Glycolysis and PDH Flashcards
1
Q
Glycolysis
A
- splitting of 1 6-carbon glucose into 2 3-carbon pyruvate molecules
- NAD+ is reduced to NADH
2
Q
Aerobic glycolysis
A
- when O2 supply is plentiful, NADH reoxidised to NAD+ via mitochondria
- pyruvate taken up by mitochondria, metabolised to CO2 and H2O via TCA cycle
3
Q
Anaerobic glycolysis
A
- NADH production exceeds the capacity of the electron transport chain
- Pyruvate is converted to lactate by lactate dehydrogenase
- Converts NADH back into NAD+, lowering the level of NADH
- Reduces intracellular pH
- Lactate diffuses out into bloodstream & is processed by the liver to produce glucose
4
Q
Significance of glycolysis
A
- only pathway that takes place in all cells of the body
- only source of energy in erythrocytes and only fuel normally used by neurones
- provides carbon skeletons for synthesis of non-essential amino acids as well as glycerol part of fat
- Most reactions of glycolytic pathway are reversible, which are used for gluconeogenesis
5
Q
Glycolysis in exercise
A
- Rapid formation of energy during short-term strenuous exercise: glycolysis could support activity up to c. 2 minutes
- Cardiac muscle is adapted for aerobic performance: doesn’t fatigue like skeletal muscle, has low glycolytic activity, rapidly damaged under ischaemic conditions
6
Q
Site of glycolysis
A
- occurs in the cytosol of all the cells of the body
7
Q
Two phases of glycolysis
A
- energy investment (reactions 1-5)
- energy generation (reactions 6-10)
8
Q
Energy investment (reactions 1-5)
A
- sugar phosphates are synthesised at the expense of ATP -> ADP
- the sugar is metabolically activated by phosphorylation
- 6C split into 2x3C sugar phosphates (triose phosphates)
9
Q
Energy generation (reactions 6-10)
A
- further activation of triose phosphates to energy-rich compounds
- reduced electron carriers are generated (NADH)
- the energy-rich compounds then transfer phosphate to ADP to form ATP = substrate-level phosphorylation
10
Q
Substrate-level phosphorylation
A
- generation of an energy-rich phosphate bond driven by the breakdown of a more energy-rich substrate
11
Q
Hexokinase reaction: phosphorylation of hexoses (mainly glucose)
A
- Hexokinase is present in most cells.
- In liver, Glucokinase is the main hexokinase (both ISOENZYMES) which prefers glucose as substrate
- It requires Mg-ATP complex as a substrate. Uncomplexed ATP - potent competitive inhibitor of this enzyme.
- Enzyme catalyses the reaction by bringing the two substrate in close proximity.
- This enzyme undergoes large conformational change upon binding with glucose. It is inhibited allosterically by G6P.
12
Q
Hexokinase
A
- high affinity for glucose
- Non-specific, can phosphorylate any of hexoses
- Present in tissues, supplies glucose to tissues even in low blood glucose concentration
- Not effected by insulin
- Allosterically inhibited by glucose
13
Q
Glucokinase
A
- low affinity for glucose
- Specific, can phosphorylate only glucose
- Present in liver only
- Helps drive movement of glucose from blood to cells after meal
- Stimulated by glucose and insulin
- Not inhibited by glucose-6-phosphate
14
Q
Phosphohexose Isomerase: Isomerization of Glucose-6-Phosphate (G6P) to Fructose-6-phosphate (F6P)
A
- catalyses the reversible isomerization of G6P - (an aldohexose) to F6P - (a ketohexose)
- requires Mg++ for its activity.
- specific for G6P and F6P
- extracellular PGI have multiple additional roles in health and disease: neural growth factor, driver of cancer cell metastasis and maturation
15
Q
Phosphofructokinase-1 (PKF-1) Reaction: Transfer of phosphoryl group from ATP to C-1 of F6P to produce Fructose 1,6 bisphosphate
A
- important irreversible, regulatory step.
- PFK-1 is one of the most complex regulatory enzymes, with various allosteric inhibitors and activators.
- ATP is an allosteric inhibitor, and Fructose 2,6 biphosphate is an activator of this enzyme.
- ADP and AMP also activate PFK-1 whereas citrate is an inhibitor.
- PFK-1 has 3 different sub-units whose distribution may be tissue-specific
16
Q
Aldolase Reaction: Cleavage of Fructose 1,6 bisphosphate into glyceraldehyde 3 phosphate and dihydroxy acetone phosphate.
A
- Conversion of aldose to ketose
- catalyses the cleavage of F1,6 biphosphate by aldol condensation mechanism.
- standard free energy change is positive in the forward direction, meaning it requires energy.
- Driven forwards due to subsequent reactions
- Ensures rapid aldolase-driven conversion
- “moonlighting” roles e.g. cell structure, endocytosis, cancer cell survival and protein mediated transport
17
Q
Triose phosphate isomerase reaction: Conversion of Dihydroxyacetone phosphate to glyceraldehyde 3 Phosphate
A
- reversible reaction catalysed by acid-base catalysis
- Histidine-95 + Glutamate-165 residues of the enzyme are involved
- Triosephosphate Isomerase Deficiency: number of rare metabolic disorders linked to glycolysis enzymes, chronic haemolytic anaemia
18
Q
Glyceraldehyde-3-phosphate dehydrogenase reaction (GAPDH): Conversion of GAP to Bisphosphoglycerate
A
- Oxidation of aldehyde derives the formation of a high energy acyl phosphate derivative.
