glycolysis and TCA

Question 1

is glucose reduced or oxidized

Answer 1

highly reduced- oxidation is employed in glycolysis to gain energy

Question 2

Why is glycolysis important

Answer 2

glycolytic breakdown of glucose is the major source of energy of RBCs and sperm, and provides energy to tissues when O2 delivery is limited, such as during exercise

Question 3

phases of glycolysis

Answer 3

1. energy investment phase/ preparatory phase- 2 ATP are used to activate glucose leading to generation of glyceraldehyde-3-P. 2. Payoff phase- energy is extracted for use

Question 4

How does glucose enter cells

Answer 4

glucose transporters. Insulin sensitive tissues (skeletal muscle and adipose tissue): Glut4 increases glucose transport into the cell following exposure to insulin. Insulin dependent tissues (liver): Glut 2 does not change in response to insulin

Question 5

List the three types of rxns that occur in glycolysis

Answer 5

1. Degradation of the carbon skeleton of glucose to yield pyruvate. 2. substrate level Phosphorylation of ADP to ATP. 3. Generation of NADH

Question 6

describe reaction 1 of glycolysis

Answer 6

Hexokinase or glucokinase catalyzes activation of D-glucose into glucose-6-phosphate, using first ATP. Ireversible rxn which traps glucose in cell (negative charge), conserves metabolic energy and lowers activation energy of next rxn

Question 7

hexokinase vs glucokinase

Answer 7

hexokinase 1: not very selective, present in all cells, low Km for sugars and inhibited by glucose-6-P. Glucokinase: selective for glucose, located in liver and pancreatic beta cells, high Km for glucose, inhibited by fructose-6-P

Question 8

compare carb metabolism in muscle vs liver

Answer 8

liver has Glut2 which maintains glucose conc close to that in the blood. Since glucokinase converts glucose to glu-6-P, this favors net flux of glucose into liver during fed state and net efflux during fasting. When glu is low, glucokinase activity is lower which favors delivery of glu to peripheral tissues containing hexokinase. muscle consumes glu and uses it for energy.

Question 9

rate limiting step of glycolysis

Answer 9

Reaction 3: fructose-6-phosphate converted to fructose 1,6-bisphosphate by Phosphofructokinase 1 using second ATP. This is the committed step of glycolysis and is irreversible

Question 10

properties of phosphofructokinase 1

Answer 10

allosteric enzyme- ATP or citrate inhibits the enzyme while AMP stimulates it. The most potent activator of PFK1 is fructose 2,6-bisphosphate which is produced by phosphofructokinase 2 from fructose-6-phosphate

Question 11

properties of phosphofructokinase 2

Answer 11

Converts fructose-6-phosphate to fructose 2,6 bisphosphate which activates PFK1 (glycolysis) and inhibits fructose 1,6 bisphosphatase (gluconeogenesis). This enzyme is bifunctional- can be a kinase or phosphatase

Question 12

compare actions of phosphofructokinase during fed and fasting states

Answer 12

Fed: High insulin and low glucagon increase F2,6BP and incrase glycolysis. Fasting: low insulin and high glucagon activates protein Kinase A > phosphorylates PFK-2 which turns into fructose 2,6 bisphosphatase > transforms F 2,6 BP into F6P > inhibits glycoslysis and promotes gluconeogenesis.

Question 13

describe the first step in energy generating phase of glycolysis

Answer 13

Glyceraldehyde-3-phosphate + 2 NAD+ + Pi (2) 1,3-bisphosphoglycerate + 2NADH + 2H+. Catalyzed by glyceraldehyde-3-phosphate dehydrogenase this is the first oxidation rxn and the first NADH generated

Question 14

describe the first substrate level phosphorylation of glycolysis

Answer 14

rxn 5: 1,3-Bisphosphoglycerate + 2ADP –> 2 3-phosphoglycerate + 2 ATP. Catalyzed by phosphoglycerate kinase. At this step, net ATP yield is 0 b/c 2 have been made and 2 have been used.

Question 15

describe the second substrate level phosphorylation of glycolysis

Answer 15

2 phosphoenol pyruvate + 2ADP –> 2 pyruvate + 2ATP. Catalyzed by pyruvate kinase. Now there is net yield of 2 ATP for each molecule of glucose

Question 16

What inhibits pyruvate kinase

Answer 16

ATP, alanine, and protein kinase A. This promotes gluconeogenesis and inhibits glycolysis when there is sufficient cellular energy

Question 17

pyruvate kinase deficiency

Answer 17

Second most common cause (after Glucose-6-Phosphate Dehydrogenase deficiency) of enzyme deficiency-linked hemolytic anemia. Normally, Glucagon results in activated protein kinase A which phosphorylates pyruvate kinase and turns off glycolysis.

