analogues, N-(5-dimethylaminonaphthalenesulphonyl)-1-palmitoyl-sn-glycero-3-phosphoet hanolamine (dansyl lysoPE) and 1-{O-[11-(5-dimethylaminonaphthalenesulphonyl)amino]undecyl}-sn- glycero-3-phosphoethanolamine (dansyl-C-11-lysoPAF), has been investigated. The results confirmed the ability of liver FABP to bind lysophospholipids with K-D values in the range of 1-2 mu M, and a 1:1 binding stoichiometry was indicated. Binding of fluorescent lysophospholipids was enhanced with the FABP mutant, R122Q, possibly due to increased flexibility of the binding cavity as a result of reduced hydrogen-bonding constraints. The fluorescent lysophospholipids also bound to albumin, with K-D values in the range 0.1-1.0 mu M, and could be displaced by oleic acid. The fluorescence characteristics of the dansyl lysophospholipid analogue dansyl-C-11-lyso-PAF suggested that this probe binds to the same site(s) on albumin as the fluorescent fatty acid probe 11-(5-dimethylaminonaphthalenesulphonylamino)-undecanoic acid (DAUDA).