Lactose and D-galactose metabolism in Staphylococcus aureus. III. Purification and properties of D-tagatose-6-phosphate kinase.
D-Tagatose-6-phosphate kinase, an inducible enzyme that functions in the metabolism of lactose and D-galactose in Staphylococcus aureus, was purified about 300-fold from an extract of D-galactose-grown cells. The enzyme catalyzed the nucleoside triphosphate-dependent phosphorylation of both D-tagatose 6-phosphate and D-fructose 6-phosphate. Although the Vmax values were equal for these two substrates, the apparent Km values differed by 10,000-fold, being 16 micro M for D-tagatose 6-phosphate and 150 mM for D-fructose 6-phosphate. The purified enzyme was free from the constitutive D-fructose-6-phosphate kinase. Phosphoryl donors used by D-tagatose-6-phosphate kinse, listed in order of decreasing rates at saturating concentrations were GTP, UTP ITP ATP, CTP, and TTP; the Km values were 0.38, 0.91, 0.17, 0.16, 18, and 20 mM, respectively. The enzyme appeared to be nonallosteric; it exhibited Michaelis-Menten kinetics and was not inhibited by high concentrations of MgATP. However, it was activated 3- to 4-fold by 33.3 mM K+, NH4+, Rb+, and Cs+, and was inhibited 31 to 65% by 33.3 mM Na+ and Li+. It was inactivated reversibly by the thiol reagent, N-ethylmaleimide. The subunit molecular weight was estimated to be 52,000, and the native enzyme appeared to be a dimer with a sedimentation coefficient of 6.8 S. Data on stability, pH optimum, and inducibility of the enzyme are also presented.
D002414:Cations, Monovalent; D004790:Enzyme Induction; D005690:Galactose / Q000378:metabolism / Q000494:pharmacology; D006600:Hexosephosphates / Q000302:isolation & purification / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007785:Lactose / Q000378:metabolism; D008970:Molecular Weight; D010770:Phosphotransferases / Q000302:isolation & purification / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor); D013211:Staphylococcus aureus / Q000201:enzymology*; D013379:Substrate Specificity
D002414:Cations, Monovalent; D006600:Hexosephosphates; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C032258:tagatose kinase; C432824:tagatose 6-phosphate kinase; D007785:Lactose; D005690:Galactose