Weill Cornell Medicine-Qatar Principles of Biochemistry Lecture 16b PDF

Principles of Biochemistry SPRING 2024 Professor: Moncef LADJIMI [email protected] Office: C-169 As faculty of Weill Cornell Medical College in Qatar we are committed to providing transparency for any and all external relationships prior to giving an academic presentation. I, Moncef LADJ...

Principles of Biochemistry SPRING 2024 Professor: Moncef LADJIMI [email protected] Office: C-169 As faculty of Weill Cornell Medical College in Qatar we are committed to providing transparency for any and all external relationships prior to giving an academic presentation. I, Moncef LADJIMI DO NOT have a financial interest in commercial products or services. Lecture 16a Carbohydrates oxidation: Glycolysis Lecture 16b Glucose Synthesis: Gluconeogenesis Glycolysis and Gluconeogenesis: A Balancing Act Additional material for this lecture may be found in: § Lehninger’s Biochemistry (8th ed), chapter 14: p. 510-539 Lecture 16b GLUCOSE SYNTHESIS: GLUCONEOGENESIS Key topics: – Synthesis of glucose from simpler compounds: gluconeogenesis – Glycolysis vs gluconeogenesis – Glucogenic amino acids CARBOHYDRATE SYNTHESIS FROM SIMPLE PRECURSORS The pathway from phosphoenolpyruvate to glucose 6phosphate 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. § Any compound that can be converted to either pyruvate or oxaloacetate can therefore serve as starting material for gluconeogenesis. This includes aminoacids 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. oxaloacetate § Glycerol (a three carbon molecule from triacylglycerols) enters gluconeogenesis at the level of glyceraldehyde-3-phosphate Notice that mammals cannot convert fatty acids to sugars. § Plants and photosynthetic bacteria are uniquely able to convert CO2 to carbohydrates, using the Calvin cycle. PRECURSORS FOR GLUCONEOGENESIS Humans can produce glucose mainly from sugars or proteins – Sugars: pyruvate, lactate, or oxaloacetate – Protein: from amino acids that can be converted to citric acid cycle intermediates (or glucogenic amino acids) – Glycerol: from breakdown of fat (used to a lesser extent) Humans cannot produce glucose from fatty acids – Product of fatty acid degradation is acetyl-CoA – Cannot have a net conversion of acetyl-CoA to oxaloacetate Plants, yeast, and many bacteria can do this, thus producing glucose from fatty acids GLYCOLYSIS VS. GLUCONEOGENESIS Opposing pathways that are both thermodynamically favorable – Operate in opposite directions end product of one is the starting compound of the other Reversible reactions are used by both pathways Irreversible reaction of glycolysis must be bypassed in gluconeogenesis – Highly thermodynamically favorable, and regulated – Different enzymes in the different pathways – Differentially regulated to prevent a futile cycle GLYCOLYSIS VS. GLUCONEOGENESIS Reactions of glycolysis: 10 reactions with 3 irreversible (in red) The opposing pathway of gluconeogenesis: - Reversible reactions are used by both pathways - 3 bypass reactions (in blue) of gluconeogenesis (bypass of irreversible glycolytic reactions:use of different enzymes in the different pathways). Glycolysis occurs mainly in the muscle and brain (cytosol). Glycolysis and gluconeogenesis are reciprocally regulated Gluconeogenesis occurs mainly in the liver (mitochondria and cytosol). FIRST BYPASS REACTION: FROM PYRUVATE TO PHOSPHOENOLPYRUVATE Requires two energy-consuming steps First step, pyruvate carboxylase (PC) converts pyruvate (using bicarbonate and ATP) to oxaloacetate – Carboxylation using a biotin cofactor – Occurs in mitochondria – Requires transport of oxaloacetate outside mitochondria (via malate because there is no oxaloacetate transporters) Second step, phosphoenolpyruvate carboxykinase (PEPCK or PCK) converts oxaloacetate to PEP – Phosphorylation from GTP and decarboxylation – Occurs in mitochondria or cytosol depending on the conditions ALTERNATIVE PATH FROM PYRUVATE