ISOFORMS OF RAT-LIVER FATTY-ACID-BINDING PROTEIN DIFFER IN STRUCTURE AND AFFINITY FOR FATTY-ACIDS AND FATTY ACYL COAS

Although native rat liver fatty acid binding protein (L-FABP) is compo sed of isoforms differing in isoelectric point, their comparative stru cture and function are unknown. These properties of apo- and holo-L-FA BP isoforms were resolved by circular dichroism, time-resolved fluores cence spectroscopy, and binding/displacement of fluorescent Ligands. B oth apo-isoforms had similar hydrodynamic radii of 18.5 Angstrom, but apo-isoform I had a greater alpha-helical content and exhibited a long er Tyr lifetime, indicative of secondary and tertiary structural diffe rences from isoform II. Isoforms I and II both had two fatty acid or f atty acyl CoA binding sites. Ligand binding decreased the isoform hydr odynamic radii by 3-4 Angstrom and increased Tyr rotational motions in a more restricted range. Fatty acyl CoAs were more effective than fat ty acids in altering the isoform structures. Scatchard analysis showed that both isoforms bound cis- parinaric acid with high affinity (K-d values 41 and 60 nM, respectively) and bound trans-parinaric acid with 2- and 7-fold, respectively, higher affinity than for cis-parinaric a cid. In contrast, isoform I had higher affinity for cis- and trans-par inaroyl CoAs (K-d values of 33 and 14 nM) than did isoform II (K-d val ues of 110 and 97 nM), thereby resulting in biphasic plots of parinaro yl-CoA binding to native L-FABP. Finally, displacement studies indicat ed that each isoform displayed distinct specificities for fatty acid/f atty acyl CoA chain length and unsaturation. Thus, rat L-FABP isoforms differ markedly in both structure and ligand binding function.