Miao_2019_J.Biol.Chem_294_6214

Lysophospholipids (LPLs) are important lipid-signaling molecules in plants, of which lysophosphatidylcholine (lysoPC) is one of the most well-characterized LPLs, having important roles in plant stress responses. It is broken down by lysophospholipases, but the molecular mechanism involved in lysoPC degradation is unclear. Recombinant Arabidopsis thaliana ACYL-CoA-BINDING PROTEIN2 (AtACBP2) has been reported to bind lysoPC via its acyl-CoA-binding domain and also LYSOPHOSPHOLIPASE 2 (AtLYSOPL2) via its ankyrin repeats in vitro To investigate the interactions of AtACBP2 with AtLYSOPL2 and lysoPC in more detail, we conducted isothermal titration calorimetry with AtACBP2(70-354), an AtACBP2 derivative consisting of amino acids 70-354, containing both the acyl-CoA-binding domain and ankyrin repeats. We observed that the interactions of AtACBP2(70-354) with AtLYSOPL2 and lysoPC were both endothermic, favored by solvation entropy and opposed by enthalpy, with dissociation constants in the micromolar range. Of note, three AtLYSOPL2 catalytic triad mutant proteins (S147A, D268A, and H298A) bound lysoPC only weakly, with an exothermic burst and dissociation constants in the millimolar range. Furthermore, the binding affinity of lysoPC-premixed AtACBP2(70-354) to AtLYSOPL2 was 10-fold higher than that of AtACBP2(70-354) alone to AtLYSOPL2. We conclude that AtACBP2 may play a role in facilitating a direct interaction between AtLYSOPL2 and lysoPC. Our results suggest that AtACBP2(70-354) probably binds to lysoPC through a hydrophobic interface that enhances a hydrotropic interaction of AtACBP2(70-354) with AtLYSOPL2 and thereby facilitates AtLYSOPL2's lysophospholipase function.