Adipocyte lipid-binding protein complexed with arachidonic acid. Titration calorimetry and X-ray crystallographic studies

J. M. LaLonde, M. A. Levenson, J. J. Roe, D. A. Bernlohr, L. J. Banaszak

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Abstract

The association of the adipocyte lipid-binding protein (ALBP) with arachidonic acid (all cis, 20:4 Δ5,8,11,14) and oleic acid (cis, 18:1 Δ9) has been examined by titration calorimetry. In addition, the crystal structure of ALBP with bound arachidonic acid has also been obtained. Crystallographic analysis of the arachidonic acid-ALBP complex along with the previously reported oleic acid-ALBP structure (Xu, Z., Bernlohr, D. A., and Banaszak, L. J. (1993) J. Biol. Chem. 268, 7874-7884) provides a framework for the molecular examination of protein-lipid association. Isothermal titration calorimetry revealed high affinity association of both unsaturated fatty acids with the protein. The calorimetric data yielded the following thermodynamic parameters for arachidonic acid: K(d) = 4.4 μM, n = 0.8, ΔG = -7370 cal/mol, ΔH = -6770 cal/mol, and TΔS = +600 cal/mol. For oleic acid, the thermodynamic parameters were K(d) = 2.4 μM, n = 0.9, ΔG = -7770 cal/mol, ΔH = -6050 cal/mol, and TΔS = +1720 cal/mol. The identification of thermodynamically dominating enthalpic factors for both fatty acids are consistent with the crystallographic studies demonstrating the interaction of the fatty acid carboxylate with a combination of Arg106, Arg126, and Tyr128. The crystallographic refinement of the protein-arachidonate complex was carried out to 1.6 Å with the resultant R factor of 0.19. Within the cavity of the crystalline binding protein, the arachidonate was found in a hairpin conformation. The conformation of the bound ligand is consistent with acceptable torsional angles and the four cis double bonds in arachidonate. These results demonstrate that arachidonate is a ligand for ALBP. They provide thermodynamic and structural data concerning the physical basis for protein-lipid interaction and suggest that intracellular lipid- binding proteins may mediate the biological effects of polyunsaturated fatty acids in vivo.

Original languageEnglish (US)
Pages (from-to)25339-25347
Number of pages9
JournalJournal of Biological Chemistry
Volume269
Issue number41
StatePublished - 1994

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