Lactose and D-galactose metabolism in Staphylococcus aureus. III. Purification and properties of D-tagatose-6-phosphate kinase.
1980; Elsevier BV; Volume: 255; Issue: 18 Linguagem: Inglês
10.1016/s0021-9258(18)43563-6
ISSN1083-351X
AutoresD. Bissett, Richard L. Anderson,
Tópico(s)Enzyme Structure and Function
Resumo~-Tagatose-6-phosphate kinase, an inducible enzyme that functions in the metabolism of lactose and D-galaCtose in Staphylococcus aumus, was purified about 300fold 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 V,, values were equal for these two substrates, the apparent K,,, values differed by 10,000-fold, being 16 CM for D-tagatose 6-phosphate and 150 n m for D-fructose 6-phosphate.The purified enzyme was free from the constitutive ~-fructose-6-phosphate kinase.Phosphoryl donors used by ~-tagatose-6-phosphate kinase, listed in order of decreasing rates at saturating concentrations, were GTP, UTP, ITP, ATP, CTP, and TTP; the K,,, values were 0.38, 0.91, 0.17, 0.16, 18, and 20 m, 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 n m K', NH4', Rb', and Cs+, and was inhibited 31 to 65% by 33.3 m M Na+ and Li'.It was inactivated reversibly by the thiol reagent, Nethylmaleimide.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.The preceding paper in this series (1) described the isomerization of D-galactose 6-phosphate to D-tagatose 6-phosphate by a specific, inducible isomerase from Staphylococcus aureus.The present paper documents that the next step in the metabolism of lactose and D-galactose in this organism is the ATP-dependent phosphorylation of D-tagatose 6-phosphate to D-tagatose 1,6-diphosphate (Fig. l), and reports the purification and some properties of the kinase that catalyzes this reaction.The following article (2) describes the enzymatic aldol cleavage of D-tagatose 1,6-diphosphate.'
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