Thermostable anthranilate synthase from the marine sulfate-reducing hyperthermophile Archaeoglobus fulgidus has been expressed in Escherichia coli, purified, and characterized. The functional enzyme is an alpha2beta2 heterotetrameric complex of molecular mass 150+/-15 kDa. It is composed of two TrpE (50 kDa) and two TrpG (18 kDa) subunits. The extrinsic factors glycerol (25%) and potassium chloride (2 M) stabilized the recombinant enzyme against thermal inactivation. In the presence of these extrinsic factors, the enzyme was highly thermostable, exhibiting a half-life of thermal inactivation of about 1 h at 85 degrees C. The kinetic constants for the enzyme under these conditions were: Km (chorismate) 84 microM, Km (glutamine) 7.0 mM, kcat 0.25 s(-1), and pH optimum 8.0. The enzyme was competitively, though non-cooperatively, inhibited by tryptophan.