IDENTIFICATION OF AUTOANTIBODY EPITOPES OF GLUTAMIC-ACID DECARBOXYLASE IN STIFF-MAN SYNDROME PATIENTS

Stiff-man syndrome is a neurologic disorder characterized by progressi ve rigidity of skeletal muscles. Deficiency of the neurotransmitter ga mma-aminobutyric acid and autoantibodies to glutamic acid decarboxylas e (GAD), the enzyme synthesizing gamma-aminobutyric acid, are closely associated with the disorder, although the relevant antigenic epitopes have not been identified. In the present study, sera from two patient s with SMS was used in an immunoblotting assay with recombinant GAD67 (M(r) 67,000) and GAD65 (M(r) 65,000) isoforms to test whether SMS ser a can recognize specific epitopes. We found that both SMS sera recogni zed the GAD65, but not the GAD67, isoform. Using 13 different syntheti c GAD peptides to block the autoantibodies, two GAD65 epitopes were id entified. One epitope recognized by both patients' sera, was blocked b y the peptide representing amino acid residues 354-368. In one patient only, blocking was also observed by a peptide representing residues 3 90-402, which includes the binding site of the GAD cofactor, pyridoxal 5'-phosphate. A single amino acid substitution in GAD65 at position 4 01 (leucine to proline) and representing the analogous GAD67 sequence in this region significantly reduced the peptide's inhibitory effect. These findings suggest that SMS GAD autoantibodies share distinct GAD6 5 linear epitopes and that some SMS patients' autoantibodies may block the active site, explaining SMS GABA deficiency.