Exam 2 Flashcards
1
Q
Acetyl-CoA. a.) What is its structure? b.) What is its metabolic importance? c.) What is its source? Where does it come from?
A
- a.) see picture - b.) It is an important intermediate and precursor molecule for TCA, FA biosynthesis, ketone body formation and cholesterol biosynthesis (leading to steroid hormones, bile acids and vit D production) - c.) It is produced as a result of glycolysis, beta-oxidation and AA deamination/oxidation. It is requires presence of vitamin B5.
1
Q
Pyruvate. a.) What is the fate of pyruvate? b.) What enzyme converts pyruvate to acetyl-CoA? c.) What enzyme converts pyruvate to oxaloacetate? d.) What determines whether pyruvate is used to form acetyl-CoA or oxaloacetate?
A
- a.) lactate, oxaloacetate, acetyl-CoA and alanine - b.) PDH complex - c.) pyruvate carboxylase - d.) pyruvate carboxylase is allosterically activated by acetyl-CoA. When there is sufficient amount of acetyl-CoA and therefore little need to divert more pyruvate into acetyl-CoA, pyruvate carboxylase converts pyruvate to oxaloacetate.
1
Q
What is DNP? What is its effect? What symptoms does it cause?
A
- It is a pesticide and poison that uncouples ETC/ox-phos. It causes sweating, flushing, nausea, inc RR, tachycardia, fever, coma, death in 1-2 days. Treatment with ice baths, oxygen and fluid/electrolyte replacement.
1
Q
Where are the following glucose transporters found? What are their functions?
A
- SGLT1: found in small intestine, responsible for actively transporting glucose from lumen into intestinal epithelia. - GLUT1: found in all tissues, responsible for basal glucose uptake - GLUT2: found in liver, intestine and beta-cells of pancreas. In liver: removes glucose from blood. In intestine: releases glucose from epithelia into blood. In pancreas: regulates secretion of insulin. Also able to move fructose. - GLUT3: found in all tissues, responsible for basal glucose uptake - GLUT4: found in muscle and adipose tissue. Increases with endurance training, induced by insulin. - GLUT5: found in small intestine, responsible for uptake of fructose into epithelial cells and movement into blood serum.
1
Q
Which enzymes of gluconeogenesis are regulated? What factors inhibit and stimulate these enzymes? Which is the main regulated step? Which are reversible/irreversible?
A
- Pyruvate carboxylase: +: acetyl-CoA; -: insulin - PEP carboxykinase: +: glucagon via cAMP; -: insulin, AMP - Fructose-1,6-bisphosphatase: +: citrate; -: F26BP, AMP (main regulated step) - Glucose-6-phosphatase: +: glucagon; -: insulin
1
Q
What is MEOS? Explain.
A
- Microsomal ethanol oxidizing system. This is a P450 cytochrome system that is induced in chronic alcohol abuse and has a higher capacity to process alcohol. Activation of this system causes oxidation of NADPH and weakens the cellular antioxidant defense mechanisms.
1
Q
Explain how lipids are transported into the mitochondria. Is it in the form of TAGs or FAs?
A
- FAs are transported into the mitochondria.
1
Q
Which intermediate during ketone body synthesis can function in another pathway (besides acetyl Co-A)? What pathway? During what environmental conditions will this occur?
A
- HMG-CoA = beta-hydroxy-beta-methylglutaryl-CoA - This molecule is used in the synthesis of cholesterol. - Will occur in well-fed times
1
Q
What is the enzyme that commits acetyl-CoA to FA synthesis? What does this reaction do?
A
- Acetyl-CoA carboxylase, which converts acetyl-CoA to malonyl-CoA
1
Q
Describe the synthesis of FAs.
A
1
Q
Draw function of glucagon presence (insulin absence) in regulation of FA synthesis.
A
1
Q
Describe the composition of lipid droplets?
A
- Lipid droplets are packages containing TAGs. These are surrounding by a protein coat composed of many proteins including perilipin. Perilipin acts as protector of the TAGs stores and prevents lipases, such as hormone sensitive lipases from degrading TAGs into FFA and glycerol. When perilipin is phosphorylated, it allows lipases to do this, but not when dephosphorylated.
1
Q
What are the three factors that lead to development of fatty liver in chronic alcoholism?
A
1.) alcohol metabolism in liver generates NADH and acetyl-CoA. High concentration of NADH blocks TCA cycle enzymes and forces acetyl-CoA into FA synthesis. 2.) Beta-hydroxyacyl dehydrogenase (beta-oxidation enzyme) requires NAD, which is at low concentration during alcohol metabolism. This means that FA breakdown by liver is slowed down and leads to accumulation of fat. 3.) Damaged liver tissue has reduced capacity to synthesize VLDLs to export FAs to adipose tissue
1
Q
Describe synthesis of prostaglandins, thromboxanes and leukotrienes.
A
1.) Prostaglandins - AA to PGG2 via COX1/2 - PGG2 to PGH2 via PGH synthase - PGH2 is precursor to other prostaglandins via PGD/PGE/PGF synthase enzymes 2.) Thromboxanes - PGH2 to TXA2 via thromboxane synthase - TXA2 to TXB2 via hydrolysis in blood 3.) Leukotrienes - AA to leukotrienes via lipoxygenase and other enzymes
1
Q
How can prostaglandin synthesis be inhibited? Explain
A
- NSAIDs and glucocorticosteroids(aka glucocorticoids) inhibit COX1/2 and prostaglandin synthase (PGH synthase)
1
Q
Name the sphingolipidoses. Name the enzyme defect, accumulated lipid and presentation.
