Phosphoenolpyruvate synthetase

Phosphoenolpyruvate Synthetase and Pyruvate, Phosphate Dikinase R. A. Cooper and H. L. Komberg... 

The existence and importance of the covalent intermediates that appear in the few such reactions in which they are involved can be rationalized in a few ways. Most can be rationalized on the basis of the principle of economy in the evolution of binding sites. In the cases of the pyrophosphoenzymes and phosphoenzymes that are intemediates in the phosphoenolpyruvate synthetase and pyruvate phosphate dikinase reactions, the same principle applies however, additional interactions are probably more important. In one instance of a phosphoenzyme, the succinyl-CoA synthetase reaction, the role of the phosphoenzyme is unclear. But even in that case an attractive rationale can be advanced as a hypothesis. 

The problem of focusing phosphorylation free energy is elegantly solved in the mechanisms of action of phosphoenolpyruvate synthetase and pyruvate phosphate dikinase. Phosphohistidyl residues are generated at the active sites of these enzymes by pyrophosphorylation of active site histidines, followed by hydrolysis... 

Phosphoenolpyruvate synthetase (PEPS), 15 Phosphoglucomutase, 1664 Phospholipase, 2676 Phospholipase-2 (PLA2), 2679 Phospholipase A2 (iPLA2), 3966 Phospholipid bilayer, 2471 Phospholipids, 993, 1277 Phosphoribosyltransferase (PRT), 1269... 

Gene activated Lipoprotein lipase fatty acid transporter protein adipocyte fatty acid binding protein acyl-CoA synthetase malic enzyme GLUT-4 glucose transporter phosphoenolpyruvate carboxykinase... 

This enzyme [EC 4.1.2.16] (also known as phospho-2-dehydro-3-deoxyoctonate aldolase, phospho-2-keto-3-deoxyoctonate aldolase, and 3-deoxy-D-manno-octulo-sonic acid 8-phosphate synthetase) catalyzes the reaction of 2-dehydro-3-deoxy-D-octonate 8-phosphate and orthophosphate to produce phosphoenolpyruvate, D-arabinose 5-phosphate, and water. 

Enzymatic synthesis relying on the use of aldolases offers several advantages. As opposed to chemical aldolization, aldolases usually catalyze a stereoselective aldol reaction under mild conditions there is no need for protection of functional groups and no cofactors are required. Moreover, whereas high specificity is reported for the donor substrate, broad flexibility toward the acceptor is generally observed. Finally, aldolases herein discussed do not use phosphorylated substrates, contrary to phosphoenolpyruvate-dependent aldolases involved in vivo in the biosynthetic pathway, such as KDO synthetase or DAHP synthetase [18,19]. 

In this one-pot procedure NeuAc 16 is generated from ManNAc 15 and pyruvic acid in situ with sialic acid aldolase and then converted irreversibly to CMP-NeuAc 17. CMP is converted to CDP with myokinase and ATP. The released ADP is converted to ATP with pyruvate kinase and PEP. CDP is then converted to CTP also with pyruvate kinase and phosphoenolpyruvate (PEP). The formed CTP reacts with NeuAc catalyzed by NeuAc synthetase to give 17. 

Muscolo, A., Panuccio, M. R., Abenavoli, M. R., Concheri, G, and Nardi, S. (1996). Effect of molecular complexity and acidity of earthworm faeces humic fractions on glutamate dehydrogenase, glutamine synthetase and phosphoenolpyruvate carboxylase in Daucus carota II cells. Biol. Fertil. Soils 22, 83-88. 

Available evidence (14,15) favors the pathway for pyruvate kinase by way of phosphorylation of pyruvate enol. Furthermore, J. Knowles and his coworkers (16,17), using chiral thiophosphates and chiral (160,170,180) phosphate have shown that pyruvate kinase transfers phosphate from phosphoenolpyruvate to ADP with stereochemical inversion at phosphorus. Since monomeric metaphosphate is presumably planar, a chemical reaction by way of that ion should proceed with racemization. In the active site of an enzyme, however, all components might be held so rigidly that racemization need not occur. Furthermore, no information is yet available on the detailed mechanism of reactions catalyzed by cytidine synthetase our own experiments, designed to distinguish among the mechanisms here discussed, are as yet incomplete. 

Other pyruvate- and phosphoenolpyruvate-dependent aldolases have been isolated and purified, but have not yet been extensively investigated for synthetic use. Those showing promise for future applications include, 3-deoxy-D-arabino-2-heptulosonic acid 7-phosphate (DAHP) synthetase (EC 4.1.2.15), 2-keto-4-hydroxyglutarate (KHG) aldolase (EC 4.1.2.31), and 2-keto-3-deoxy-D-gluconate (KDG) aldolase (EC 4.1.2.20). DAHP synthetase has been used... 

