BCH 5045 Graduate Survey of Biochemistryhort.ifas.ufl.edu/faculty/guy/bch5045/Lecture Files/Lecture 45.pdf · Then CO2 reacts with biotin, forming carboxybiotinyl-enzyme. Th\ long

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BCH 5045

Graduate Survey of Biochemistry

Instructor: Charles Guy Producer: Ron Thomas Director: Glen Graham

Lecture 45

Slide sets available at: http://hort.ifas.ufl.edu/teach/guyweb/bch5045/index.html

http://hort.ifas.ufl.edu/teach/guyweb/bch5045/index.html�



• LEHNINGER • PRINCIPLES OF BIOCHEMISTRY

• Fifth Edition

David L. Nelson and Michael M. Cox

© 2008 W. H. Freeman and Company

CHAPTER 14 Glycolysis, Gluconeogenesis, and the

Pentose Phosphate Pathway



Seven of the ten reactions of glycolysis are physiologically reversible and three are not under physiological conditions. What controls which direction a reaction will go? What about reactions 1, 3 and 10, can they be reversed?



Glycolytic Intermediate Rat Liver nmol/mg DNA

Mouse Muscle mmol/kg d. m.

Human Blood nmol/ml

ATP 1423 24 1128

ADP 299 3.2 126

AMP 8 0.1 50

UDP-Glucose 195 - -

Glucose-1-P 4 0.1 -

Glucose-6-P 81 1.7 27

Fructose-6-P 22 0.3 11

Fructose-1,6-BP 9 0.3 5

Dihydroxyacetone Phosphate 26 - -

Glyceraldehyde-3-P - 0.1 4

Glycerol-1-P 55 0.8

3-Phosphoglycerate 114 - 48

2-Phosphosglycerate - - 7

Phosphoenolpyruvate 64 - 12

Pyruvate 62 0.4 71

Lactate 351 5.1 1190 1From Faupel, Seitz and Tarnowski, (1972) Arch. Biochem. Biosphys. 148, 509-522; 2Harris, Hultman and Nordesio (1974) Scand. J. clin. Lab. Invest. 33, 109-120; 3Minakami, Suzuki, Saito and Yoshikawa (1965) J. Biochem. 58, 543-550.



Glucose in the brain is used to produce energy, perhaps its most critical function, but is also needed for regulatory, protective (against ROS) and anabolic (protein and lipid synthesis) processes. In the brain, surprisingly glucose metabolism can deliver energy quickly and efficiently for cell functions independent or without the operation of oxidative phosphorylation. Energy production from glucose in an aerobic environment is termed aerobic glycolysis. Recent studies suggest there is regional variation in the rate of aerobic glycolysis within a resting brain, and possibly high rates of aerobic glycolysis may be linked to the formation of amyloid-β plaques. High rates of aerobic glycolysis have been observed in the prefrontal and lateral parietal cortex, and posterior cingulate cortex just to list a few regions that make up the default mode network that is active when one is awake but not performing a task.

This slide is not in the lecture video



In contrast, the cerebellum and inferior temporal gyrus, including the hippocampus, have low aerobic glycolysis rates. Amyloid-β plaques are often found in the default mode network regions in the early stages of Alzheimer's disease. In a study of patients with Alzheimer's disease and people with elevated levels of the amyloid-β protein that were cognitively normal, PET was used to assess amyloid-β deposition and mapped against the spatial distribution aerobic glycolysis levels. There was a high spatial correlation between levels of amyloid-β deposition and aerobic glycolysis, and the correlation was higher in people with Alzheimer's than in cognitive normal people with elevated amyloid-β. Please keep in mind that correlation does not prove cause and effect.

Vaishnavi, S. N. et al. (2010) Regional aerobic glycolysis in the human brain. Proc. Natl Acad. Sci. Vlassenko, A. G. et al. Spatial correlation between brain aerobic glycolysis and amyloid-β deposition. Proc. Natl Acad. Sci.