A
1.) Tay-Sachs Disease Enzyme defect: beta-hexosaminidase A Accumulated lipid: ganglioside GM2 Presentation: mental retardation, blindness, cherry red spot on macula, death before age 3 2.) Gaucher Disease Enzyme defect: beta-glucosidase (beta-cerebrosidase) Accumulated lipid: glucocerebroside Presentation: liver and spleen enlargement, erosion of long bones 3.) Fabry Disease Enzyme defect: alpha-galactosidase Accumulated lipid: ceramide trihexoside Presentation: skin rash, kidney failure 4.) Niemann-Pick Disease Enzyme defect: sphingomyelinase Accumulated lipid: sphingomyelin Presentation: liver and spleen enlargement, mental retardation 5.) Sandhoff Disease Enzyme defect: beta-hexosaminodase A and B Accumulated lipid: GM2 ganglioside and globosides Presentation: similar to Tay-Sachs, progresses more rapidly 6.) Metachromatic Leukodystrophy Enzyme defect: arylsulfatase Accumulated lipid: sulfatide Presentation: mental retardation
1
Q
Where is cholesterol synthesized, what is the rate-limiting step/enzyme, what is the precursor used, in what other pathway is this precursor seen? What enzyme is used to make this precursor? Is this the same enzyme used in the previous pathway? Explain. During what environmental conditions is the precursor made in both pathways?
A
- Cholesterol synthesis takes place in the liver, intestine and reproductive tissues - Rate-limiting step = HMG-CoA reductase - Precursor = HMG-CoA – seen during synthesis of ketone bodies - Precursor synthesized by cytosolic HMG-CoA synthase isoform that is active in well-fed state. Mitochondrial HMG-CoA synthase isoform synthesizes the precursor when in starving/fasting state.
1
Q
How can cholesterol synthesis be inhibited in a clinical setting? Explain how this works.
A
- Use of statins - Statins inhibit the HMG-CoA reductase enzyme and prevents synthesis of mevalonic acid, a precursor molecule to farnesyl-PP, a precursor to cholesterol.
1
Q
T2D patients present with high serum LDL even when serum glucose is well controlled. Where does this lipid abnormality originate? Why does it occur?
A
- Type 2 diabetics are insulin insensitive - As a result of insulin signaling stating to the body that glucose is in abundance and should be taken up, fatty acids are still being released from adipose tissue and the liver has to repackage large amounts of these, which in turn means that LDL levels are high.
2
Q
What is the malate-aspartate shuttle? Where does it occur? Draw it.
A
- It is a shuttle mechanism to move electrons from NADH in cytoplasm to the mitochondrion. It occurs in liver and heart.
2
Q
What are/is disorder(s) that prevent proper use of galactose? Explain.
A
- 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).
2
Q
Explain how F26BP affects gluconeogenesis. Include details about the enzymes involve, pathways activated and substrates affected.
A
- Take home message: high concentrations of F26BP inhibit gluconeogenesis, low concentrations of F26BP stimulate gluconeogenesis - F26BP is produced by enzyme PFK2 (produces F26BP from F6P) - Insulin stimulates PFK2 via cAMP leading to increased concentration of F26BP, which causes inhibition of gluconeogenesis via pyruvate carboxylase, PEP carboxykinase and glucose-6-phosphatase - Glucagon inhibits PFK2 via cAMP leading to decreased concentration of F26BP, which causes stimulation of gluconeogenesis via PEP carboxykinase and glucose-6-phosphatase
2
Q
Explain why NADH concentration increases while NADPH concentration decreases in alcohol-metabolizing cells.
A
- The main step to handling ethanol in the liver is through two dehydrogenases (alcohol DH and aldehyde DH), both, which produce NADH. - The other way to handle ethanol is through the higher capacity MEOS system, which as a means to process the alcohol, also oxidizes NADPH.
2
Q
List a few mucopolysaccharidoses. Describe defect and symptoms. Genetics?
A
- Hunter’s: defect in iduronate sulfatase, accumulation of dermatan sulfate and heparan sulfate. Symptoms = skeletal abnormalities, mental retardation. X-linked recessive. - Hurler-Scheie: defect in alpha-iduronase, accumulation of dermatan sulfate and heparan sulfate. Symptoms = skeletal abnormalities, mental retardation. Autosomal recessive. - Sanfilippo’s: defect of heparan sulfate degradation. Symptoms = mild physical defects, severe mental retardation. Autosomal recessive.
2
Why aren’t ketone bodies used in liver or kidney?
- These tissues don’t express acetoacetate:succinyl-CoA transferase to generate the intermediate: acetoacetyl-CoA that generates acetyl-CoA.
2
Describe important of asymmetric distribution of lipids between lipid bilayers. What is typical distribution? Implication in RBCs
- Outer leaflet is rich in PC and sphingomyelin (both have choline) - Inner leaflet is rich in PE and PS - Stress activates enzymes known as flippases that flips lipids between leaflets - Eg. When PS appears in outer leaflet of RBCs, RBCs are destroyed by macrophages.
2
What are the function of leukotrienes?
- promotion of SM contraction - anaphylaxis
4
What enzyme(s) require(s) thiamin (B1)?
- Alpha-ketoglutarate dehydrogenase complex and PDH complex.
4
Explain what reaction(s) the cell can perform when there is a need for NADPH and ATP. Ribose is not required.
4
Explain the effect of insulin-induced dephosphorylation on glycogen phosphorylase and glycogen synthase.
- Glycogen phosphorylase is inactivated - Glycogen synthase is activated
4
Explain process of beta-oxidation. Include enzymes, substrates, cofactors and products.
4
Draw function of insulin in regulation of FA (TAG) synthesis.
4
Describe role of chylomicron, where synthesized, association with apoproteins, source of apoproteins, destination of contents and formation / fate of chylomicron remnant.
5
What is biotin? Which vitamin is it?
- Biotin is a B vitamin, also known as vitamine B7. It is required by pyruvate carboxylase.
