Aspartokinase I-homoserine dehydrogenase I of Escherichia coli K12 (lambda). Activation by monovalent cations and an analysis of the effect of the adenosine triphosphate-magnesium ion complex on this activation process.
The dehydrogenase activity of the aspartokinase I-homoserine dehydrogenase I complex isolated from Escherichia coli K12 is subject to a cooperative activation by K+ or Rb+, which is characterized by a Hill coefficient of approximately 2. Ionic strength has little effect on the Hill coefficient for this activation process; however, high ionic strength appears to increase the enzyme's affinity for K+ and decrease its affinity for Rb+. The Vmax of the K+-activated dehydrogenase is greater than that of the Rb+-activated dehydrogenase. The results of a study of the competition between K+ and Rb+ in the activation process suggest the presence of an activated species containing both K+ and Rb+. The cooperative activation by K+ is antagonized by Na+ via a process that is noncooperative with respect to Na+. The MgATP-2- complex, a substrate for the kinase activity of aspartokinase I-homoserine dehydrogenase I, has a marked effect on the K+ activation of the dehydrogenase activity. Kinetic studies of this effect of MgATP-2- on the K+ requirement of the dehydrogenase at pH 8.9 indicate that: (a) activation by a monovalent cation is essential in the presence as well as in the absence of MgATP-2-; (b) the concentration of K+ required to activate fully the dehydrogenase is reduced in the presence of MgATP-2-; (c) activation of the dehydrogenase by K+ is noncooperative in the presence of MgATP-2-; and (d) the maximum velocity for the dehydrogenase catalyzed oxidation of homoserine is greater in the presence of MgATP-2- than in its absence. Based on these results, a simple model consistent with these data is proposed. Destruction of the kinase activity and the threonine sensitivity of the aspartokinase-homoserine dehydrogenase complex by treatment with 5,5'-dithiobis(2-nitrobenzoic acid) or by incubation at pH 9 also converts the K+ activation of the dehydrogenase from a cooperative to a noncooperative process. Marked protection of the enzyme against loss of threonine sensitivity at pH 9 is afforded by MgATP-2- plus K+ and homoserine. The apparent molecular radius of the enzyme complex as determined by gel filtration at pH 8.85 in the presence of threonine or MgATP-2- plus K+ and homoserine is dependent on the enzyme concentration. The observed apparent molecular radii of 70 A at high enzyme concentrations and 61 A at low enzyme concentrations are consistent with the enzyme's undergoing a concentration-dependent dissociation from a tetrameric to a dimeri
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000429:Alcohol Oxidoreductases / Q000378:metabolism*; D001224:Aspartic Acid; D001565:Benzoates / Q000494:pharmacology; D001665:Binding Sites; D002794:Choline / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000201:enzymology*; D006714:Homoserine; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D008433:Mathematics; D009097:Multienzyme Complexes / Q000378:metabolism*; D009574:Nitro Compounds / Q000494:pharmacology; D009994:Osmolar Concentration; D010770:Phosphotransferases / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology*; D011485:Protein Binding; D011487:Protein Conformation; D012413:Rubidium / Q000494:pharmacology
D001565:Benzoates; D009097:Multienzyme Complexes; D009574:Nitro Compounds; D001224:Aspartic Acid; D006714:Homoserine; D000255:Adenosine Triphosphate; D000429:Alcohol Oxidoreductases; D010770:Phosphotransferases; D008274:Magnesium; D012413:Rubidium; D002794:Choline; D011188:Potassium