Chapter 18 Flashcards
Glycolysis vs. Citric Acid Cycle
Glycolysis:
- Glucose to pyruvate
- Occurs in cytoplasm
- Net yield of the reaction is 2-ATP
Citric Acid Cycle:
- Acetyl entry: acetyl unit enters as acetyl-CoA
- Occurs in mitochondrial matrix
- Generates high transfer potential, allows for generation of ATP
Acetyl CoA
- Under aerobic conditions, pyruvate enters the mitochondria through conversion into acetyl-CoA
- The two-carbon acetyl unit is processed into two molecules of CO2
- High energy electrons are generated to be used for formation of ATP
Notes:
- Acetyl-CoA = fuel for citric acid cycle
- Glycolytic pathway, we are going from glucose to pyruvate. CAC kicks off by converting the pyruvate into acetyl-CoA. Acetyl-CoA will go through oxidation in CAC with aim to generate ATP at the end
Pyruvate Dehydrogenase Complex
- Pyruvate dehydrogenase complex = mitochondrial matrix enzyme that oxidatively decarboxylates pyruvate to form acetyl-CoA
- Reaction is an irreversible link between glycolysis and citric acid cycle
- Irreversible enzyme complex has 3 distinct enzymatic activities (reactions):
- Oxidative decarboxylation
- Transacetylation
- Re-oxidation
- The 3 irreversible enzymatic reactions are performed by three distinct regions of the enzyme: E1, E2, E3
- Each reaction is further facilitated by a specific prosthetic group:
–> TPP (thiamine pyrophosphate) in E1
–> Lipomide in E2
–> FAD (flavin adenine dinucleotide) in E3
SUMMARY:
Enzyme pyruvate dehydrogeanse –> TPP (Region E1) –> oxidative decarboxylation of pyruvate
Enzyme dihydrolipoyl transacetylase –> Lipomide (Region E2) –> transacetylation; transfer of acetyl group to CoA
Enzyme dihydrolipoyl dehydrogenase –> FAD (Region E3) –> re-oxidation; regeneration of oxidized form of lipoamide
Overview of Pyruvate Dehydrogenase Reactions
Reaction involves: 3-enzymes, 4-main reactions, 5-coenzymes
4-main reactions involve the following steps:
1. Decarboxylation
2. Oxidation
3. Transfer to CoA
4. Re-oxidation (to reset enzyme)
5 coenzymes include:
- Thiamine pyrophosphate (TPP)
- Lipoic acid
- FAD
- CoA
- NAD+
1.Decarboxylation: E1
E1-component of pyruvate dehydrogenase catalyzes decarboxylation
Pyruvate –> combines w/ ionized form of thiamine pyrophosphate (TPP) –> is decarboxylated –> hydroxyethyl-TPP generated
- TPP is derived from vitamin thiamine (vitamin-B1)
- TPP = coenzyme (1st one)
2.Oxidation: E1
Hydroxyacetyl-TPP –> E1 component oxidizes hydroxyexthyl group that’s attached to TPP into acetyl –> while this is happening, it’s being transferred to lipoamide of E2 (dihydrolipoyl transacetylase component)
- Reaction is catalyzed by E1 and yields acetyl-lipoamide
- Disulfide group of lipoamide is reduced to its disulfhydryl form in reaction
- Lipoamide = derivative of lipoic acid
- Lipoic acid = coenzyme (2nd one)
3.Transfer to CoA: E2
Acetyl-ipomide –> E2 transfers acetyl group from acetyl-lipomide to coenzyme A (CoA) –> left with acetyl-CoA + dihydrolipomide
- Coenzyme A = coenzyme (3rd one)
4.Reoxidation: E3
dihydrolipomide –> E3 oxidizes dihydrolipomide to lipomide w/ help from FAD and NAD+
- To participate in another reaction cycle, dihydrolipoamide must be re-oxidized
- Dihydrolipoamide is not able to accept acetyl group in step 2 (oxidation) unless its re-set
- Needs to “re-oxidize” back to lipoamide to restore enzyme functionality
Structure/Function of Pyruvate Dehydrogenase
- The 3 enzymes of pyruvate dehydrogenase complex are structurally integrated
- Lipoamide arm allows rapid movement of substrates and products from one active site of complex to another
- Subunit of transacetylase consists of 3 domains:
- lipoamide-binding domain
- Domain for interaction w/ E3
- Large transacetylase catalytic domain
Summary of steps:
- Pyruvate is decarboxylated to form the hydroxyethyl-thiamine pyrophosphate (TPP) intermediate
–> First product, CO2 exits
–> Active site of E1 complex is deep within hydrophobic channel - Lipoamide arm of E2 moves into active site of E1
- E1 catalyzes transfer of 2-carbon acetyl group to lipoamide group to form acetyl–lipoamide complex through oxidation of hydroxyethyl group attached to TPP
- E2 catalyzes transfer of acetyl moiety to CoA to form product acetyl CoA
–> Dihydrolipoamide arm then swings to active site of E3 - E3 catalyzes oxidation of dihydrolipoamide acid and transfer of protons and electrons to NAD+ to complete reaction cycle
Fate of Acetyl CoA
- Formation of acetyl-CoA from pyruvate dehydrogenase complex = key irreversible step in metabolism of glucose in mammalian cells
- After, acetyl-CoA has two principal fates:
- Metabolism by CAC
- Incorporation into fatty acids
Regulation of pyruvate dehydrogenase
Covalent modification:
- Regulation of pyruvate dehydrogenase complex occurs at key E1 site of regulation
- A kinase associated with the complex phosphorylates and inactivates E1.
- A phosphatase, also associated with the complex, removes the phosphate and thereby activates the enzyme
Regulation by energy charge:
- ATP, acetyl CoA, and NADH inhibit the complex
- ADP and pyruvate stimulate the complex
- Allosteric regulation occurs
–> Directly by acetyl CoA and NADH (products of PDH)
–> Indirectly through PDH kinase and PDH phosphatase
NOTES:
- high energy in cell = inhibit pyruvate dehydrogenase complex
- low energy in cell = activate pyruvate dehydrogenase complex
- high levels of ATP, Acetyl-CoA, and/or NADH within cell = pyruvate dehydrogenase complex inhibited through activation of kinase
- high levels of ADP and pyruvate = stimulate activation of dehydrogenase complex through inhibition of kinase
- Phosphatase activity will also be activated by calcium levels, which is important in muscles where intracellular calcium will be high for actively contracting muscle cells
- Intracellular calcium also associated w/ epinephrine signaling in liver
- Insulin can also activate the activity. Allosteric regulation occurs directly by acetyl CoA and NADH (products of PDH) and indirectly through PDH kinase and PDH phosphatase
Vitamin B1 deficiency
- Beriberi, a neurological and cardiovascular disorder, is caused by a dietary deficiency of thiamine
- Thiamine deficiency results in insufficient pyruvate dehydrogenase activity because thiamine pyrophosphate (TPP) cannot be formed
