Pyruvate kinase, reaction catalyzed

The enzyme that catalyzes the conversion of PEP to pyruvate is pyruvate kinase. Liver pyruvate kinase is stimulated allosterically by fructose-1,6-diphosphate, AMP, ADP, and glyceraldehyde-3-phosphate. It is inhibited by alanine, ATP, NADH, and, more importantly, by cAMP- and Ca2 calmodulin-controlled phosphorylation. High blood glucagon levels thus inhibit the activities of both PFK II and pyruvate kinase in the liver through phosphorylation. Transcription of pyruvate kinase is also decreased by glucagon and increased by insulin. Muscle pyruvate kinase is not subject to cAMP or Ca2+ regulation. The pyruvate kinase reaction is practically irreversible. 

Pyruvate kinase irreversibly catalyzes the formation of ATP and pyruvate from phosphoenolpyruvate (PEP) and ADP. This reaction is named for the direction that is opposite to the flow of the process. Since the high-energy... 

The two reactions are coupled by the enzyme pyruvate kinase, which catalyzes reaction... 

Despite these reservations, use of either the relative maximal capacity or the equilibrium/nonequilibrium reaction approach identifies the same enzymes as catalyzing potentially rate-controlling reactions. For example, in most mammalian tissues both methods suggest that the potential exists for control of glycolytic flux at the hexokinase, phosphofructokinase, and pyruvate kinase reactions (cf. Newsholme and Start, 1973). 

The second ATP-synthesizing reaction of glycolysis is catalyzed by pyruvate kinase, which brings the pathway at last to its pyruvate branch point. Pyruvate kinase mediates the transfer of a phosphoryl group from phosphoenolpyru-vate to ADP to make ATP and pyruvate (Figure 19.27). The reaction requires... 

Transfer of the phosphoryl group to ADP in step 10 then generates ATP and gives enolpyruvate, which undergoes tautomerization to pyruvate. The reaction is catalyzed by pyruvate kinase and requires that a molecule of fructose 1,6-bis-phosphate also be present, as well as 2 equivalents of Mg2+. One Mg2+ ion coordinates to ADP, and the other increases the acidity of a water molecule necessary for protonation of the enolate ion. 

Glycolysis A net formation of two results from the formation of lactate from one molecule of glucose, generated in two reactions catalyzed by phospho-glycerate kinase and pyruvate kinase, respectively (Figure 17-2). 

Glycolysis is regulated by three enzymes catalyzing nonequilibrium reactions hexokinase, phosphoffuc-tokinase, and pyruvate kinase. 

Three nonequilibrium reactions catalyzed by hexoki-nase, phosphofructokinase, and pyruvate kinase prevent simple reversal of glycolysis for glucose synthesis (Chapter 17). They are circumvented as follows ... 

CK catalyzes the reversible phosphorylation of creatine in the presence of ATP and magnesium. When creatine phosphate is the substrate, the resulting creatine can be measured as the ninhydrin fluorescent compound, as in the continuous flow Auto Analyzer method. Kinetic methods based on coupled enzymatic reactions are also popular. Tanzer and Gilvarg (40) developed a kinetic method using the two exogenous enzymes pyruvate kinase and lactate dehydrogenase to measure the CK rate by following the oxidation of NADH. In this procedure the main reaction is run in a less favorable direction. 

The Jirst indirect route in glucose synthesis involves the formation of phosphoenolpyruvate from pyruvate without the intervention of pyruvate kinase. This route is catalyzed by two enzymes. At first, pyruvate is converted into oxaloacetate. This reaction occurs in the mitochondria as the pyruvate molecules enter them, and is catalyzed by pyruvate carboxylase according to the scheme... 

Pyruvate kinase the last enzyme in aerobic glycolysis, it catalyzes a substrate-level phosphorylation of ADP using the high-energy substrate phosphoenolpyruvate (PEP). Pyruvate kinase is activated by fructose 1,6-bisphosphate from the PFK-1 reaction (feedforward activation). 

