title: Extrinsic factors potassium chloride and glycerol induce thermostability in recombinant anthranilate synthase from Archaeoglobus fulgidus.
abstract: 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.
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, Howard University, 520 W. Street, N.W., Washington, DC 20059, USA. wbyrnes@howard.edu
authors.forename: W Malcolm
authors.initials: WM
authors.lastname: Byrnes
authors.forename: Vincent L
authors.initials: VL
authors.lastname: Vilker
mesh_terms: D000878:Anthranilate Synthase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019611:Archaeoglobus fulgidus / Q000201:enzymology* / Q000235:genetics; D004795:Enzyme Stability / Q000187:drug effects; D019766:Genes, Archaeal; D005990:Glycerol / Q000494:pharmacology; D007700:Kinetics; D008970:Molecular Weight; D011189:Potassium Chloride / Q000494:pharmacology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D014364:Tryptophan / Q000494:pharmacology
chemical_list: D011994:Recombinant Proteins; D011189:Potassium Chloride; D014364:Tryptophan; D000878:Anthranilate Synthase; D005990:Glycerol