Gv. Richieri et al., THERMODYNAMICS OF FATTY-ACID-BINDING TO ENGINEERED MUTANTS OF THE ADIPOCYTE AND INTESTINAL FATTY-ACID-BINDING PROTEINS, The Journal of biological chemistry, 273(13), 1998, pp. 7397-7405
We constructed 18 single amino acid mutants of the adipocyte fatty aci
d-binding protein (A-FABP) and 17 of the intestinal fatty acid-binding
protein (I-FABP), at locations in the fatty acid (FA) binding sites,
For each mutant protein, we measured thermodynamic parameters that cha
racterize FA binding, Binding affinities ranged from about 200-fold sm
aller to 30-fold larger than the wild type (WT) proteins, Thermodynami
c parameters revealed that binding affinities often inaccurately repor
ted changes in protein-FA interactions because changes in the binding
entropy and enthalpy were usually compensatory and larger than the bin
ding free energy, FA-FABP interactions were quite different for I-FABP
and A-FABP proteins, Binding affinities were larger and decreased to
a greater degree with increasing FA solubility for most of the I-FABP
as compared with the A-FABP proteins, consistent with a more hydrophob
ic binding site for the I-FABP proteins, In A-FABP, Ala substitutions
for Arg(106) and Arg(126), which interact with the FA carboxylate, red
uce affinities by about 100-fold, but in I-FABP, R106A increases affin
ities up to 30-fold, Moreover, in A-FABP, the thermodynamic parameters
predict that the FA carboxylate location switches from the 126-positi
on in R106A to the 106 position in R126A. Finally, the A-FABP proteins
, in contrast to the I-FABP proteins, reveal significant heat capacity
changes (Delta C-p) upon FA binding, and substitutions at residues Ar
g(106), and Arg(126) reduce the magnitude of Delta C-p.