Glycolysis And Hemolysis

Question 1

What is the fate of absorbed glucose?

Answer 1

Glucose from the bloodstream enters the cells via specific transporters (GLUT) by facilitated diffusion)

GLUT 2- liver
GLUT 1 and 3- in neurons and brain
GLUT 1 in erythrocytes
GLUT 4 in adipose tissue and muscle; responsive to insulin. Number of GLUT 4 increase in presence of insulin, when the plasma glucose level is elevated

Question 2

How is glucose 6 phosphate formed?

Answer 2

-Glucokinase: liver has high Km for glucose (more active when plasma glucose is elevated). Also present in B-cells of pancreas.

Other tissues (RBC)- contain Hexokinase: low km for glucose

Question 3

What are the intracellular fates of Glucose 6 phosphate?

Answer 3

Pyruvate and HMP shunt

Question 4

What is the purpose of glycolysis ?

Answer 4

Principal pathways for generating ATP(energy)
-In brain and erythrocytes

Generates ATP in presence and abundance of O2 (aerobic and anaerobic) and in presence and absence of mitochondria

Red blood cells have no mitochondria
-Glycolysis is the only source of ATP in these cells

Glycolytic enzymes are present in the cytosol
Remember: Enzymes of pentose phosphate pathway are also present in cytosol

Question 5

Give an overview of the phases of glycolysis

Answer 5

Step 1:
Glucose—> fructose 1,6 biphosphate- uses 2 ATP

Step 2:
Cleavage of 6C sugar into 3 carbon intermediates:
Fructose 1,6 biphosphate—> dihydroxyacretone phosphate+ glyceraldehyde 3-phosphate

Step 3:energy generation phase
2 ATP formed
 NADH formed from each
Glyceraldehyde 3-phosphate—> Pyruvate 
Dihydroxyacetone phosphate—> Pyruvate

Question 6

What are the irreversible/ regulated reactions of glycolysis ?

Answer 6

• Two phosphorylation reactions require ATP (energy investment phase)- requires 2 ATP
• Glucokinase/Hexokinase
• PFK- 1: irreversible reaction of glycolysis; regulated step (allosteric regulated enzyme )
• Pyruvate kinase: is also irreversible

Question 7

What are the energy generation: substrate level phosphorylation reactions ?

Answer 7

• Reactions directly form ATP from ADP
• DO NOT require the electron transport chain or mitochondria or oxygen
• ONLY reactions that form ATP in cells that lack mitochondria and in hypoxia conditions
• Forms ATP directly from high energy compounds (1,3-Bisphosohoglycerate and Phosphoenolpyruvate)

Question 8

Explain the fates of Pyruvate

Answer 8

1. Aerobic (abundance of oxygen and mitochondria)—>. Acetyl CoA(Pyruvate dehydrogenase)—> enters TCA cycle
2. Anaerobic (no mitochondria OR no oxygen)—> lactate (lactate dehydrogenase-cytosolic enzyme)—> goes to liver via cori cycle

Absence of mitochondria (red blood cells, lens, cornea, leukocytes)

Poorly vascularized tissues or lack of oxygen (actively contracting white skeletal muscle, cardiac muscle hypoxia) to lactate (lactic acid)

Question 9

Describe the anaerobic fate of Pyruvate

Answer 9

• NADH formed by glyceraldehyde 3-phosphate dehydrogenase reaction is reoxidized to form NAD+
  
  • Allows glycolysis to proceed even in absence of mitochondria

-LDH is cytosolic enzyme

• Direction of reaction depends on NADH/NAD+ ratio
  
  • Higher NADH levels, favors lactate formation

Identify states that result in high NADH/NAD+ ratio

Question 10

Summarize lactate formation by anaerobic glycolysis in muscle

Answer 10

• In actively contracting skeletal muscle, NADH formed as a result of glycolysis and TCA cycle, exceeds oxidative capacity of ETC, resulting in a high NADH/NAD+ ratio
  
  • Favors lactate formation via lactate dehydrogenase
  • Lactate (lactic acid) in muscle causes a drop in pH and causes cramps

Cardiac muscle has low NADH/NA+ ratio and is generally aerobic (abundant mitochondria and good vascularization)

• Converts lactate to Pyruvate and later to Acetyl CoA, to enter TCA cycle
• Cardiac muscle hypoxia results in lactate formation (myocardial infarction)

Question 11

What is the cori cycle?

