L16: Glycolysis/CHO metabolism II

Question 1

What is substrate-level phosphorylation? At what step / reaction is this occurring in glycolysis?

Answer 1

• Substrate-level phosphorylation refers to the production of ATP within the cytoplasm from ADP, without the need to move electron carriers to the mitochondrial for ETC and oxphos.

Question 1

Describe the clinical presentation of pyruvate kinase deficiency.

Answer 1

• In this disorder, RBCs are deprived of ATP (only get 1 ATP instead of 2 from glycolysis), which leads to lysis of the cells (chronic hemolytic anemia) because the membrane potential cannot be maintained. These patients present at pale, jaundiced, fatigued, SOB, tachycardic. They have splenomegaly, excess of iron in blood and have gallstones. Severe cases require pts to have regular blood transfusions.

Question 2

What is the main regulated step in glycolysis?

Answer 2

• Phosphofructokinase

Question 3

How does ADP affect regulation of glycolysis?

Answer 3

• ADP stimulates PFK1 stimulating glycolysis

Question 3

What are disorders that prevent proper use of fructose? Explain each

Answer 3

• Essential fructosuria: absence of fructokinase (normally in liver), prevents uptake of fructose. Benign. Fructose can still be used by muscle or become excreted. - Hereditary fructose intolerance: deficiency in aldolase B enzyme. F1P accumulates in liver causing depletion of liver phosphate pools, preventing liver from breaking down glycogen, causing liver to be damaged by accumulation of glycogen, lack of Pi and synthesis of ATP. Pts are required to avoid fructose and sucrose (disaccharide of fructose and glucose)

Question 4

Explain how F26BP affects glycolysis. Include details about the enzymes involve, pathways activated and substrates affected.

Answer 4

• Take home message: high concentrations of F26BP stimulate glycolysis; low concentrations of F26BP inhibit glycolysis - F26BP is produced by enzyme PFK2 (produces F26BP from F6P) - F26BP activates PFK1 leading to increased production of F16BP - F26BP inhibits fructose bisphosphatase leading to increased production of F16BP - Insulin stimulates PFK2 via cAMP leading to increased concentration of F26BP, which stimulates PFK1, inhibits bisphosphatase and stimulating glycolysis - Glucagon inhibits PFK2 via cAMP leading to decreased concentration of F26BP, which means that PFK1 is inhibited, bisphosphatase is stimulated and glycolysis is inhibited - AMP stimulates PFK2, causing increased concentrations of F26BP, which stimulates PFK1, inhibits bisphosphatase and stimulated glycolysis

Question 5

How does AMP affect regulation of glycolysis?

Answer 5

• AMP stimulates PFK1 stimulating glycolysis

Question 5

Where is fructose metabolized? What enzymes are responsible for this? What are the clinical implications of fructose metabolism?

Answer 5

• Fructose is primarily metabolized in the liver via enzyme fructokinase - Also minimally metabolized in muscle via hexokinase. - Clinical implications: fructokinase reaction is insulin indepenent, meaning diabetics can tolerate dietary fructose as well as “normal” individuals and second, fructose from diet is metabolized immediately and gives a quicker boost of energy than glucose.

Question 6

What is hereditary fructose intolerance?

Answer 6

• deficiency in aldolase B enzyme. F1P accumulates in liver causing depletion of liver phosphate pools, preventing liver from breaking down glycogen, causing liver to be damaged by accumulation of glycogen, lack of Pi and synthesis of ATP. Pts are required to avoid fructose and sucrose (disaccharide of fructose and glucose)

Question 7

How does insulin affect regulation of glycolysis?

Answer 7

• insulin stimulates hexokinase stimulating glycolysis - insulin stimulates PFK1 via F26BP (increases its concentration_ stimulating glycolysis - insulin stimulates pyruvate kinase stimulating glycolysis

Question 7

Describe/Draw how fructose is degraded.

Answer 7

Question 8

What is the glycerol phosphate shuttle system? Where does it occur? Draw it.

Answer 8

• It is a shuttle mechanism to move electrons from NADH in cytoplasm into the mitochondrion. It occurs in brain and muscle.

Question 10

How does glucagon affect regulation of glycolysis?

Answer 10

• Glucagon inhibits hexokinase inhibiting glycolysis - Glucagon inhibits PFK1 via F2BP (decreases its concentration) inhibiting glycolysis

Question 11

How does ATP affect regulation of glycolysis?

Answer 11

• ATP inhibits PFK1 inhibiting glycolysis - ATP inhibits pyruvate kinase inhibiting glycolysis

Question 12

What is the Cori Cycle? What is its function/purpose?

Answer 12

• Cori cycle refers to process of utilizing lactate, produced via glycolysis in RBCs and muscle cells, by the liver to generate more glucose via gluconeogenesis.

Question 14

How does acetyl-CoA affect regulation of glycolysis?

Answer 14

• acetyl-CoA inhibits hexokinase inhibiting glycolysis

Question 16

What are the regulatory points in glycolysis? What factors are stimulating (+) and inhibiting (-)? What is the rate-limiting step in glycolysis?