Presenter

Presentation Notes

FIGURE 14-15 Carbohydrate synthesis from simple precursors. The pathway from phosphoenolpyruvate to glucose 6-phosphate is common to the biosynthetic conversion of many different precursors of carbohydrates in animals and plants. The path from pyruvate to phosphoenolpyruvate leads through oxaloacetate, an intermediate of the citric acid cycle, which we discuss in Chapter 16. Any compound that can be converted to either pyruvate or oxaloacetate can therefore serve as starting material for gluconeogenesis. This includes alanine and aspartate, which are convertible to pyruvate and oxaloacetate, respectively, and other amino acids that can also yield three- or four-carbon fragments, the so-called glucogenic amino acids (Table 14-4; see also Figure 18-15). Plants and photosynthetic bacteria are uniquely able to convert CO2 to carbohydrates, using the glyoxylate cycle (p. 639). FIGURE 14-16 Opposing pathways of glycolysis and gluconeogenesis in rat liver. The reactions of glycolysis are on the left side, in red; the opposing pathway of gluconeogenesis is on the right, in blue. The major sites of regulation of gluconeogenesis shown here are discussed later in this chapter, and in detail in Chapter 15. Figure 14-19 illustrates an alternative route for oxaloacetate produced in mitochondria.



Presenter

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FIGURE 14-17a Synthesis of phosphoenolpyruvate from pyruvate. (a) In mitochondria, pyruvate is converted to oxaloacetate in a biotin-requiring reaction catalyzed by pyruvate carboxylase. FIGURE 14-17b Synthesis of phosphoenolpyruvate from pyruvate. (b) In the cytosol, oxaloacetate is converted to phosphoenolpyruvate by PEP carboxykinase. The CO2 incorporated in the pyruvate carboxylase reaction is lost here as CO2. The decarboxylation leads to a rearrangement of electrons that facilitates attack of the carbonyl oxygen of the pyruvate moiety on the γ phosphate of GTP. FIGURE 14-18 Role of biotin in the pyruvate carboxylase reaction. The cofactor biotin is covalently attached to the enzyme through an amide linkage to the ε-amino group of a Lys residue, forming a biotinyl-enzyme. The reaction occurs in two phases, which occur at two different sites in the enzyme. At catalytic site 1, bicarbonate ion is converted to CO2 at the expense of ATP. Then CO2 reacts with biotin, forming carboxybiotinyl-enzyme. The long arm composed of biotin and the Lys side chain to which it is attached then carry the CO2 of carboxybiotinyl-enzyme to catalytic site 2 on the enzyme surface, where CO2 is released and reacts with the pyruvate, forming oxaloacetate and regenerating the biotinyl-enzyme. The general role of flexible arms in carrying reaction intermediates between enzyme active sites is described in Figure 16-17, and the mechanistic details of the pyruvate carboxylase reaction are shown in Figure 16-16. Similar mechanisms occur in other biotin-dependent carboxylation reactions, such as those catalyzed by propionyl-CoA carboxylase (see Figure 17-11) and acetyl-CoA carboxylase (see Figure 21-1).



Presenter

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FIGURE 14-19 Alternative paths from pyruvate to phosphoenolpyruvate. The relative importance of the two pathways depends on the availability of lactate or pyruvate and the cytosolic requirements for NADH for gluconeogenesis. The path on the right predominates when lactate is the precursor, because cytosolic NADH is generated in the lactate dehydrogenase reaction and does not have to be shuttled out of the mitochondrion (see text).





The Pentose Phosphate Pathway or the Reductive Pentose Pathway has two major functions: the production of reducing potential in the form of NADPH, and the synthesis of five carbon sugars, notably D-ribose. Thus there are two ways of viewing this metabolic pathway, that part which generates NADPH and the portion that leads to the production of 4, 5 and 7 carbon sugars. The first step of the pathway begins when G-6-P is oxidized to 6-phospho-gluconolactone producing NADPH. The lactone is hydrated to form 6-phospho-gluconate which is then oxidized and decarboxylated to form a second molecule of NADPH and D-ribulose-5-phosphate. The Ru-5-P is isomerized to D-ribose-5-P. If five carbon sugars are not needed, then they are recycled back into hexose phosphates. This part of the pathway is a recapitulation of part of the Calvin Cycle, the cycle important in photosynthesis. I will have more to say about that when we discuss photosynthesis.



The second part of the pathway produces the 4, 5, and 7 carbon sugars by combining the activities of four enzymes, isomerase, epimerase, transketolase and a transaldolase. Like glycolysis all of the sugars of the PPP are sugar phosphates.

Presenter

Presentation Notes

FIGURE 14-20 General scheme of the pentose phosphate pathway. NADPH formed in the oxidative phase is used to reduce glutathione, GSSG (see Box 14-4) and to support reductive biosynthesis. The other product of the oxidative phase is ribose 5-phosphate, which serves as a precursor for nucleotides, coenzymes, and nucleic acids. In cells that are not using ribose 5-phosphate for biosynthesis, the nonoxidative phase recycles six molecules of the pentose into five molecules of the hexose glucose 6-phosphate, allowing continued production of NADPH and converting glucose 6-phosphate (in six cycles) to CO2.