Biotin acts to induce glucokinase, phosphofructokinase, and pyruvate kinase (key enzymes of glycolysis), phosphoenolpyruvate carboxykinase (a key enzyme of gluconeogenesis), and holocarboxylase synthetase, acting via a cell-surface receptor linked to formation of cGMP and increased activity of RNA polymerase. The activity of holocarboxylase synthetase (Section 11.2.2) falls in experimental biotin deficiency and increases with a parallel increase in... 

Comprehensive Biological Catalysis—a Mechanistic Reference Volume has recently been published. The fiiU contents list (approximate number of references in parentheses) is as follows S-adenosylmethionine-dependent methyltransferases (110) prenyl transfer and the enzymes of terpenoid and steroid biosynthesis (330) glycosyl transfer (800) mechanism of folate-requiring enzymes in one-carbon metabohsm (260) hydride and alkyl group shifts in the reactions of aldehydes and ketones (150) phosphoenolpyruvate as an electrophile carboxyvinyl transfer reactions (140) physical organic chemistry of acyl transfer reactions (220) catalytic mechanisms of the aspartic proteinases (90) the serine proteinases (135) cysteine proteinases (350) zinc proteinases (200) esterases and lipases (160) reactions of carbon at the carbon dioxide level of oxidation (390) transfer of the POj group (230) phosphate diesterases and triesterases (160) ribozymes (70) catalysis of tRNA aminoacylation by class I and class II aminoacyl-tRNA synthetases (220) thio-disulfide exchange of divalent sulfirr (150) and sulfotransferases (50). 

Scheme L Synthesis of a2,64inked sialyl-N-acetyllactosamine using a one-pot multi-enzyme system with in situ regeneration of CMP-Neu5Ac. Abbreviations for enzymes CSS, CMP-sialic acid synthetase NMK, nucleoside monophosphate kinase PK, pyruvate kinase PPase, pyrophosphatase. Abbreviations for compounds PEP, phosphoenolpyruvate ADP, adenosine 5 -diphosphate ATP, adenosine 5 -triphosphate CMP, cytidine 5-monophosphate CDP, cytidine 5 -diphosphate CTP, cytidine 5-triphosphate LacNAc, N-acetyllactosamine NeuSAc, N-acetylneuraminic acid PPi, inorganic pyrophosphate.

The cascade begins with stoichiometric amounts of phosphoenolpyruvate (PEP), 8-allyl-A-acetyl lactosamine 120, NeuAc 1, and catalytic quantities of ATP and CMP. Initially, CMP is converted to CDP by nucleoside monophosphate kinase (NMK) in the presence of ATP. The CDP produced reacts with PEP under pyruvate kinase (PK) catalysis to form CTP. Next, CMP-NeuAc synthetase catalyzes the in situ formation of the sialyl donor from NeuAc and CTP. The pyrophosphate byproduct is decomposed to inorganic phosphate by inorganic pyrophosphatase (PPase). Subsequently, the a-2,6-sialyltransferase accomplishes the sialyation of the lactosamine acceptor 120 and produces the ttansferase inhibitor CMP as a by-product. The CMP concentrations are kept low by conversion to CDP, and in so doing the problem of product inhibition is minimized. The cycle afforded 21% of the sialylated ttisac-charide 121, which is remarkable considering the complexity of the system and number of synthetic steps that can be avoided. 

Synthesis of iV-acetylneuraminic acid (Neu5Ac) in vivo is catalyzed by Neu5Ac synthetase (EC 4.1.3.19) through the irreversible condensation of phosphoenolpyruvate (PEP) and A/-acetylmannosamine (15) (Scheme 2) [30-32], This enzyme has not yet been isolated and its catalytic activity might be interesting field for exploration. 

The 3-deoxy-D-ara6mo-2-heptulosonic acid 7-phosphate (DAHP) synthetase (EC 4.1.2.15) is an enzyme involved in the shikimic pathway of aromatic amino acids biosynthesis in bacteria and plants, where catalyzes the construction of 3-deoxy-D-ara6/ o-2-heptulosonic acid 7-phosphate from phosphoenolpyruvate and D-erythrose 4-phosphate [6]. Although 3-deoxy-D-ara6/H0-2-heptulosonic acid 7-phosphate (DAHP) synthetase has not been widely investigated it has been employed for the DAHP synthesis on preparative scale from D-fructose in multienzyme system [68], This one-pot synthesis was subsequently even more simplified by the results of further studies which indicated that it was more efficient and economical to use the whole cells containing a DAHP synthetase plasmid [69]. 

Development of the chemical synthesis of ulosonic acids which mimics the enzymatic condensation is an interesting target [70]. As a model a reaction leading to KDO 8-phosphate (22) has been evaluated. In this reaction mediated by KDO 8-P synthetase D-arabinose 5-phosphate (A5P, 21) is reacted with phosphoenolpyruvate (PEP) (Scheme 7) [71]. 

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