Answer 16

• Regulatory points: a.) Hexokinase (+: insulin; -: G6P, acetyl-CoA, glucagon) b.) phosphofructokinase 1 (+: hormones via F26BP, ADP, AMP; -: ATP and citrate) c.) pyruvate kinase (+: F16BP and insulin; -: ATP) - Rate-limited step = phosphofructokinase 1

Question 17

How is NADH shuttled into the mitochondrion?

Answer 17

• Glycerol phosphate shuttle (used in muscle and brain) - Malate-aspartate shuttle (used in liver and heart)

Question 17

Can NADH move directly into the mitochondria from the cytoplasm?

Answer 17

• No, requires shuttle. Either glycerol phosphate shuttle or malate-aspartate shuttle.

Question 18

Under what conditions will lactic acid fermentation occur in the body? Why? What enzyme is involved? Why is this problematic in the body?

Answer 18

• Concentration of NAD to NADH are tightly monitored. Reason: NAD+ is limited in concentration. NADH must be recycled to NAD+ in order for glycolysis to continue. Lactic acid fermentation takes the electrons from NADH and places them onto pyruvate, generating lactate. This is done by lactate dehydrogenase. - Lactic acid buildup leads to lactic acidosis. Cori cycle tries to circumvent this from occurring.

Question 19

Describe the effect of arsenic on glycolysis.

Answer 19

• Arsenic mimics phosphate. In its presence, Glyceraldehyde-3-phosphate DH reaction yields arsenic product instead of 13BPG. The arsenic product is unstable and hydrolyzes to 3PG. No ATP is gained from the process, where previously the conversion to 3PG yields a single ATP. Net ATP from glycolysis with one molecule of glucose is 0. This is problematic for RBCs who rely solely on glycolysis for their energy production. - Pts present with SOB and dizziness. CBC reveals hemolytic anemia and elevated urine arsenic levels. Treatment includes discontinuation of arsenic and therapeutic red-cell exchange.

Question 20

What is the malate-aspartate shuttle? Where does it occur? Draw it.

Answer 20

• It is a shuttle mechanism to move electrons from NADH in cytoplasm to the mitochondrion. It occurs in liver and heart.

Question 22

Describe/Draw how galactose is degraded.

Answer 22

• Galactose is C4 epimer of glucose, so purpose of this reaction is to epimerize back to glucose.

Question 23

Which reactions in glycolysis use ATP, produce ATP and produce NADH? What is the net production of ATP from 1 molecule glucose?

Answer 23

• use of ATP: hexokinase (produce G6P from glucose) and phosphofructokinase (produce F16BP from F6P) - produce ATP: phosphoglycerate kinase (produce 3PG from 13BPG) and pyruvate kinase (produce pyruvate from phosphoenolpyruvate) - produce NADH: glyceraldehyde phosphate DH (produce 13BPG from dihydroxyacetonephosphate) - net ATP: 2 - net NADH: 1

Question 24

How does G6P affect regulation of glycolysis?

Answer 24

• G6P inhibits hexokinase inhibiting glycolysis

Question 25

What is essential fructosuria?

Answer 25

• absence of fructokinase (normally in liver), prevents uptake of fructose. Benign. Fructose can still be used by muscle or become excreted.

Question 26

What are/is disorder(s) that prevent proper use of galactose? Explain.

Answer 26

• Galactosemia - Three types: a.) Classic galactosemia: most common / severe form where there is a galactose-1 phosphate uridyl transferase deficiency b.) Deficiency of galactokinase c.) Deficiency of UDP-galactose epimerase - Results in accumulation of galactose1-phosphate in liver and other tissues (CNS, kidney). Newborns present with milk intolerance and signs of liver failure, cataracts and intellectual disability. Also present with jaundice, lethargy and hepatomegaly. Diagnosis is by detection of galactose or galactose-phosphate in urine. Pts must receive a galactose-free diet (no lactose either, which is a glucose, galactose disaccharide).

Question 28

Draw the glycolytic pathway. Include all the enzymes, where ATP is made or used and where reducing equivalents are produced.

Answer 28

Question 29

What problems in glycolysis affect RBCs?

Answer 29

• Arsenic poisoning - Pyruvate kinase deficiency

Question 30

How does citrate affect regulation of glycolysis?

Answer 30

• Citrate inhibits PFK1 inhibiting glycolysis

Question 31

What are all the molecules that inhibit/stimulate enzymes for the purposes of regulating glycolysis?

Answer 31

• insulin, glucagon, acetyl-CoA, G6P, F26BP, ADP, AMP, ATP, citrate

Question 32

What is galactosemia?

Answer 32

• There are 3 kinds depending on the enzyme deficiency: Galact-1-P uridyl transferase, galactokinase or UDP-galactose epimerase. - Results in accumulation of galactose1-phosphate in liver and other tissues (CNS, kidney). Newborns present with milk intolerance and signs of liver failure, cataracts and intellectual disability. Also present with jaundice, lethargy and hepatomegaly. Diagnosis is by detection of galactose or galactose-phosphate in urine. Pts must receive a galactose-free diet (no lactose either, which is a glucose, galactose disaccharide).