5
Describe how xenobiotics are detoxified.
- Example: acetaminophen. - UGT catalyzes conjugation of UDP-glucuronate with acetaminophen, making it water soluble, allowing it to be excreted in urine
7
How does carnitine deficiency present? What are the consequences?
- Without carnitine, FAs cannot be imported into mitochondria to be used as energy source. - Pts present as fasting non-ketotic hypoglycemia. Ketones are unusually low in blood or / and urine.
8
Describe synthesis and sulfation of glycosaminoglycans.
- Core proteins are manufactured in ER - Transferred to Golgi where glycosyltransferases add disaccharides to them. - Sulfotransferases add sulfate groups to them (can be unpredictable) - Exported in membrane-bound vesicle, finds linker and attached to polysaccharide backbone
9
List endogenous uncoupling proteins. What are their functions?
- UCP1-4. These are found in brown fat, skeletal muscle, brain and other parts of the body and function to increase body temperature.
9
Describe/draw the reactions of glycogen degradation (glycogenolysis).
9
Explain the process of FA elongation from a 16 carbon molecule. Why is this necessary? Where does it occur?
- Cytoplasm has FA synthase that synthesizes FAs to 16 carbons = palmitate (16:0), which is a saturated molecule. - Many lipid structures in the body are longer than 16 carbons - Elongation occurs in mitochondrial as shown here:
9
Explain the process of FA desaturation. Why is this necessary? Where does it occur?
- Many lipids in body are not saturated. - Desaturation of FAs occur in endoplasmic reticulum as shown here:
11
What are the precursors for gluconeogenesis?
- Lactate, - Amino acids - Glycerol
12
What are the products of the non-oxidative phase of the pentose phosphate pathway? Why are these important?
- 2 F6P and 1 glyceraldehyde-3-phosphate - These are intermediates in the glycolytic / gluconeogenic pathways and can be synthesized from pentoses depending on the need of the cell.
13
Describe the clinical presentation of pyruvate kinase deficiency.
- 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.
14
Draw the glycolytic pathway. Include all the enzymes, where ATP is made or used and where reducing equivalents are produced.
15
Describe the effect of arsenic on glycolysis.
- 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.
16
Examples of exogenous uncouplers to ETC/ox-phos. Symptoms? Treatment?
- DNP (2,4 dinitrophenol) is a pesticide and poison. It causes sweating, flushing, nausea, inc RR, tachycardia, fever, coma, death in 1-2 days. Treatment with ice baths, oxygen and fluid/electrolyte replacement. - Aspirin in very high doses
17
Describe role of VLDL, IDL and LDL. Include where each are synthesized/formed, association with apoproteins, source of apoproteins, destination of contents and fate of lipoprotein.
19
Briefly explain the role of F26BP in gluconeogenesis and glycolysis.
- High F26BP concentrations: glycolysis is stimulated, gluconeogenesis is inhibited - Low F26BP concentrations: glycolysis is inhibited; gluconeogenesis is stimulated
21
What is the function of diacylglycerol acyltransferase (DGAT) inhibitors?
- Inhibits synthesis of triacylglycerol
21
How do gallstones form?
- Bile contains cholesterol, bile salts and phospholipids. These aid in emulsification of dietary fats. Cholesterol is prone to precipitating in gallbladder and duct works as its water solubility is low. When in high concentration, it particularly precipitates out causing formation of cholesterol stones known as gallstones, which can obstruct the gallbladder and ducts causing painful backup of bile of pancreatic enzymes.
23
Describe/Draw how fructose is degraded.
24
Describe/draw the reactions of glycogen formation (glycogenesis)
25
What apoproteins do VLDL associate with? IDL? LDL? Where?
- VLDL associates with ApoB100 in hepatocytes - VLDL in serum associates with apoE and apoC obtained from HDL - IDL remains associated with all 3 per above - LDL remains associated with all 3 per above
26
Describe/Draw the oxidative/non-oxidative phase reactions of the pentose phosphate pathway.
27
Describe how arachidonic acid is synthesized and inserted into phospholipids.
28
Explain what reaction(s) the cell can perform when there is a need for ribose, but not for NADPH. ATP is not required.
29
How do serum lactic acidoses occur as a result of defects to complexes I-IV, administration of DNP, administration of cyanide and lack of oxygen?
- Defects in complexes I-IV would result in “back up” of intermediate substrates / electron carrier molecules, including NADH. This would result in lack of NAD+, which is limited in supply. End result is that pyruvate builds up in the cell and is reduced to lactate via LDH enzyme. Lactate is an acid and therefore a lactic acidosis is occurring here. - DNP, cyanide and lack of oxygen cause the exact same thing to occur.
31
Explain what reaction(s) the cell can perform when there is a need for NADPH and glucose. ATP is not required.
33
What reaction does galactokinase catalyze?
- Galactose + ATP =(ez: galactokinase) Galactose-1-P
35
Pyruvate carboxylase. a.) What is the function of this reaction? b.) Write out the reaction is catalyzes. Include all energy components and coenzymes. c.) What is an allosteric activator of this enzyme? Why is that important?
- It is an anaplerotic reaction to regenerate oxaloacetate when levels are low. - Pyruvate + ATP + HCO3 = oxaloacetate + ADP + Pi + H+ (ez: pyruvate carboxylase, coenzyme: biotin) - Acetyl-CoA. When there is sufficient amount of acetyl-CoA and therefore little need to divert more pyruvate into acetyl-CoA, pyruvate carboxylase converts pyruvate to oxaloacetate.
35
Thiamin deficiency. a.) What vitamin is deficient? Where is this required? b.) Who does it affect? c.) What are the early symptoms? d.) What are the advanced / severe deficiencies of thiamin? Which groups are affected? What are the symptoms?