TO PHOSPHOENOLPYRUVATE (STARTS WITH LACTATE) Glucose GTP Cytosolic Gluconeogenic enzymes The relative importance of the two pathways depends on the availability of lactate or pyruvate and the cytosolic requirements for NADH for gluconeogenesis. Lactate is the precursor, when [lactate] is high and NADH is needed in the cytosol. NADH is thus generated in the lactate dehydrogenase reaction and does not have to be shuttled out of the mitochondrion. Pyruvate is the precursor, when [pyruvate] or [alanine] are high. In this case NADH is shuttled out of mitochondria. [pyruvate] or [alanine] are high [lactate] is high and NADH is needed in the cytosol This also illustrates an alternative route for oxaloacetate produced in mitochondria. SYNTHESIS OF OXALOACETATE FROM PURUVATE In mitochondria, pyruvate is converted to oxaloacetate in a biotin-requiring reaction catalyzed by pyruvate carboxylase. BIOTIN IS A CO2 CARRIER Role of biotin (Vitamin B7) 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 biotinylenzyme. 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 carboxybiotinylenzyme to catalytic site 2 on the enzyme surface, where CO2 is released and reacts with the pyruvate, forming oxaloacetate and regenerating the biotinyl-enzyme. Similar mechanisms occur in other biotin-dependent carboxylation reactions, such as those catalyzed by propionyl-CoA carboxylase and acetyl-CoA carboxylase. FROM OXALOACETATE TO PHOSPHOENOLPYRUVATE - In the cytosol, oxaloacetate is converted to phosphoenolpyruvate by cytosolic PEP carboxykinase. - In mitochondria, oxaloacetate is converted to phosphoenolpyruvate by mitochondrial 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 g phosphate of GTP. SECOND AND THIRD BYPASSES Catalyze reverse reaction of opposing step in glycolysis Are irreversible themselves Second bypass: – Fructose 1,6-bisphosphate à Fructose 6-Phosphate By fructose bisphosphatase-1 Coordinately/oppositely regulated with PFK Third bypass: – Glucose 6-phosphate à Glucose By glucose 6-phosphatase SEQUENTIAL REACTIONS IN GLUCONEOGENESIS FROM PYRUVATE Pyruvate + 𝐇𝐂𝐎" 𝟑 + ATP → oxaloacetate + ADP + Pi ×2 Oxaloacetate + GTP ⇌ phosphoenolpyruvate + CO2 + GDP ×2 Phosphoenolpyruvate + H2O ⇌ 2-phosphoglycerate ×2 2-Phosphoglycerate ⇌ 3-phosphoglycerate ×2 3-Phosphoglycerate + ATP ⇌ 1,3-bisphosphoglycerate + ADP ×2 1,3-Bisphosphoglycerate + NADH + H+ ⇌ glyceraldehyde 3-phosphate + NAD+ + Pi ×2 Glyceraldehyde 3-phosphate ⇌ dihydroxyacetone phosphate Glyceraldehyde 3-phosphate + dihydroxyacetone phosphate ⇌ fructose 1,6bisphosphate Fructose 1,6-bisphosphate → fructose 6-phosphate + Pi Fructose 6-phosphate ⇌ glucose 6-phosphate Glucose 6-phosphate + H2O → glucose + Pi Sum: 2 Pyruvate + 4ATP + 2GTP + 2NADH + 2H+ + 4H2O → glucose + 4ADP + 2GDP + 6Pi + 2NAD+ Note: The bypass reactions are in red. All other reactions are the reversible steps in glycolysis GLUCONEOGENESIS IS EXPENSIVE BUT ESSENTIAL 2 Pyruvate + 4 ATP + 2 GTP + 2 NADH + 2 H+ + 4 H2O à Glucose + 4 ADP + 2 GDP + 6 Pi + 2 NAD+ Gluconeogenesis is expensive and costs: – 4 ATP, 2 GTP, and 2 NADH But it is physiologically necessary: – in the brain, nervous system, and red blood cells which generate ATP ONLY from glucose – when glycogen stores (mostly in liver and muscles) are depleted, as during starvation or vigorous exercise. CITRIC ACID INTERMEDIATES AND ALL BUT TWO AMINO ACIDS ARE GLUCOGENIC q All aminoacids derived from proteins (except Lys and Leu) are ultimately catabolized to pyruvate or to intermediates in the Citric Acid Cycle (CAC). Such amino acids can undergo net conversion to glucose and are said to be glucogenic. q Thus, gluconeogenesis allows the net synthesis of glucose from pyruvate but also from 4 and 5carbon intermediates of the CAC (a-ketoglutarate, succinyl-CoA, fumarate and oxaloacetate), to which amino acids are converted. Lys and Leu are not glucogenic Remember to prepare for next lecture: Lehninger’s Biochemistry (8th ed), §chapter 14: p. 546-552