Molecular specificity of a medium chain acyl-CoA synthetase for substrates and inhibitors: conformational analysis.

Amino acid conjugation is an important route of detoxification of xenobiotic and endogenous carboxylic acids. The specificity of the purified medium chain acyl-CoA synthetase catalyzing the first reaction of amino acid conjugation was investigated further for substrates and inhibitors. Molecular modeling techniques were applied to derive the molecular characteristics of substrates and inhibitors for the medium chain acyl-CoA synthetase. The purified enzyme accepted not only straight medium chain fatty acids but also aromatic acids. Of the arylacetic acids, activity was obtained with naphthylacetic acids, whereas introduction of a methyl group at the alpha-position caused loss of activity. High activity was also observed with cyclohexanoic acid. Diflunisal, 2-hydroxydodecanoic acid, and nalidixic acid inhibited the medium chain acyl-CoA synthetase activity for hexanoic acid, with Ki values of 0.8, 4.4, and 12.3 microM, respectively. The inhibitory carboxylic acids were competitive with respect to hexanoic acid. The hydroxyl or ketone (oxo) groups at the beta-position of carboxylic acids were an important determinant for inhibitory activity. All substrates and inhibitors contained a flat hydrophobic region coplanar to the carboxylate group. In addition, the substrates had negative values for charge on the carbon in the beta-position of carboxylic acids.