- a.) Vitamin B1. Required for PDH and alpha-KGDH enzymes to function. - b.) Primarily affects alcoholics, elderly and low income groups who are more likely to have poor diets. - c.) loss of appetite, constipation, nausea, fatigue, depression, peripheral neuropathy - d.) Wernicke-Korsakoff = advanced deficiency frequently seen in alcoholics: mental confusion, ataxia, loss of eye coordination Beri-Beri = severe deficiency seen in populations that heavily rely on polished rice diets: neuromuscular symptoms – muscle weakness, muscle atrophy, fatigue, peripheral neuropathy, lactic acidemia
36
What are the consequences of PDH deficiency? Population affected? Symptoms?
- Leads to chronic lactic acidosis, primary in kids. It leads to severe neurological problems and is frequenctly fatal.
38
What are disorders that prevent proper use of fructose? Explain each
- 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)
40
Explain mechanism that brings acetyl-CoA to cytoplasm from mitochondrion that preceeds FA synthesis?
42
What apoproteins do HDL associate with? Where?
- Nascent HDL associates with apoE,C and A in hepatocytes.
43
How do high levels of NADH prevent gluconeogenesis from proceeding?
- Malate-oxaloacetate shuttle. When levels of NADH are high, it will donate its electrons to oxaloacetate to form malate, which moves into mitochondria and reconverts to oxaloacetate and NADH in the matrix. This ensures low oxaloacetate in the cytosol and inhibition of gluconeogenesis.
44
What is the function of cyanide? What are treatments?
- Cyanide inhibits cytochrome oxidase (complex IV) by binding Fe3+ and preventing reduction to Fe2+. It is fast acting and lethal. Causes lactic acidosis. Treatment: nitrites (oxidize Hb so cyanide binds to Hb instead of cytochrome oxidase) and Na thiosulfate (converts cyanide to non toxic SCN- form).
44
Describe reactions that form triacylglycerols.
45
ETC. a.) What are the components? What are the enzyme complexes involved? Name them b.) What reaction does each component catalyzed? List reduced substrates and oxidized products. c.) What is the sequence of flow through these carriers? Draw and indicate the reactions of each of the complexes. Include movement of protons. What coenzymes and metals do each of the complexes require? d.) Which components are proton pumps? e.) There are other dehydrogenases besides complexes I through IV contained in the mitochondrion. What are they?
- a.) dehydrogenase enzymes (complexes), heme-iron proteins (aka cytochromes), iron-sulfur proteins, CoQ/ubiquinone Dehydrogenase enzymes are: Complex 1: NADH-CoQ reductase Complex 2: Succinate-CoQ reductase Complex 3: CoQH2-cytochrome c reductase Complex 4: Cytochrome c oxidase - b.) Complex 1: NADH + H+ + CoQ = NAD+ + CoQH2 Complex 2: Succinate + CoQ = Fumarate + CoQH2 Complex 3: CoQH2 + 2 cyt c (Fe3+) = CoQ + 2 cyt c (Fe2+) + 2H+ Complex 4: 4 cyt c (Fe2+) + 4H+ + o2 = 4 cyt c (Fe3+) + 2H2o - c.) see picture - d.) Protons are moved to the intermembrane space via complex I, II and IV - e.) Fatty acyl-CoA DH and glycerol-3-P DH. These both reduce CoQ and are not part of the std I-IV complexes
47
Beri-beri. a.) What is it? What is it caused by? b.) What are the symptoms associated with it? c.) What are the two forms of this disorder? d.) What organ/tissue(s) are most affected by this disorder? e.) Do RBCs contribute to this condition? Explain. f.) Which populations are susceptible to this disorder?
- a.) It is a severe thiamin (B1) dietary deficiency. - b.) Neuromuscular symptoms – muscle weakness, muscle atrophy, fatigue, peripheral neuropathy and lactic acidemia. - c.) Dry beri-beri which affects neuromuscular function, does not lead to fluid retention; Wet beri-beri causes peripheral edema and cardiac failure. - d.) It affects the nervous system and musculoskeletal system primarily. - e.) No, as they do not contain mitochondria, where enzymes such as PDH complex and aKGDH that participate in linking glycolysis to TCA cycle and TCA cycle function respectively. - f.) Populations that have high intake of polished rice diets.
48
What is problematic about unsaturated and branched FAs in beta-oxidation? How are these handled?
- Unsaturated FAs: dbl bonds have to be moved around so enzymes used in beta-oxidation can recognize molecules and process them. a.) If dbl bond bw C3,4, isomerizes to trans-delta2-enoyl CoA b.) If dbl bond bw C4,5, reduced to trans-delta3-enoyl CoA, then isomerized to trans-delta2-enoyl CoA - Branched FAs: result from degradation of chlorophylls a.) requires peroxisomal alpha-oxidation, which starts with hydroxylation and ends with release of CO2
49
What are the functions of prostaglandins/thromboxanes?
- inflammatory mediators (vasodilation) - stimulation of contraction of uterus - platelet aggregation
51
Which reactions in glycolysis use ATP, produce ATP and produce NADH? What is the net production of ATP from 1 molecule glucose?
- 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
52
What is Wernicke-Korsakoff syndrome? What are the symptoms?
- It is an advanced thiamin (B1) deficiency that typically occurs in alcoholics. Symptoms include mental confusion, ataxia and loss of eye coordination.
53
What is the purpose of the pentose phosphate pathway? What are the products from this pathway used for?
- Way for generating pentoses and NADPH - Pentoses are used for synthesis of: DNA, RNA, ATP, NADH, FADH, Coenzyme A - NADPH are used for synthesis of: FAs, cholesterol, NTs, nucleotides and reduction of oxidized glutathione and P450 monooxygenases
55
Why is gluconeogenesis not a reversal of glycolysis? Explain. Explain what reactions occur to circumvent this issue.
