Gluconeogenesis Pyruvic carboxylase

OVERSATURATION OXALOACETATE DECARBOXYLASE Oxaloacetate, synthesis in gluconeogenesis, PYRUVATE CARBOXYLASE PHOSPHOENOLPYRUVATE CARBOXYKI-NASE (PYROPHOSPHATE)... 

The main site of gluconeogenesis is the liver, although it also occurs to a far lesser extent in the kidneys. Very little gluconeogenesis occurs in brain or muscle. Within liver cells, the first enzyme of gluconeogenesis, pyruvate carboxylase, is located in the mitochondrial matrix. The last enzyme, glucose 6-phosphatase is bound to the smooth endoplasmic reticulum. The other enzymes of the pathway are located in the cytosol. 

The four key enzymes in gluconeogenesis (pyruvate carboxylase, PEP car-boxykinase, fructose-1,6-bisphosphatase, and glucose-6-phosphatase) are affected to varying degrees by allosteric modulators. For example, fructose-1,6-bisphos-phatase is activated by ATP and inhibited by AMP and fructose-2,6-bisphosphate. Acetyl-CoA activates pyruvate carboxylase. (The concentration of acetyl-CoA, a product of fatty acid degradation, is especially high during starvation.)... 

Several enzymes are common to both the glycolytic and gluconeogenic pathways, but four enzymes catalyse steps that only occur in gluconeogenesis pyruvate carboxylase (reaction 3.6), phosphoenol-... 

Figure 5.31 Reversal of the reaction of pyruvate kinase for gluconeogenesis — pyruvate carboxylase and phosphoenoipyruvate carboxykinase.

Pyruvate carboxylase is the most important of the anaplerotie reactions. It exists in the mitochondria of animal cells but not in plants, and it provides a direct link between glycolysis and the TCA cycle. The enzyme is tetrameric and contains covalently bound biotin and an Mg site on each subunit. (It is examined in greater detail in our discussion of gluconeogenesis in Chapter 23.) Pyruvate carboxylase has an absolute allosteric requirement for acetyl-CoA. Thus, when acetyl-CoA levels exceed the oxaloacetate supply, allosteric activation of pyruvate carboxylase by acetyl-CoA raises oxaloacetate levels, so that the excess acetyl-CoA can enter the TCA cycle. 

Acetyl-CoA is a potent allosteric effector of glycolysis and gluconeogenesis. It allosterically inhibits pyruvate kinase (as noted in Chapter 19) and activates pyruvate carboxylase. Because it also allosterically inhibits pyruvate dehydrogenase (the enzymatic link between glycolysis and the TCA cycle), the cellular fate of pyruvate is strongly dependent on acetyl-CoA levels. A rise in... 

Step 1 of Figure 29.13 Carboxylation Gluconeogenesis begins with the carboxyl-afion of pyruvate to yield oxaloacetate. The reaction is catalyzed by pyruvate carboxylase and requires ATP, bicarbonate ion, and the coenzyme biotin, which acts as a carrier to transport CO2 to the enzyme active site. The mechanism is analogous to that of step 3 in fatty-acid biosynthesis (Figure 29.6), in which acetyl CoA is carboxylated to yield malonyl CoA. 

The free glucose produced by this reaction is supplied to the blood from the tissues. As exemplified by gluconeogenesis, one may easily envision the economical organization of these metabolic routes, since, apart from four special gluconeogenesis enzymes-pyruvate carboxylase, phosphopyruvate carboxylase, fructose bisphosphatase, and glucose 6-phosphatase-individual glycolytic enzymes are also used in the gluconeogenesis. 

Most patients with pyruvate-carboxylase deficiency present with failure to thrive, developmental delay, recurrent seizures and metabolic acidosis. Lactate, pyruvate, alanine, [3-hydroxybutyrate and acetoacetate concentrations are elevated in blood and urine. Hypoglycemia is not a consistent finding despite the fact that pyruvate carboxylase is the first rate-limiting step in gluconeogenesis. 

Biotiii Pyruvate carboxylase Gluconeogenesis MCC (rare) excessive consumption... 

Pyruvate dehydrogenase is inhibited by its product acetyl CoA. This control is important in several contexts and shoidd be considered along with pyruvate carboxylase, the other mitochondrial enzyme that uses pyruvate (introduced in gluconeogenesis, Chapter 14, Figure 1-14-5). 