Presenter

Presentation Notes

FIGURE 14-21 Oxidative reactions of the pentose phosphate pathway. The end products are ribose 5-phosphate, CO2, and NADPH.



Presenter

Presentation Notes

FIGURE 14-22 Nonoxidative reactions of the pentose phosphate pathway. (a) These reactions convert pentose phosphates to hexose phosphates, allowing the oxidative reactions (see Figure 14-21) to continue. Transketolase and transaldolase are specific to this pathway; the other enzymes also serve in the glycolytic or gluconeogenic pathways. (b) A schematic diagram showing the pathway from six pentoses (5C) to five hexoses (6C). Note that this involves two sets of the interconversions shown in (a). Every reaction shown here is reversible; unidirectional arrows are used only to make clear the direction of the reactions during continuous oxidation of glucose 6-phosphate. In the light-independent reactions of photosynthesis, the direction of these reactions is reversed (see Figure 20-10).



Presenter

Presentation Notes

FIGURE 14-23a The first reaction catalyzed by transketolase. (a) The general reaction catalyzed by transketolase is the transfer of a two-carbon group, carried temporarily on enzyme-bound TPP, from a ketose donor to an aldose acceptor. FIGURE 14-23b The first reaction catalyzed by transketolase. (b) Conversion of two pentose phosphates to a triose phosphate and a seven-carbon sugar phosphate, sedoheptulose 7-phosphate.



Presenter

Presentation Notes

FIGURE 14-24 The reaction catalyzed by transaldolase. FIGURE 14-25 The second reaction catalyzed by transketolase.



Presenter

Presentation Notes

FIGURE 14-26 Carbanion intermediates stabilized by covalent interactions with transketolase and transaldolase. (a) The ring of TPP stabilizes the carbanion in the dihydroxyethyl group carried by transketolase; see Figure 14-14 for the chemistry of TPP action. (b) In the transaldolase reaction, the protonated Schiff base formed between the ε-amino group of a Lys side chain and the substrate stabilizes the C-3 carbanion formed after aldol cleavage.



Presenter

Presentation Notes

FIGURE 14-27 Role of NADPH in regulating the partitioning of glucose 6-phosphate between glycolysis and the pentose phosphate pathway. When NADPH is forming faster than it is being used for biosynthesis and glutathione reduction (see Figure 14-20), [NADPH] rises and inhibits the first enzyme in the pentose phosphate pathway. As a result, more glucose 6-phosphate is available for glycolysis.



It can be said that for many organisms, there are two primary routes for glucose metabolism, glycolysis and the Pentose Phosphate Pathway (PPP). While glucose-6-phosphate can be the initial substrate for the the direct entry into the PPP, glycolytic metabolites fructose-6-phosphate (F6P) and glyceraldehyde-3-phosphate (G3P) derived from glucose can also serve as the building blocks for the sugar phosphate intermediates of the second phase of the PPP. Based on thermodynamic considerations the non-oxidative second phase of the PPP would be presumably fully reversible meaning inputs of F6P and G3P may just as easily enter as exit the PPP. However, sedoheptulose-1,7-bisphosphatase (SBPase) appears to catalyze the committed reaction that allows metabolite flow from glycolysis into the PPP which is often directed into ribose-5-phosphate production, a process known as ribogenesis.




Clasquin et al. has concluded that the riboneogenic pathway in yeast based on their findings has striking similarity to the Calvin Cycle of photosynthesis. A sedoheptulose-1,7-bisphosphatase (SBPase) catalyzes the dephosphorylation sedoheptulose-1,7-bisphosphate to form sedoheptulose-7-phosphate, that is then converted to ribulose-1,5-bisphosphate, which is the primary the substrate for fixation of carbon dioxide in photosynthesis.

Clasquin et al (2011) Ribogenesis in yeast. Cell 145, 969-980


This and the previous slide is on exam III.

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BCH 5045 Graduate Survey of Biochemistryhort.ifas.ufl.edu/faculty/guy/bch5045/Lecture Files/Lecture 45.pdf · Then CO2 reacts with biotin, forming carboxybiotinyl-enzyme. Th\ long