- Glycolysis has 3 irreversible reactions: hexokinase, phosphofructokinase 1 and pyruvate kinase. In order for glucose to be synthesized, ie. gluconeogenesis, 4 new enzymes not seen in glycolysis are seen: Pyruvate carboxylase, PEP carboxykinase, Fructose-1,6-bisphosphatase and glucose-6-phosphatase.
57
Explain how ethanol is metabolized? What is its impact on CHO metabolism?
- Alcohol metabolism occurs in the liver by two mechanisms: 1.) two DH reactions (main pathway) and 2.) MEOS: microsomal ethanol oxidizing system: induced by chronic alcohol abuse. - Both DH reactions reduce NAD to NADH, which interferes with CHO metabolism and utilization. - When NADH is high in the cytoplasm, pyruvate and oxaloacetate are converted into lactate and malate respectively via lactate dehydrogenase and malate dehydrogenase respectively. Pyruvate and oxaloacetate are both intermediates in gluconeogenesis and glycolysis. - Lactate levels rise (more than normally) and cannot be processed in the Cori cycle, causing accumulation in blood, leading to metabolic acidosis. Body responds by increasing respiration in order to attempt restoral of acid/base balance. - Failure to complete Cori cycle also leads to epinephrine and glucagon release, which induces a stress response. - NADH inhibits key enzymes of TCA and shunts acetyl-CoA into FA/ketone body biosynthesis leading to a fatty liver over time.
59
What are the effects of insulin, glucagon and epinephrine on glycolysis, glycogenolysis and glycogenesis in liver and muscle? Tabulate.
62
TCA cycle. a.) What are the 2 major functions of the TCA cycle? b.) What is the overall balanced reaction of the TCA cycle? c.) Draw the reactions of the TCA cycle. Include enzymes, cofactors. d.) What is the fuel for the TCA cycle? e.) Which reaction(s) liberate CO2? f.) Which reaction(s) liberate NADH? g.) Which reaction(s) liberate FADH2? h.) Which reaction(s) involve substrate-level phosphorylation? i.) How many ATPs/ATP-equivalents are produced from a molecule of acetyl Co-A? From pyruvate? From glucose? j.) Explain mechanisms for TCA cycle regulation. What enzymes are regulated?
- a.) Serves to generate ATP and to generate biosynthetic precursors. - b.) CH3COSCoA + 2H2O + 3NAD+ + FAD + GDP + Pi = 2CO2 + 3NADH + 3H+ + FADH2 + CoASH + GTP - c.) see picture - d.) Acetyl-CoA - e.) isocitrate to alpha-ketoglutarate; alpha-ketoglutarate to succinyl-CoA - f.) isocitrate to alpha-ketoglutarate; alpha-ketoglutarate to succinyl-CoA; malate to oxaloacetate - g.) succinate to fumarate - h.) succinyl-CoA to succinate - i.) since 3 ATPS per NADH and 2 ATPS per FADH - 12 per acetyl CoA, 15 from pyruvate (considering PDH reaction) and 30 from glucose - j.) Regulation of the TCA cycle occurs through regulation of isocitrate dehydrogenase and alpha-ketoglutarase dehydrogenase complex. Isocitrate dehydrogenase activity increases when conc of ADP are high and decreases when NADH conc are high. Alpha-ketoglutarate dehydrogenase complex activity decreases when concentrations of succinyl-CoA and NADH are high.
64
How is the PDH complex similar or dissimilar to the alpha-ketoglutarate DH complex?
- Each complex has 3 enzyme subunits. Each produces CO2. Substrate for PDH is 3C alpha-keto acid (pyruvate), while substrate for aKGDH is 5C alpha-keto acid (alpha-ketoglutarate). Product for PDH is 2C (acetyl-CoA), while produce for aKGHD is 4C (succinyl-CoA). Regulation of PDH occurs via phosphorylation, while aKGDH is not regulated via phosphorylation. Reactions catalyzed by both are irreversible as a lot of free energy is lost.
66
List the enzymes involved in glycogen synthesis. Explain how each is regulated. What are the factors that stimulate and inhibit each.
- Hexokinase: +: insulin; -: G6P - Glucokinase: +: insulin - Phosphoglucomutase: not regulated - UDP-glucose pyrophosphorylase: not regulated - Glycogen synthase: +: insulin; -: glucagon, epinephrine - Branching enzyme: not regulated
69
What is Crigler-Najjar/Gilbert syndrome? Symptoms.
- Glucuronic acid is a prerequisite for bilirubin excretion. - UGT, which is an enzyme that catalyzes the conjugation of bilirubin with glucuronic acid leads to buildup of bilirubin and syndromes known as Crigler-Najjar/Gilbert. - Jaundice
69
Types of phospholipases and their functions?
- Phospholipase A1: removal of FAs from C1 of phospholipid - Phospholipase A2: removal of FAs from C2 of phospholipid, usually arachidonic acid - Phospholipase C: removal of phosphate head group from C3 of PIP2 phospholipid creating DAG and IP3
70
How does beta-oxidation differ in peroxisomes from other tissues?
- Beta-oxidation in peroxisomes involves preferential catabolism of LCFAs and produces hydrogen peroxide instead of FADH as seen in beta-oxidation in other tissue. Peroxisomes use oxidase enzyme
71
Explain the function and regulation of HSL.
- HSL=hormone sensitive lipase - In adipose tissue, HSL releases FFAs into blood by cleaving TAGs into glycerol and FAs - Inhibited by insulin
73
Describe synthesis of ketone bodies
74
What is the committed step in ketone body formation?
- Conversion of HMG-CoA to Acetoacetate via HMG-CoA lyase, which produces and acetyl-CoA. Procession through this step will commit the pathway to forming ketone bodies.
