Modification of the fatty acid binding profile of liver fatty acid binding protein (L-FABP)

Abstract

Low molecular-weight fatty acid binding protein (L-FABP) was purified from rat liver by a combination of gel filtration and affinity chromatography. The purified protein had a molecular weight of 14,000 Daltons as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fast protein liquid chromatography (FPLC) gel filtration chromatography. Isoelectric focusing of delipidated preparations gave a major protein band with a pl of 7.5. Delipidated L-FABP was used to determine binding constants for individual saturated, monounsaturated, and polyunsaturated fatty acids. For fatty acids of chain length greater than C14 there was no apparent selectivity based on chain length or degree of unsaturation. When delipidated L-FABP was incubated with an equimolar (0.3 mmol/L) mixture of fatty acids; 16:0, 18:1 (n-9), 18:2(n-6), 20:3(n-6), 20:4(n-6), and 22:6(n-3) were bound at equivalent levels (0.18 mol/mol L-FABP). However, stearic acid was bound to a greater extent (approximately two fold) and 18:3(n-6) and 18:3(n-3) were bound to a lesser extent (50%). 12:0, 14:0, and 20:5(n-3) bound poorly to L-FABP. Thus, under these conditions fatty acid binding protein exhibits a selectivity that is not apparent from individual binding constants. When rats were maintained on different diets for 6 weeks, the concentration of L-FABP was reduced only in animals maintained on a fat-free diet. The apparent binding capacity of L-FABP (0.71 mol fatty acid per mol L-FABP) was the same for all diets. However, the endogenous fatty acid composition of L-FABP was strongly influenced by dietary fatty acid composition.