In Clostridium thermocellum, a thermophilic anaerobic bacterium able to rapidly ferment cellulose to ethanol, pyruvate kinase (EC 2.7.1.40) is absent based on both the genome sequence and enzymatic assays. Instead, a new pathway converting phosphoenolpyruvate to pyruvate via a three-step pathway involving phosphoenolpyruvate carboxykinase, NADH-linked malate dehydrogenase, and NADP-dependent ma...

Kinetic properties of rat liver pyruvate kinase type I at pH7.5 and 6.5 were studied with physiological ranges of substrates, modifiers and Mg(2+) concentrations at increasing enzyme concentrations, including the estimated cellular concentrations (approx. 0.1mg/ml). Enzyme properties appear unaffected by increased enzyme concentration if phosphoenolpyruvate, fructose 1,6-diphosphate and inhibit...

l h e pyruvate dehydrogenase kinase-catalyzed inactivation of the pyruvate dehydrogenase complex was studied using dialyzed, soluble proteins from mitochondria purified from green leaf tissue of Pisum sativum 1. seedlings. At subsaturating ATP concentrations, K+ or NH4+, but not Na+, stimulated the pyruvate dehydrogenase kinase by lowering the K,,,(ATP). Micromolar concentrations of NH4+ were r...

In isolated rat adipocytes, the insulin stimulation of pyruvate dehydrogenase can be partially inhibited by inhibitors of PI3K (phosphoinositide 3-kinase) and MEK1/2 (mitogen-activated protein kinase/extracellular signal-regulated kinase kinase). In combination, U0126 and wortmannin completely block the insulin stimulation of pyruvate dehydrogenase. It is concluded that the effect of insulin on...

Endogenous kinase activity of highly purified pyruvate dehydrogenase complex from bovine kidney is markedly inhibited by N-ethylmaleimide and by certain disulfides. Inhibition by disulfides is highly specific and is reversed by thiols. 5,5'-Dithiobis(2-nitrobenzoate) is the most potent inhibitor, showing significant inhibition at a concentration as low as 1 microM. Cystamine, oxidized glutathio...

and optical properties of rabbit muscle pyruvate kinase activated with Co(II), Ni(II), Mg(II), and Mn(I1) were studied. The stoichiometry of metal binding to enzyme was found to be 4 metal ions per tetrameric enzyme for Co(I1) and Ni(I1) by carrying out circular dichroic titrations. Cu(I1) and Fe(I1) were inactive. Ca(I1) and Zn(I1) were not activating, and were inhibitory with respect to all o...

When cod fish muscle oxalacetate decarboxylase catalyzes the decarboxylation of oxalacetate in the presence of NaBH4, L-lactate results from the reduction of enzyme-bound pyruvate. However, D-lactate results when borohydride reduces the binary enzyme-pyruvate complex formed by adding pyruvate from solution, as reported by others. This observation suggests that there are alternate mechanisms for...

This paper presents a simple solution to the problem of approximating the calculated curve of reaction progress to the measured curve which is usually disturbed by initial oscillation of auxiliary lactate dehydrogenase (LDH) reaction. The experiments leading to the determination of the apparent Km for phosphoenolpyruvate (PEP) and Vm were performed. For precise estimation of kinetic parameters ...

Control of gluconeogenesis from lactate was studied by titrating rat liver cells with lactate and pyruvate in a ratio of 1 O : l in a perifusion system. At different steady states of glucose formation, the concentration of key gluconeogenic intermediates was measured and plotted against gluconeogenic flux (Jglucoee). Complete saturation was observed only in the plot relating JglUcto the extrace...