Accumulation of free tryptophan in azuki bean (Vigna angularis) induced by expression of a gene (OASA1D) for a modified α-subunit of rice anthranilate synthase

Anthranilate synthase (AS) is a key metabolic enzyme of the tryptophan (Trp) biosynthetic pathway in plants and microbes, and its activity is regulated by feedback inhibition by Trp. A rice gene (OASA1D) for a modified α-subunit of anthranilate synthase that is resistant to such feedback inhibition was introduced into azuki bean [Vigna angularis (Willd.) Ohwi & Ohashi] by Agrobacterium-mediated transformation followed by selection with hygromycin B. Five independent azuki bean lines expressing OASA1D in which the transgene was fixed in the T2 generation were obtained. The free Trp level in seeds of these lines (1.3–3.3 mg/g of dry weight) was increased compared with that in wild-type seeds (0.2 mg/g). In contrast, the amount of protein-bound (fixed) Trp in seeds was not affected by expression of the transgene. The AS activity in the transgenic seeds showed a reduced sensitivity to feedback inhibition by Trp or the Trp analog 5-methyltryptophan, indicating that the rice OASA1D subunit was able to interact functionally with the β-subunit of azuki bean AS. These results reveal that transformation with OASA1D is a feasible approach to increase Trp synthesis and accumulation in mature seeds of grain legumes.