The process of converting an enol to a ketone. Pyruvate kinase catalyzes a ketonization reaction in the conversion of the enolpyruvate intermediate to pyruvate. See... 

Figure 6-1. The steps of glycolysis. Feedback inhibition of glucose phosphorylation by hexokinase, inhibition of pyruvate kinase, and the main regulatory, rate-limiting step catalyzed by phosphofructoki-nase (PFK-I) are indicated, pyruvate formation and substrate-level phosphorylation are the main outcomes of these reactions. Regeneration of NAD occurs by reduction of pyruvate to lactate during anaerobic glycolysis.

FIGURE 13-3 Hydrolysis of phosphoenolpyruvate (PB3). Catalyzed by pyruvate kinase, this reaction is followed by spontaneous tautomerization of the product, pyruvate. Tautomerization is not possible in PER and thus the products of hydrolysis are stabilized relative to the reactants Fiesonance stabilization of P, also occurs, as shown in Figure 13-1. 

The formation of ATP (or GTP) at the expense of the energy released by the oxidative decarboxylation of a-ketoglutarate is a substrate-level phosphorylation, like the synthesis of ATP in the glycolytic reactions catalyzed by glyceraldehyde 3-phosphate dehydrogenase and pyruvate kinase (see Fig. 14-2). The GTP formed by succinyl-CoA synthetase can donate its terminal phosphoryl group to ADP to form ATP, in a reversible reaction catalyzed by nucleoside diphosphate kinase (p. 505) ... 

Seven of the reactions of glycolysis are reversible and are used for gluconeogenesis in the liver and kidneys. Three reactions are physiologically irreversible and must be circum vented. These reactions are catalyzed by the glycolytic enzymes pyruvate kinase, phos phofructokinase, and hexokinase. 

The 500-residue subunits of pyruvate kinase consist of four domains,891 the largest of which contains an 8-stranded barrel similar to that present in triose phosphate isomerase (Fig. 2-28). Although these two enzymes catalyze different types of reactions, a common feature is an enolic intermediate. One could imagine that pyruvate kinase protonates its substrate phosphoenolpyruvate (PEP) synchronously with the phospho group transfer (Eq. 12-42). However, the enzyme catalyzes the rapid conversion of the enolic form of pyruvate to the oxo form (Eq. 12-43) adding the proton sterospecifically to the si face. This and other evidence favors the enol as a true intermediate... 

If we add the equations for the reactions catalyzed by pyruvate carboxylase, phosphoenolpyruvate carboxykinase, and nucleoside diphosphate kinase, we obtain the overall reaction for conversion of pyruvate to phosphoenolpyruvate. 

Some of the reactions of PO3- parallel enzymatic reactions promoted by adenosine triphosphate (ATP). Pyruvate kinase catalyzes the equilibration of ATP and pyruvate with adenosine diphosphate (ADP) and phosphoenol pyruvate (11,12). In a formal sense, this reaction resembles the preparations of enol phosphate (eqs. 6 and 7). Cytidine triphosphate synthetase catalyzes the reaction of uridine triphosphate with ammonia to yield cytidine triphosphate (13). In a formal sense, this reaction resembles the replacement of the ester carbonyl group of ethyl acetate by the nitrogen of aniline (eq. 8). 

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. 

In the last reaction, pyruvate kinase catalyzes the physiologically irreversible transfer of the phosphoryl group from PEP to ADP to form ATP and pyruvate. 

Fig. 1. Comparison of gluconeogenesis and glycolysis. The three steps of glycolysis that are irreversible are numbered. (1) Flexokinase in glycolysis is reversed by glucose 6-phosphatase in gluconeogenesis (2) PFK in glycolysis is reversed by fructose 1,6-bisphosphatase in gluconeogenesis (3) pyruvate kinase in glycolysis is reversed by two sequential reactions in gluconeogenesis catalyzed by pyruvate carboxylase and PEP carboxykinase.

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