Unit 19: TCA/Citric acid/ Krebs cycle; PDH rxn; Beta oxidation Flashcards
describe mitochondria generally
- where energy conversion happens
- Hundreds to thousands per cell , depending
on cell type
– Many fused to one another
– Some cells do not have mitochondria - has a Double membrane
– Outer membrane controls entry of
large molecules
– Electron transport chain (ETC) and ATP synthesis enzymes located in inner membrane - need over 1000 different proteins for maintenance and function
- most are encoded by nuclear genome and imported. some are encoded by mtDNA
describe mitochondria inner membrane
- High content of cardiolipin and very protein-rich
- Impermeable to ions and small charged molecules
- Contains electron transport chain (ETC) and ATP Synthase
- Contains protein importers (e.g., TIM), transporters and shuttles for small molecules and ions
describe mitochondria Outer membrane
- Molecular sieve with channels formed by porins
- Permeable for molecules < 5 kDa (in open state)
- Enzymes for mitochondrial lipid synthesis located here
- Import receptors and translocases for proteins (TOM complex) here
describe mitochondria Intermembrane space
- Similar to cytoplasm in composition (small molecules)
- Various enzymes exist here
- the space between the outer membrane and inner membrane
describe mitochondria Matrix
Contains enzymes for several pathways
- pyruvate oxidation, TCA, amino acids, fatty acids, etc.
Contains mtDNA, transcription and translation components
- inner most compartment
the TCA is the complete oxidation of fuel carbon backbone to CO2 with ____
electrons bound to carriers
Acetyl CoA (C2) is also know as
activated acetate
what is the first molecule to enter the TCA cycle
acetyl CoA
what are the catabolic and anabolic functions of TCA cycle
Catabolic:
- Provides electrons for oxidative phosphorylation
(3 NADH and 1 FADH2 per full cycle)
- Provides energy via substrate- level phosphorylation (1 step, 1 GTP)
anabolic:
- Provides precursors for building blocks
is the TCA cycle Amphibolic?
yes, has both catabolic and amphibolic pathways
what coupled cycles provide most of the energy from glucose oxidation
the TCA cycle and oxidative phosphorylation (ETC + ATP synthase)
do Fatty acids, glucose and Amino acids all feed into the TCA cycle?
yes, via acetyl coA
describe the PDH reaction
- pyruvate transported into mitochondria matrix via mitochondrial pyruvate carrier (MPC)
- Oxidative decarboxylation to acetyl CoA by PDH enzyme complex in the matrix
- irreversible rxn
what is the net reaction of the PDH rxn
pyruvate + CoA + NAD = Acetyl CoA + CO2 + NADH + H+
what is the ∆𝐺′° of the PDH rxn
-8.0 kcal/mol
what is the PDH complex made of
E1: Pyruvate dehydrogenase (TPP)
E2: Dihydrolipoyl transacetylase (Lipoamide) + CoA
E3: Dihydrolipoyl dehydrogenase (FAD) + NAD+
what are the 5 coenzymes of the complex
TPP,
Lipoamide + CoA,
FAD + NAD+
what are the co-substrates of the PDH rxn?
CoA and NAD+
- they are not tightly bound. they pick up molecules and leave
what are the prostethic groups of the PDH rxn
TPP (Pyruvate dehydrogenase) (E1)
Lipoamide (Dihydrolipoyl transacetylase) (E2)
FAD - dihydrolipoyl dehydrogenase (E3)
what are the apoenzymes of the PDH rxn
Pyruvate dehydrogenase
Dihydrolipoyl transacetylase
Dihydrolipoyl dehydrogenase
what does Pyruvate dehydrogenase do
takes away the CO2 and electrons
what does Dihydrolipoyl transacetylase do
something with the acetyl
what does Dihydrolipoyl dehydrogenase do
take the electrons away
describe general mechanism of PDH rxn
pyruvate decarboxylase
1. decarboxylation of Pyruvate
2. oxidation of acetyl group w/ simultaneous transfer to lkpoamide + formation of thioester bond
Dihydrolipoyl transacetylase
3. transfer of acetyl group from lipoamide to CoA + Preserves thioester bond
Dihydrolipoyl dehydrogenase
4. regeneration of lipoamide by oxidation of dihydrolipoamide
– Prosthetic FAD group as primary electron acceptor
– Electrons then transferred to NAD+
what is the advantage of multi enzyme complex in performing consecutive reactions
Facilitates ‘substrate channeling’
Maximizes efficiency
Minimizes side reactions
causes of PDH deficiency
- Vitamin deficiencies (thiamine, niacin) due to nutritional deficits, alcohol abuse
– Mutations of E1 component
– Arsenic poisoning (inhibits lipoamide-containing enzymes)
consequences of PDH deficiency
- Lactic acidosis due to conversion of pyruvate to lactic acid
- Severe CNS symptoms due to insufficient levels of ATP in CNS (relies on TCA cycle for energy production)
- especially in mitochondria dense cells
sources for fatty acids
triglycerides (adipose tissue)
dietary fats
acetyl CoA
storage form of fatty acids
triglycerides-non polar storage in lipid droplets (adipocytes)
enzyme that liberates FA
lipase- hydrolyzes the ester bond between oxygen and carboxyl group of hydrocarbon tail
how are fatty acids activated
by binding to CoA
triglycerides are __ for transport
solubilized
- bile salts for dietary lipids
- lipoprotein particles for blood transport of triglycerides
- albumin for FA
how are FA activated
-bind to CoA
- enzyme is acetyl CoA synthase
- a mitochondrial outer membrane protein
why is FA oxidation is called beta oxidation
b/c the beta carbon undergoes oxidation to a carbonyl group
what is the fate of Glycerol from TG cleavage?