Question 18

fates of pyruvate

Answer 18

In presence of O2 and mitochondria: oxidation to Acetyl CoA. Hypoxic/ no mitochondria conditions: pyruvate is converted to lactic acid then heart muscle converts lactate to pyruvate which is then oxidized. Lactate dehydrogenase catalyzes pyruvate/lactate interconversion

Question 19

Alternative fates of pyruvate in fed vs fasting state (aerobic environment)

Answer 19

fed: converted to alanine, or in cases of excess carb intake can enter mitochondria and increase amount of Acety CoA available for fatty acid synthesis. Fasting: in liver, pyruvate (from lactate) is converted to oxaloacetate and used for gluconeogenesis.

Question 20

How does pyruvate enter TCA cycle

Answer 20

Pyruvate is transported from cytoplasm into mitochondria where Pyruvate dehydrogenase catalyzes pyruvate transformation into acetyl CoA. (aerobic conditions)

Question 21

pyruvate dehydrogenase components

Answer 21

Made of 3 enzymes and five coenzymes (coenzyme A, thiamine pyrophosphate (TPP), prosthetic groups, flavin adenine dinucleotide (FAD), nicotinamide adenine, dinucleotide (NAD), and lipoate)

Question 22

Vitamins involved in pyruvate dehydrogenase cofactors

Answer 22

thiamine (Vit B1) TPP
riboflavin (Vit B2) FAD
niacin NAD
pantothenate coenzyme Athiamine (Vit B1) TPP
riboflavin (Vit B2) FAD
niacin NAD
pantothenate coenzyme A

Question 23

thiamine deficiency

Answer 23

Wernickes encephalopathy- inability to oxidize pyruvate due to thiamine deficit. Affects brain most b/c this organ derives most of its energy from aerobic oxidation of glucose.

Question 24

regulation of pyruvate dehydrogenase

Answer 24

allosteric regulation - AMP, CoA and NAD+ activate it. ATP, NADH deactivate it. Second level occurs by phosphorylation of PDH by protein kinase which inactivates PDH, or de-phosphorylation of PDH by phospho-protein phosphatase which activates PDH. calcium activates the phosphatase

Question 25

regulation of pyruvate dehydrogenase in fed vs fasting state

Answer 25

fed: active in de-phospho state, when insulin and ADP are high and glucagon is low. Glucagon causes phosphorylation and insulin causes dephosphorylation. Fasted: PDH is inactive in phosphorylated state, when ATP is high and pyruvate is low.

Question 26

Function of TCA cycle

Answer 26

Oxidation of glucose to pyruvate has not recovered all of the energy stored in glucose. Acetyl CoA is completely oxidized in the presence of O2 to generate the ATP necessary for energy requiring cellular processes

Question 27

Overall result of TCA cycle

Answer 27

For each turn one Acetyl CoA enters and 2 CO2 leave. Oxaloacetate is regenerated at end so no net removal occurs.

Question 28

List key steps of TCA cycle

Answer 28

acetyl CoA > citrate (citrate synthase) > alpha-ketoglutarate + CO2 + NADH > succinyl CoA + second CO2 + NADH >succinate + GTP (substrate level phosphorylation) > fumarate (succinate dehydrogenase) > malate > oxaloacetate + third NADH >oxaloacetate continues cycle

Question 29

What intermediate leaves TCA cycle to form fatty acids in de novo lipogenesis

Answer 29

citrate

Question 30

List compounds which serve as entrance points into TCA cycle for some amino acids

Answer 30

alpha ketoglutarate and succinyl CoA and fumarate

Question 31

Which part of the TCA cycle is also part of the electron transport chain?

Answer 31

oxidation of succinate to fumarate via succinate dehydrogenase. Enzyme bound FAD is the electron acceptor and the electrons from FADH2 are directly passed to coenzyme Q in electron transport chain.

Question 32

Location of enzymes in TCA cycle

Answer 32

all are in mitochondrial matrix

Question 33

role of TCA cycle in anabolic pathways

Answer 33

During fasting, gluconeogenic precursors are converted to malate, which leaves the mitochondria to enter the pathway of gluconeogenesis in the cytosol. Plus, other TCA intermediates are channeled into aa, fatty acid, etc, synthesis

Question 34

regulation of TCA cycle

Answer 34

Enzymes in the TCA cycle are regulated by allosteric and covalent modifications

Question 35

Which enzymes are most regulated in the TCA cycle

Answer 35

enzymes that catalyze the 3 strongly exergonic steps: citrate synthase, isocitrate dehydrogenase and alpha-ketoglutarate dehydrogenase