- Pi is incorporated in this reaction without any expense of ATP.
- NAD+: cofactor in this reaction which acts as an oxidizing agent. The free energy released in the oxidation reaction is used in acyl phosphate formation
- Appears strongly linked to control of cell survival or apoptosis in response to multiple factors e.g. oxidative stress, starvation and toxic compounds
19
Q
Phosphoglycerate kinase Reaction: Transfer of phosphoryl group from 1,3 bisphosphoglycerate to ADP generating ATP
A
- catalyses the formation by proximity effect. ADP-Mg bind on one domain and 1,3BPG binds on the other and a conformational change brings them together similar to hexokinase.
- coupled reaction generating ATP from the energy released by oxidation of 3-phosphoglyceraldehyde
- generates ATP by substrate-linked phosphorylation
- Linked to roles in interaction with DNA/RNA, cell death and viral replication
20
Q
Phosphoglycerate Mutase Reaction: Conversion of 3-phosphoglycerate to 2-phosphoglycerate (2-PG)
A
- transfer of the phosphoryl group form enzyme to 3-PG, generating enzyme bound 2,3-biphosphoglycerate (2,3BPG) intermediate
- phosphoryl group from the C-3 of the intermediate is transferred to the enzyme and 2-PG is released.
- traces of 2,3BPG present in most cells, but in erythrocytes, it is present in significant amount, affects oxygen affinity to Hb
21
Q
Enolase Reaction: Dehydration of 2-phosphoglycerate (2-PG) to phosphoenolpyruvate (PEP)
A
- increases the standard free energy change of hydrolysis of phosphoanhydride bond
- rapid extraction of proton from C-2 position by general base on enzyme, generating a carbanion. The abstracted proton is readily exchanges with solvent.
- second rate limiting step involves elimination of -OH group generating PEP
22
Q
Pyruvate Kinase Reaction: Transfer of phosphoryl group from PEP to ADP generating ATP and Pyruvate
A
- second substrate level phosphorylation reaction of glycolysis
- couple the free energy of PEP hydrolysis to synthesis of ATP
- requires Mg++ and K+
- also linked to roles in cell regulation
23
Q
Products of glycolysis
A
- 1Gluc + 2NAD+ + 2ADP + 2Pi = 2pyruvate + 2ATP + 2NADH
- 2ATP generated can directly be used for doing work or synthesis
24
Q
Products of glycolysis in aerobic conditions
A
- 2 NADH are oxidized in mitochondria
- free energy released is enough to synthesize 6 molecules of ATP by oxidative phosphorylation
- pyruvate is catabolized further in mitochondria through PDH and citric acid cycle where all the carbon atoms are oxidized to CO2
- free energy released is used in the synthesis of ATP, NADH and FADH2
25
Q
Effects of hormones in glycolysis
A
- Insulin stimulate Hexokinase & Glucokinase by converting glucose to glu-6-PO4
- Insulin stimulate Phosphofructokinase converting fru-6-PO4 to Fru-1,6 bisphosphate
- Glucagon stimulate liver glu-6-PO4 by converting glu-6-PO4 to glucose & fru-1,6- bisphosphate.
- Fru-1,6- bisphosphate is converted to fru-6-PO4
26
Q
Inhibitors
A
- Iodoacetate inhibit Gly-3-PO4 dehydrogenase involved in gly-3-PO4 to 1,3-bisphosphoglycerate
- Arsenate inhibit synthesis of ATP in the conversion of 1,3 bisphosphoglycerate to 3-phosphoglycerate.
- Fluoride inhibit enolase in conversion of 2-Phosphoglycerate to phosphoglycerate
27
Q
Substrate limited regulation of glycolysis
A
- When concentrations of reactant and products in the cell are near equilibrium, substrate availability drive reaction rate
28
Q
Enzyme limited regulation of glycolysis
A
- When concentration of substrate and products are far away from the equilibrium, then it is activity of enzyme that decides rate of reaction
29
Q
3 steps in glycolysis that have enzymes which regulate the flux of glycolysis
A
- hexokinase (HK)
- phosphofructokinase (PFK)
- pyruvate kinase
30
Q
Oxidation of pyruvate
A
- the generation of an activated 2C fragment = the acetyl group of acetyl CoA
31
Q
Conversion of pyruvate to acetyl-CoA
A
- Pyruvate enters the mitochondrial matrix
- Undergoes oxidative decarboxylation
- Pyruvate + NAD+ + CoA –>
acetyl CoA + NADH + CO2 - Sequence of reactions catalysed by pyruvate dehydrogenase complex (3 enzymes + 5 coenzymes)
- A virtually irreversible reaction
32
Q
What 3 principal enzymes does pyruvate dehydrogenase complex contain?
A
- E1: pyruvate dehydrogenase
- E2: dihydrolipoamide transacetylase
- E3: dihydrolipoamide dehydrogenase
33
Q
Coenzymes in PDH complex
A
- thiamine pyrophosphate
- lipoic acid
- coenzyme A
- flavin adenine dinucleotide
- nicotinamide adenine dinucleotide
34
Q
Diet and PDH complex activity
A
- maybe a rapid potential to affect PDC activity in muscle (and other tissues) by changing dietary energy sources
- High fat diet lower activity vs high CHO diet in <5 days (“favours” fat oxidation)
- May support CHO sparing during prolonged physical activity
- Higher fat diets not endorsed by sports nutrition expert bodies
35
Q
Where does acetyl-CoA enter?
A
- citric acid cycle, which has a central role in cell metabolism, to oxidise organic metabolites