76
What are the products of the oxidative phase of the pentose phosphate pathway?
- Ribose-5-phosphate, 2 NADPH and CO2
78
Discuss regulation of FA (TAG) synthesis, degradation and mobilization/release
1. ) FA synthesis
- Well-fed state:
a. ) insulin: induces expression of acetyl-CoA carboxylase, FA synthase, malic enzyme and G6PD long-term basis
b. ) insulin: activates acetyl-CoA carboxylase and glycolysis (both via phosphoprotein phosphatase, which removes phosphates) – short term basis
c. ) citrate: activates FA synthase
- Starvation
a. ) absence of insulin: levels of acetyl-CoA carboxylase, FA synthase, malic enzyme and G6PD fall
b. ) glucagon: caused cAMP and PKA activity to rise.
c. ) PKA: inactivates acetyl-CoA carboxylase by phosphorylation, inhibits glycolysis
2. ) FA degradation
- Limiting step = bringing FA into mitochondria via carnitine shuttle / CPTI/II. This is inhibited by malonyl-CoA in well-fed state when FA synthesis is occurring.
3. ) FA mobilization
- High glucagon/low insulin stimulates mobilization: In fasting state, glucagon raises cAMP levels and activates PKA. PKA activates perilipin and hormone sensitive lipase both by phosphorylation (remember, glucagon acts via phosphorylation). This results in FAs being clipped off glycerols allowing them to become FFAs diffuse out of adipose tissue and move to other tissues. In other tissues they move into mitochondria via CPT I shuttle mechanism which are active as malonyl-CoA is low (this should only be occurring in starved state) and proceed through beta-oxidation.
79
What apoproteins do chylomicrons associate with? Where?
- Chylomicrons in enterocytes associate with ApoB-48 - After entering into serum, CM now associates with ApoC and ApoE, which is receives from HDL
81
List and describe the glycogen storage diseases (GSD). What is occurring in each and what is the clinical presentation?
- GSD-I: aka Von Gierke’s disease: deficiency in G-6-phosphatase. Leads to hypoglycemia and increased glycogen stores in liver. This ez is only found in liver. - GSD-II: aka Pompe’s disease: deficiency in alpha-glucosidase (aka acid maltase). It is required for the degradation of glycogen, which with the deficiency, slowly accumulates in the lysosome. Over time, this process eventually kills affected cells. Death occurs by cardiac or respiratory failure. - GSD-III: aka Cori’s disease: deficiency in debranching enzyme. Glycogen granules grow large because only the non-branched, outer layers of the deposits can be degraded. Presents in early childhood or infancy with hepatomegaly, hypoglycemia and growth retardation. Can also present with muscle weakness. Some pts develop cardiomyopathy. - GSD-V: aka McArdle’s disease: deficiency in glycogen phosphorylase in muscle. Muscle is unable to use glycogen. Leads to greater than normal glycogen stores in muscles. During periods of high glucose demand, ie. exercise, patients experience muscle cramps. - GSD-VI: aka Hers disease: deficiency in glycogen phosphorylase in liver. Presents with mild to moderate hypoglycemia, mild ketosis, growth retardation and prominent hepatomegaly.
82
What is the role of phospholipids in prevention of respiratory distress syndrome in premature infants?
- Surface of lung epithelium is covered with secretion that prevents alveolar membranes from sticking together (surfactant) - This surfactant is rich in phospholipids, particularly dipalmitoyl phosphatidylcholine (sometimes called dipalmitoyl lecithin). - This surfactant is produced at about 31 weeks gestation. - Infants born prematurely have a lecithin/sphingomyelin ratio below 2, indicating they have insufficient amounts of surfactant, which puts them at risk for respiratory distress syndrome (RDS). - Artificial surfactant mixtures treat premature infants. - Also they are given corticosteroid growth hormones to stimulate lung maturation.
83
Describe pathology of familial hypercholesterolemia. What is the presentation?
- Autosomal dominant disorder resulting from defects in LDL receptor. As a result, serum lipid concentrations are elevated and result in high levels of LDL in the blood serum. As a result of the overflow pathway into macrophages, atherosclerotic changes in blood vessels occur leading to higher incidences of CVD in the 3rd and 4th decades of life. - These individuals present with xanthomas (deposition of lipis) to joints and tendons. Lab studies indicate elevelated cholesterol and triglyceride levels.
86
Provide examples of defects in oxidative phosphorylation.
- LHON: Leber’s Hereditary Optic Neuropathy is due to mutations in subunits of complex I and is characterized by sudden-onset blindness in young adults. - MERRF: Myoclonic Epilepsy and Ragged Red Fibers is due to abnormally shaped mitochondria with dimished cytochrome oxidase (complex IV) activity as the result of a point mutation in the mitochondrial gene for lysine tRNA. It is characterized by myoclonus and ataxia with generalized seizures. - MELAS: Mitochondrial Encephalopathy, Lactic acidosis and Stroke-like activity is due to abnormally shaped mitochondria with normal cytochrome oxidase activity that is caused by a point mutation in the gene for leucine tRNA. Early symptoms include muscle weakness, pain, recurrent headaches, loss of appetite, vomiting and seizures. Stroke-like episodes begin before age 40. Episodes involve transient hemiparesis (temporary muscle weakness on one side of body), altered consciousness, vision abnormalities, seizures and severe headaches resembling migraines. Episodes lead to brain damage and in some cases dementia.
87
How is glycerol (or G3P) synthesized? What is this a precursor to?
- Glycerol-phosphate-dehydrogenase takes DHAP and converts it into glycerol-3-phosphate, which is precursor to TAGs.
88
What pentose phosphate pathway deficiency(ies) seen in human? What is/are the clinical presentation(s)?
