Subunit structure of anthranilate synthase from Neurospora crassa: Preparation and characterization of a protease-free form

The multifunctional enzyme complex anthranilate synthase from Neurospora crassa has been purified to homogeneity by a new procedure which yields a stable preparation of the enzyme. Unlike earlier preparations of the enzyme, anthranilate synthase prepared by this technique is not degraded during incubation at 37 °C or during freeze-thaw treatment. Purified anthranilate synthase contains two subunits of M_r 84,000 (β-subunit) and 76,000 (α-subunit), which are shown, by partial proteolysis, to be unrelated in sequence. Immunoprecipitation studies demonstrate that freshly prepared crude extracts of Neurospora contain anthranilate synthase subunits identical in size with those of the purified enzyme. The β-subunit is shown to be the product of the trp1 gene, and the a-subunit, of the trp2 gene.