glycolysis - G3P
FA oxidation in mitochondria vs. peroxisome
mitochondria:
- FA less than 20 carbons
- activation in cytoplasm and transported into mt
- b-oxidation to acetyl CoA
- complete oxidation to CO2 by TCA
- ATP regeneration by oxphos
peroxisome:
- for very long FA and branched
- activation in cytoplasm and transported into peroxisomes
- b-oxi to short FA and acetyl CoA
- direct reduction of O2 to H2O
- detox by Catalase
- export of NADH and Acetyl CoA; no ATP
how are FA shuttled into one of the cellular compartments where oxidation takes place
via the carnitine shuttle
- it’s inhibited by malonyl-CoA
- a product of acetyl-Coa carboxylation at high levels of acetyl CoA
net rxn of Mt FA Oxidation
Palmitate + 8 CoA + 1 ATP + 7 NAD + 7 FAD + 7 H2O = 8 acetyl Coa + 1 AMP + 2 Pi + 7 NADH + H + 7 FADH2
need 8 CoA, happens 7 times
where does the acetyl CoA go and the NADH + FADH2 go
Acetyl CoA goes to TCA
NADH, FADH2 goes to OxPhos
describe ketone bodies
produced by liver from acetyl CoA
– Alternate pathway for acetyl CoA metabolism
– Secreted into circulation
Fuel source for skeletal and
heart muscle, kidney
– Acetoacetate, hydroxybutyrate can be used by brain under starvation conditions
Produced in excess in diabetes, leading to acidosis (ketosis)
function of the TCA cycle
produce NADH, FADH2
is the TCA anaerobic or aerobic
TCA is aerobic but operates under anaerobic conditions
- O2 is required to progress for regenerating NAD/FAD via ETC
What is the acceptor molecule for acetyl-CoA
oxaloacetate
describe 1st rxn of TCA
-condensation rxn of acetyl CoA w/ oxaloacetate
-enzyme citrate synthase
-produces Citrate
- irreversible
- highly exergonic through hydrolysis energy-rich intermediate
what pathway and enzymes are involved in replenishing oxaloacetate
gluconeogenesis
Pyruvate carboxylase and PEP carboxylkinase
-pyruvate carboxylase is activated by acetyl CoA
replenishing rxns are called
anaplerotic
irreversible steps of the TCA
step 1: enzyme citrate synthase
Step 4: isocitrate to a-ketoglutarate; enzyme isocitrate dehydrogenase
step 5: a-ketoglutarate to succinyl CoA; enzyme a-ketoglutarate dehydrogenase complex
full cycle of TCA generates:
10 ATP/full cycle
– 1 GTP in TCA cycle via substrate level phosphorylation
– ~2.5 ATP/NADH+H+ via oxidative phosphorylation
– ~1.5 ATP/FADH2 via oxidative phosphorylation
TCA net rxn
Acetyl CoA + 3 NAD + FAD + GDP + Pi + 2 H2O = 2 CO2 + CoA + 3 NADH + H + FADH2 + GTP
controls of PDH
Inhibit:
- high levels of ATP, Acetyl CoA and NADH
Activate:
- high levels of AMP, CoA, Pyruvate
reversible modification (phosphorylation, dephosphorylation)
controls E1. phosphorylation inactivates
controls of TCA
Isocitrate DH
– Inhibited by NADH
– Activated by NAD+
– Inhibited by ATP
– Stimulated by ADP
- Activated by Ca2+ (muscle)
α-Ketoglutarate DH
– Inhibited by HIGH succinyl CoA and NADH
– Activated by NAD+
– Inhibited by ATP
- Activated by Ca2+ (muscle)
Citrate Synthase
– Inhibited by citrate, succinyl CoA
(prokaryotic, mammalian in vitro)
Prokaryotes: – Inhibited by ATP and Stimulated by ADP
pathway integration
Low energy charge
– Glycolysis, PDH and TCA becomes active
– Oxaloacetate reacts with Acetyl-CoA to citrate
High energy charge
– Glycolysis, PDH and TCA
becomes less active
– Conversion of oxaloacetate to glucose (gluconeogenesis)
general steps for Beta oxidation of saturated FA and enzymes
- Dehydrogenation (oxidation
- Acyl-CoA-Dehydrogenase - Hydration
- Enoyl CoA hydratase - Dehydrogenation (oxidation)
- L-3 hydroxyacyl CoA DH - Cleavage (thiolysis)
- β-ketothiolase
which enzyme of the TCA is also a part of the ETC
Succinate dehydrogenase (complex II)
which TCA intermediate is regenerated through anaplerotic rxn
Oxaloacetate
what TCA rxns synthesize NADH, FADH2 and GTP?
Step: NADH
what TCA rxn are substrate level phosphorylation
step 6: Succinyl CoA to Succinate; enzyme Succinyl CoA synthetase
what TCA rxns are oxidative decarboxylation
step 4: isocitrate to a-ketoglutarate via isocitrate DH
step 5: a-ketoglutarate to succinyl CoA; enzyme a-ketoglutarate DH complex
molecules synthesized for TCA intermediates
- FA, Sterols
- amino acids
- porphyrins, heme