- Beriberi: result of thiamine deficiency that in addition to affecting PDH and alphaKGDH, also affects the transketolase reactions of the non-oxidative phase of the PPPathway preventing rearrangement of carbon structures and leading to an imbalance in the CHO pool. Prevalent in low income communities, but most especially in alcoholic patients. Alcoholism reduces thiamine uptake and storage.
90
Explain how bilirubin is excreted. What occurs when there is a defect in this?
- Glucuronic acid is a prerequisite for bilirubin excretion. - UGT, which is an enzyme that catalyzes the conjugation of bilirubin with glucuronic acid leads to buildup of bilirubin and syndromes known as Crigler-Najjar/Gilbert.
90
Describe utilization of glucose by brain, RBCs, brain, liver, muscle and adipose.
- RBCs: glucose through glycolysis to produce lactate (used in liver), G6P into PPP - Brain: glucose through glycolysis to produce acetyl-CoA (oxphos), G6P into PPP. No production of lactate - Muscle/heart: glucose through glycolysis to produce both lactate (used in liver, ? produced in heart) and acetyl-CoA (oxphos), G6P into PPP, G1P as intermediate into glycogenolysis and glycogen synthesis - Adipose tissue: same as muscle/heart, except no lactate production AND acetyl-CoA used for FA synthesis in addition to oxphos. - Liver: glucose through glycolysis to produce pyruvate (to acetyl-CoA to oxphos or FA synthesis), pyruvate to OAA (to gluconeogenesis), G6P to PPP, G1P in glycogenolysis and glycogen synthesis. \* no exit of glucose to serum unless from liver.
91
What are the 2 sources of cytosolic NADPH?
- PPP via G6PD - Conversion of malate to pyruvate via malic enzyme
93
How is the pentose phosphate pathway regulated?
- Glucose-6-Phosphate Dehydrogenase is inhibited by NAPDH. When NADPH is high, G6P can be used elsewhere.
95
Describe role of HDL. Include where synthesized, association with apoproteins, source of apoproteins, destination of contents and fate of this lipoprotein.
96
PDH complex. a.) Describe its structure? b.) Where is it located? What is its function? c.) What are its coenzymes, vitamins? d.) What are its substrates and products? Show the balanced reaction it catalyzes. e.) Explain mechanisms for regulation of PDH complex. Explain when it is active and inactive.
- a.) PDH is a 3 enzyme complex. It contains a pyruvate decarboxylase (aka pyruvate dehydrogenase), a dihydrolipoyl transacetylase and a dihydrolipoyl dehydrogenase - b.) It is located in the mitochondrial matrix and serves to link glycolysis to the TCA cycle - c.) It requires 5 coenzymes: TPP, FAD/FADH2, NAD/NADH2, CoASH and lipoamide and 4 B-vitamins: thiamin (e) – B1, riboflavin – B2, niacin – B3 and pantothenic acid – B5. - d.) CH3COCOO- + NAD+ + CoASH = CH3COSCoA + CO2 + NADH + H+. The reaction is not reversible as the delta Go’ is too high. Too much free energy is lost. - e.) PDH is inactive when phosphorylated and is therefore activated by PDH phosphatase. PDH is deactivated when phosphorylated by PDH kinase. It is activated during times when ATP energy is needed: low concentrations of CoASH, pyruvate, ADP and NAD+ (substrates of reaction). These states deactivate the kinase, leading to active form of PDH. It is deactivated during times when ATP energy is not required: high concentrations of acetyl-CoA and NADH (products of reaction). These states activate the kinase, leading to inactive form of PDH.
97
What is the glycerol phosphate shuttle system? Where does it occur? Draw it.
- It is a shuttle mechanism to move electrons from NADH in cytoplasm into the mitochondrion. It occurs in brain and muscle.
98
What are disorders of FA degradation?
- Primary carnitine deficiency - Acyl-CoA dehydrogenase deficiency – eg. VLCAD, LCAD, MCAD and SCAD. MCAD is most common. - Refsum disease – alpha-oxidation deficiency - Peroxisome biogenesis disorders (PBD)
100
What is the function of farnesyl PP?
- converted into dolichol-PP for protein glycosylation - converted into coQ for electron transport - used for prenylation (post-translational) of proteins which influence water solubility of proteins
102
Describe/draw the synthesis of glucose from non-CHO precursors (gluconeogenesis).
103
List the enzymes involved in glycogenolysis. Explain what factors regulate each, ie. what inhibit and stimulate each?
- Glycogen phosphorylase: +: glucagon, epinephrine; -: insulin - Glucose-6-phosphatase: +: glucagon, epinephrine; -: insulin - Debranching enzyme: not regulated
104
Describe the synthesis of farnesyl PP.
105
Explain what reaction(s) the cell can perform when there is a need for both NADPH and ribose. ATP is not required.
107
Explain how F26BP affects glycolysis. Include details about the enzymes involve, pathways activated and substrates affected.
- 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
108
Explain the effect of glucagon-induced phosphorylation on glycogen phosphorylase and glycogen synthase.
- Glycogen phosphorylase is activated - Glycogen synthase is inactivated.
109
How does NADH:NAD+ ratio determine predominant forms of ketone bodies in liver?
- If NAD+ concentration is low in liver/kidney, acetoacetate will be converted to beta-hydroxybutyrate, causing an increased concentration of NAD+ and decrease in NADH.
111
How is cholesterol synthesis regulated? Explain.
- It is regulated at the HMG-CoA reductase reaction. 1.) AMP high (starved state): phosphorylation by AMP-dependent kinase, which inactivates enzyme and cholesterol synthesis is low during times of starvation 2.) Insulin high (well-fed state): desphosphorylation by protein phosphorylase activates enzyme, which means that cholesterol synthesis is high during times of well-fed 3.) Cholesterol inhibits the enzyme by feedback inhibition, which means that when cholesterol levels are high, cholesterol synthesis is low
113
How do FFAs travel in blood? Why is this transport method important?
