Phosphoenolpyruvate pyruvate

Blsphosphoglycerate ) 3-Phosphoglycerate ( 2-Phosphoglycerate Phosphoenolpyruvate Pyruvate Lactate... 

The activity of the ATP-forming enzyme complex V is usually assessed by determining the reverse reaction ATP — ADP + Pi. The reaction is coupled to reactions catalyzed by pyruvate kinase (ADP + phosphoenolpyruvate —> pyruvate + ATP) and lactate dehydrogenase (pyruvate + NADH — lactate -F NAD+). This final reaction can be followed spectrophotometrically by measuring NADH at 340 nm. The activity of complex V (ATPase) can be derived from the rate of NADH conversion in the presence and absence of the specific complex V inhibitor oligomycin. 

In practice, for most synthetic applications, either acetyl phosphate/acetate kinase or phosphoenolpyruvate/pyruvate kinase are used to regenerate ATP. Because of the ease of preparing AcP, AcP/AcK is the most economical method for large-scale work. Its application is, however, limited to fast phosphorylation reactions where the hydrolysis of AcP is not important. The PEP/pyruvate kinase system is used in instances where the requirement for a strong, stable phosphorylating reagent outweighs the relative inconvenience of preparation of PEP. 

Phosphoenolpyruvate/pyruvate kinase. Phosphoenolpyruvate (PEP 2)/pyruvate kinase (PK E.C. 2.7.1.40) is the most efficient system for the regeneration of ATP from ADP. The phosphorylating agent PEP can be prepared in a mole scale [47L Starting from crude pyruvic acid, the crystalline monopotassium salt PEP-K is synthesized in a three-step procedure. For transformations on a scale <1 mol, PEP can be prepared from commercially available 3-phosphoglyceric acid in an enzyme-catalyzed reaction1641. This method is more expensive than the chemical preparation, but is more convenient because it requires less time and produces less organic waste (see Section 13.2.1.2 Fig. 13-5). 

Investigations on primary active transport are also performed with vesicles. For inside-out vesicles no additional experimental problems arise, since the enzymatic site of the transport system located at the cytoplasmic site of the membrane is freely accessible to the substrate of the exergonic driving reaction from the incubation medium. Right-side-out vesicles, however, have to be prepared in the presence of the driver substrate, e.g. ATP, or a system for its regeneration, e.g. ATP/ADP + phosphoenolpyruvate + pyruvate kinase. 

Elucidation of the metabolic pathway repertoire of C, glutamicum was initiated soon after its discovery [4-6], related to the high importance of the central metabolism for amino acid fermentation. Evidence on the presence of the major catabolic routes, such as the Embden-Meyerhof-Parnas (EMP) pathway, the pentose phosphate (PP) pathway, the tricarboxylic acid (TCA) cycle, and the glyoxylate shunt had already been provided by the end of the 1950s [4, 5, 7] (Figure 6.1). However, it took more than 30 years for a more detailed resolution of the central metabolic network mainly related to the complex structure of the phosphoenolpyruvate/pyruvate - oxaloacetate/malate node [8-11]. Altogether,... 

Alcohols and carboxylic acids acetate, ascorbate, benzoate, benzoquinone, catechol, chlorophenol, ethanol, fatty acids, formate, glycerol, gluconate, glycolate, glyoxylate, hexanol, hydroquinone, hy-droxybutyrate, isocitrate, lactate, malate, methanol, oxalate, phenol, phosphoenolpyruvate, pyruvate, succinate, salicylate. 

Phosphoenolpyruvate, pyruvate, acetyl coenzyme A, and a number of amino acids involved in the biosynthesis of cyanogenic glycosides, alkaloids, glucosinolates, and antibiotics are linked to the final portion of the glycolytic sequence and immediately prior to entry into the Krebs cycle. A shift from normal sustained growth to that in which there is a decreased uptake of acetyl coenzyme A in the Krebs cycle may reflect a diminished requirement for nucleotide triphos-... 

In these assays, the initial reaction product is colourless, but a second enzyme is included to convert this into a product measurable by spectrophotometry. For example, for an ATPase assay, the product ADP can be used to drive NADH oxidation (measurable by A340) in the presence of phosphoenolpyruvate + pyruvate kinase (which generate pyruvate in the presence of ADP) and lactate dehydrogenase (which converts NADH to NAD in the presence of p30uvate). 

Anaplerotic reactions refer to C3-carboxylation and C4-decarboxylation around the phosphoenolpyruvate-pyruvate-oxaloacetate node, which interconnect the TCA cycle with glycolysis. These reactions result in direct oxaloacetate formation or depletion. Carboxylation of phosphoenolpyruvate catalyzed by phosphoenolpyruvate carboxylase and that of pyruvate by pyruvate carboxylase contribute to its formation. Accordingly, decarboxylation of oxaloacetate catalyzed by phosphoenolpyruvate carboxykinase and oxaloacetate decarboxylase form phosphoenolpyruvate and... 

Pyruvic acid is important as an intermediate in sugar metabolism. This acid is formed as the final product of the glycolytic pathway from 3-phosphoglycerate via phosphoenolpyruvate. Pyruvic acid is also formed through the oxidative pentose phosphate cycle. The degradation of glucose in the cycle yields the C3 product glyceraldehyde 3-phosphate, which can be oxidized to pyruvate. Pyruvic acid is the principal precursor for the biosynthesis of amino acids such as alanine, as well as leucine and valine. 

The carbon flow from 3-phosphoglycerate, phosphoenolpyruvate, pyruvate and acetyl-CoA. Even if the synthesis of aromatic amino acids by shikimate pathway /28,29,30,31/ and also prenyl-PP synthesis via mevalonate /32,33,34/ has been established in chloroplasts by identification of respective plastidic enzymes, it is still a matter of discussion from where PEP origins to supply DAHP synthesis of the shikimate pathway and from where pyruvate is delivered to supply the plastidic pyruvate dehydrogenase complex (for isolation see Treede and Heise, this Conference). Because phosphoglycerate mutase (PGM) to form 2-PGA from 3-PGA could not be detected in chloroplasts /35/ and acetyl-CoA is preferably synthesized from added acetate by the actetyl-CoA synthetase /36/, particularly in spinach chloroplasts, it was argued that chloroplasts are dependent on import of these substrates from the external site. Evidence for PEP formation from 3-PGA within the chloroplast could be obtained by three different approaches (D. Schulze-Siebert, A. Heintze and G. Schultz, in preparation D. Schulze-Siebert and G. Schultz, in preparation, for plastidic isoenzyme of PGM in Ricinus see /37/ and in Brassica /38/). 

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