Between meals when fatty acids are oxidized in the liver for energy, accumulating acetyl CoA activates pyruvate carboxylase and gluconeogenesis and inhibits PDH, thus preventing conversion of lactate and alanine to acetyl CoA. 

The FADHj and NADH are oxidized in the electron transport chain, providing ATP. In musde and adipose tissue, the acetyl CoA enters the citric acid cyde. In liver, the ATP may be used for gluconeogenesis, and the acetyl CoA (which cannot be converted to glucose) stimulates gluco-neogenesis by activating pyruvate carboxylase. 

Answer B. Acetyl CoA activates pyruvate carboxylase and gluconeogenesis during fasting. 

A problem for gluconeogenesis is that pyruvate carboxylase, which produces oxaloacetate from pyravate, is present in the mitochondria but phosphoenolpyruvate carboxylase, at least in human liver, is present in the cytosol. For reasons given in Chapter 9, oxaloacetate cannot cross the mitochondrial membrane and so a transporter is not present in any cells. Hence, oxaloacetate is converted to phosphoenolpyruvate which is transported across the membrane (Figure 6.25). 

Figure 6.25 The intracellular location of the gluconeogenic enzymes. The gluconeogenic enzymes are located in the cytosol, except for pyruvate carboxylase which is always present within the mitochondria phosphoenolpyruvate carboxykinase is cytoplasmic in some species including humans. Consequently phosphoenolpyruvate must be transported across the inner mitochondrial enzyme by a transporter molecule in order for gluconeogenesis to take place.

The answer is B. While all of the listed conditions are consistent with lethargy and developmental defects, the lactic acidosis rules out pyruvate kinase deficiency. Thiamine and niacin deficiencies are unlikely due to the lack of effect of vitamin supplementation. Excess pyruvate is the source of the elevated alanine in the serum. The clinical findings are thus consistent with pyruvate carboxylase deficiency, which is associated with severe hypoglycemia due to fasting due to impaired gluconeogenesis. 

Diabetes - insulin dependent Methyl malonic, propionic or isovaleric acidaemias Pyruvate carboxylase and multiple carboxylase deficiency Gluconeogenesis enzyme deficiency glucose-6-phosphatase, fructose-1,6-diphosphatase or abnormality of glycogen synthesis (glycogen synthase) Ketolysis defects Succinyl coenzyme A 3-keto acid transferase ACAC coenzyme A thiolase... 

Pyruvate carboxylase is stimulated by acetyl-CoA, increasing the rate of gluconeogenesis when the cell already has adequate supplies of other substrates (fatty acids) for energy production. 

Gluconeogenesis is regulated at the level of pyruvate carboxylase (which is activated by acetyl-CoA) and FBPase-1 (which is inhibited by fructose 2,6-bisphosphate and AMP). 

Carboxylation of pyruvate to oxaloacetate (OAA) by pyruvate carboxylase is a biotin-dependent reaction (see Figure 8.24). This reaction is important because it replenishes the citric acid cycle intermediates, and provides substrate for gluconeogenesis (see p. 116). 

The first "roadblock" to overcome in the synthesis of glucose from pyruvate is the irreversible conversion in glycolysis of pyruvate to phosphoenolpyruvate (PEP) by pyruvate kinase. In gluconeogenesis, pyruvate is first carboxylated by pyruvate carboxylase to oxaloacetate (OAA), which is then converted to PEP by the action of PEP-carboxykinase (Figure 10.3). 

Allosteric activation of hepatic pyruvate carboxylase by acetyl CoA occurs during fasting. As a result of excessive lipolysis in adipose tis sue, the liver is flooded with fatty acids (see p. 328). The rate of for mation of acetyl CoA by p-oxidation of these fatty acids exceeds the capacity of the liver to oxidize it to C02 and H20. As a result, acetyl CoA accumulates and leads to activation of pyruvate carboxylase. [Note Acetyl CoA inhibits pyruvate dehydrogenase (see p. 108). Thus, this single compound can divert pyruvate toward gluconeogenesis and away from the TCA cycle.]... 

During a fast, the liver is flooded with fatty acids mobilized from adipose tissue. The resulting elevated hepatic acetyl CoA produced primarily by fatty acid degradation inhibits pyruvate dehydrogenase (see p. 108), and activates pyruvate carboxylase (see p. 117). The oxaloacetate thus produced is used by the liver for gluconeogenesis rather than for the TCA cycle. Therefore, acetyl Co A is channeled into ketone body synthesis. 

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