- They bind to hydrophobic pockets in albumin. - Don’t need to synthesize FFA transporters when we are in a starved state and need FFAs as albumin is always present.
114
Name the ketone bodies. How are they used?
- Ketone bodies: acetoacetate, acetone (result of spontaneous decarboxylation of acetoacetate) and beta-hydroxybutyrate. - Ketone bodies have the ability to cross membranes and can leave liver and travel to peripheral tissue. - Beta-hydroxybutyrate gets oxidized to acetoacetate via beta-hydroxybutyrate dehydrogenase (yields NADH) - CoA from succinyl-CoA is transferred to acetoacetate yielding acetoacetyl-CoA via enzyme acetoacetate:succinyl-CoA transferase - Thiolase produces 2 x acetyl-CoA from acetoacetyl-CoA, which function in the TCA cycle to produce energy. - Acetone cannot be converted to acetyl-CoA
115
Explain enzymatic defect leading to the development of acute fatty liver of pregnancy.
- Fetus releases hydroxyacyl metabolites into the mother’s circulation as a result of long chain hydroxyacyl-CoA dehydrogenase (LCHAD) deficiency in the fetus. FAs accumulate in mother’s liver and they present with acute liver failure in pregnancy (jaundiced).
116
What is the committed step in TAG synthesis? What are TAG synthesis precursors used for?
- Committed step is conversion of phosphatidic acid to 1,2 diacylglycerol via phosphatidate phosphatase. - Phosphatidic acid can be shunted into lipid membrane synthesis pathways instead of going down TAG synthesis pathway.
117
What gluconeogenic deficiency(ies) seen in human? What is/are the clinical presentation(s)?
- G6PD deficiency: defect that lowers the activity of PPPathway and cells have lower NADPH levels. These levels impair glutathione reduction and deprive RBCs of antioxidant protection. Affected pts are very sensitive to hydrogen peroxide, which are produced in infections (by macrophages), by drugs (eg. Primaquine that is used to treat malaria and pneumocystis pneumonia) and during consumption of fava beans (known as favism) as these cause oxidatie stress and trigger hemolytic crisis and hemolytic anemia. Most prevalent in African and Mediterranean Americans. It is x-linked recessive.
118
What are the regulatory points in glycolysis? What factors are stimulating (+) and inhibiting (-)? What is the rate-limiting step in glycolysis?
- 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
119
What molecule does FA synthesis begin with? What are the second, third… molecules used?
- FA synthesis begins with acetyl-CoA and then joins with malonyl CoA during round 1 - Round 2-X begins with malonyl-CoA.
121
What is the role of glycogenin vs glycogen synthase?
- Glycogenin: acts as a primer protein and generates short glucose chains via autoglycosylation of glycogenin - Glycogen synthase: takes over from glycogenin and transfers glucose residues from UDP-glucose - Both form alpha-1,4-bonds
122
How do problems in gluconeogenesis cause fasting hypoglycemia and metabolic acidosis?
- Pyruvate can be oxidized to acetyl-CoA and moved into TCA or can be oxidized to lactate by lactate dehydrogenase. - When issues occur with gluconeogenic enzymes, much pyruvate can be oxidized to lactate as opposed to taking gluconeogenic pathways, leading to high levels of serum lactate and metabolic acidosis. - For example, when pyruvate carboxylase functions correctly, pyruvate is converted to oxaloacetate and is prevented from forming lactate and is now dedicate to moving through the gluconeogenesis pathway.
123
What is galactosemia?
- 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).
125
How can oxaloacetate initiate gluconeogenesis?
- Malate-oxaloacetate shuttle. Oxaloacetate can be moved from the mitochondria to the cytoplasm via this shuttle and gluconeogenesis can occur.
126
What is an anaplerotic reaction? What is the importance of these reactions for the TCA cycle? What is an important anaplerotic reaction for the TCA cycle? Include enzyme and coenzymes required. Are there other reactions?
- Anaplerotic reactions are those that replenish substrates. The TCA cycle is not an end-product producing reaction. It re-generates oxaloacetate in order for the cycle to occur again. However; TCA cycle intermediates, including oxaloacetate, are constantly being drawn away from the cycle as they serve as intermediates for biosynthetic pathways and therefore the oxaloacetate concentrations decrease preventing expanding rates of the TCA cycle should the need occur. - Pyruvate + ATP + HCO3 = oxaloacetate + ADP + Pi + H+ (ez: pyruvate carboxylase, coenzyme: biotin) - Yes, any reaction that generates an intermediate of the TCA cycle is a potential anaplerotic reaction. There is a reaction involving glutamate and glutamate dehydrogenase that generates more alpha-ketoglutarate, which adds to oxaloacetate in subsequent reactions.
127
Describe the synthesis of cholesterol from farnesyl PP.
128
What is the function of lipoxygenase inhibitors? What downstream molecule is being targeted with these inhibitors? What disorder/illness can these alleviate? How?
- These target synthesis of leukotrienes from AA via inhibition of the lipoxygenase enzyme. - Alleviates asthma by reducing smooth muscle contraction.
129
Types of apoproteins and functions?
- Apoprotein A: activates LCAT (lecithin:cholesterol acyltransferase), which generates cholesterol esters from cholesterols. Extracts lipids from membranes for reverse transport. - Apoprotein B: structural protein, interacts with lipoprotein receptors and mediates uptake of particle into target cells - Apoprotein C: modulate lipoprotein lipase (LPL) activity, which liberates FFAs and glycerol from lipoproteins - Apoprotein E: bind to receptors to allow removal of remnant particles from circulation
130
Under what conditions will lactic acid fermentation occur in the body? Why? What enzyme is involved? Why is this problematic in the body?
- 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.