Answer 11

The fate of lactate

Lactate from muscle and red blood cells, diffuses out of these cells and is taken to liver where it is used for gluconeogenesis (cori cycle)

Question 12

Describe the energetic of glycolysis under anaerobic conditions

Answer 12

Abaerobic:
Glucose—>. 2 moles of lactate

2 NADH + H+ (formed in glyceraldehyde 3 phosphate dehydrogenase reaction) are used in lactate dehydrogenase reaction

4 ATP (formed in substrate level phosphorylation reactions) (2 ATP from each mol of glyceraldehyde 3 phosphate)

• 2 ATP used in investment phase
• Total = 2 ATP (anaerobic glycolysis)

Question 13

What are the inhibitors of glycolysis?

Answer 13

• In clinical lab, blood collected in fluoride containing tubes for estimation of plasma glucose
• If fluoride is not added, plasma glucose is used by RBC and WBC, resulting in erroneously low values

Question 14

Explain the significance of glycolysis

Answer 14

Tissues in which complete oxidation of glucose takes place (brain, skeletal muscle), glucose is converted to Pyruvate, later converted to Acetyl CoA, that enters TCA cycle for complete oxidation of glucose

Tissues of eye (lack mitochondria): cornea, lens
Tumor tissues uses glycolysis
In red blood cells , glycolysis (anaerobic conditions)
-major source of energy (ATP)
-forms 2,3 Biphosphoglycerate

Question 15

What what are the metabolic changes in cancer cells?

Answer 15

Cancer cells use glycolysis as source of ATP(energy)- ‘Warburg effect’

Glucose analog ‘flurodeoxyglucose(FDG)’ used in positron emission tomography
-increased uptake and utilization of FDG by tumor

Inhibitors of glycolysis are explored for use as therapeutic agents in cancers

Question 16

What is the significance of 2,3 BPG?

Answer 16

• 2,3-BPG binds to hemoglobin and helps to unload Oxygen in tissues
• 2,3-BPG levels are increased at high altitude
• decreases Hb oxygen affinity

Question 17

How is 2,3-BPG made?

Answer 17

1,3 -BPG is usually converted to 3-phosphoglycerate by phosphokinase

But mutate converts 1,3-BPG to 2,3-BPG

Question 18

What is the significance of glycolysis in RBCs: energy?

Answer 18

Glycolysis is the only source of energy (ATP) fir RBCs

• mature RBCs lack mitochondria and are completely dependent on glycolysis for ATP formation
• ATP for RBCs is required for maintenance of Na-K ATPase and maintenance of osmotic balance within RBCs

Hexokinase or Pyruvate kinase deficiency result in hemolytic

Question 19

What is hemolytic anemia?

Answer 19

• Erythrocyte Pyruvate kinase deficiency
• ATP required in RBCs to maintain electrolyte concentration of RBCs (operation of Na+-K+ ATPase)

Pyruvate kinase deficiency: second most common form of hemolytic anemia due to RBC enzyme deficiency (most common: G6PD deficiency)

Variants of enzyme have altered kinetic properties, or amount of the enzyme may be lower

Compare 2,3-BPG levels in Pyruvate kinase and Hexokinase deficient RBCs

Question 20

What is the mechanism of hemolysis in Pyruvate kinase/Hexokinase deficiency?

Answer 20

1. Defect in glycolysis (Pyruvate kinase/Hexokinase deficiency)
2. Reduced energy generation
3. Erythrocytes are energy deficient
4. Na+/K+. ATPase is compromised (Remember: 40-50% energy in resting cells is used for maintenance of Na+/K+ ATPase)
5. Osmotic imbalance (Na+ content in affected RBCs higher than in normal cells)
6. RBC lysis

Question 21

Lactic acidosis is observed when…

Answer 21

• increased NADH/NAD ratio: increases anaerobic respiration
• Inherited deficiency of Pyruvate dehydrogenase

-Decreased blood supply due to cardiac failure: Anaerobic metabolism in peripheral tissues. Defect in Cori cycle and lactate is not efficiently transported to liver

• Pulmonary embolism (poor oxygenation)
• Hemorrhage and circulatory shock

Question 22

What is the source 9f regulation of glycolysis in the muscle?

Answer 22

Energy deficient state(contracting muscle)—> increased AMP—> PFK-1

Energy excess state —> increased ATP

PFK-1: allosterically regulated enzyme

Regulation in muscle:
High levels of ATP(inhibits glycolysis)
High levels of AMP(stimulates glycolysis)