Extrinsic factors potassium chloride and glycerol induce thermostability in recombinant anthranilate synthase from Archaeoglobus fulgidus

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 ₂₂ 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°C. The kinetic constants for the enzyme under these conditions were: K_m (chorismate) 84 M, K_m (glutamine) 7.0 mM, k_cat 0.25 s^–1, and pH optimum 8.0. The enzyme was competitively, though non-cooperatively, inhibited by tryptophan.