Preparation of domoic acid analogues using a bioconversion system, and their toxicity in mice.

Domoic acid (1, DA), a member of the natural kainoid family, is a potent agonist of ionotropic glutamate receptors in the central nervous system. The chemical synthesis of DA and its derivatives requires considerable effort to establish a pyrrolidine ring containing three contiguous stereocenters. Recently, a biosynthetic cyclase for DA, DabC, was identified. This enzyme cyclizes the linear precursor of isodomoic acid A (IA) to IA, a bioactive DA analogue. In this study, we developed a bioconversion system to obtain DA analogues from linear substrates prepared by simple chemical synthesis using DabC expressed in Escherichia coli , in vivo . Three IA analogues with various substitutions at the C7'-geranyl terminus were prepared using this system: two minor natural analogues, 7'-methyl-IA (5) and 7'-hydroxy-IA (6), and one new unnatural analogue, 7'-amide-IA (7). In addition, the toxicity of these DA analogues in mice was examined by intracerebroventricular injection. Most of the mice injected with 5 (3 nmol) and 6 (3 nmol) did not show any adverse symptoms, whereas the mice injected with 7 (3 nmol) showed typical symptoms induced by DA (1, 0.7 nmol) and IA (2, 3 nmol). These results suggest that the 7'-carbonyl group in the side chain of IA (2) is crucial for its toxicity. The docking studies of DA, IA (2), 5, 6, and 7 to GluK1 supported these results.