Mitochondria, Pyruvate Dehydrogenase Complex, TCA Cycle

overall purpose of pyruvate dehdyrogenase complex (PDH) and tricarboxylic acid cycle (TCA pathway)

to extract maximum energy from glucose by converting it into:

- CO₂ (fully oxidized carbon)

- NADH/FADH₂ (energy carriers)

- small amount of ATP/GTP

3 names for the TCA cycle

- tricarboxylic acid (TCA) cycle

- citric acid cycle

- krebs cycle

why tricarboxylic

because intermediates contain 3 carbons (-COOH) groups

what is the overall equation for glucose metabolism

glucose + O₂ → CO₂ + H₂O

where does energy go

stored in ATP

stored in NADH/FADH₂

why is NADH considered "high energy"

contains high energy e-

these e- will later drive ATP production

4 parts of mitochondria

outer membrane

inner membrane

intermembrane space

matrix

outer membrane

directly adjacent to cytosol

permeable to metabolites (proteins & small molecules)

fat metabolism enzymes (acyl-coA synthetase, glyceral phosphate acytransferase)

porous

inner membrane

directly adjacent to matrix

NOT permeable, highly selective

many transporters (ex. pyruvate, malate, asp)

electron carriers in resp. chain

enzymes (ATP synthase)

what is special about the inner membrane

highly selective

contains:

- ETC

- ATP synthase

folded into cristae → ↑ SA

what processes occur in the mitochondrial matrix

TCA cycle

pyruvate dehydrogenase

beta- oxidation:

- fatty acids → acetyl CoA

name key mitochondrial diseases

- MERRF (Myoclonic Epilepsy associated with Ragged Red Fiber)

- MELAS ( Mitocohondrial Encephalopathy with Lactic Acidosis and Stroke-like episodes)

- LHON (Leber Hereditary Optic Neuropathy)

- GRACILE ( Growth Retardation, Aminoaciduria, Cholestasis, Iron overload, Lactic acidosis, Early death)

-Leigh syndrome

- CODAS (Cerebral, Ocular, Dental, Auricular, and Skeletal Anomalies Syndrome)

common symptoms of mitochondrial diseases

lactic acidosis due to reliance on glycolysis

neurological issues

muscle weakness/fatigue

what other conditions are thought to involve mitochondrial dysfunction

- Parkinsons

- alzheimer

- cancer

- aging

why is lactic acidosis a common symptom in mitochondrial diseases

without mitochondrial function, pyruvate must be directed to lactate to regenerate NAD+

what happens if mitochondria don't work

-only glycolysis runs

- pyruvate → lactate

- needed to regenerate NAD+

what are two major fates of pyruvate

- acetyl-CoA (via pyruvate dehydrogenase PDH) → TCA & fatty acid synthesis

- oxaloacetate via pyruvate carboxylase → gluconeogeneiss & TCA

pyruvate dehydrogenase complex (PDH)

- irreversible

- acetyl CoA feedback inhibits

- provides substrate for TCA and fatty acid synthesis

pyruvate carboxylase

- irreversible

- acetyl CoA activates

- provides substrates for gluconeogenesis (GNG)

what does pyruvate dehydrogenase (PDH) do

converts pyruvate (3 C) → acetyl-CoA (2C)

releases:

- CO₂

- NADH

is PDH reversible

no

links glycolysis → TCA

major regulatory step

cofactors required for PDH

1. thiamine pyrophosphate (B1)

2. lipoate

3. CoA

4. FAD

5. NAD+

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pyruvate decarboxylase E1 cofactor

thiamine pyrophosphate (vitamin B1)

dihydrolipoyl transacetylase E2 cofactor

lipoate, coenzyme A

dihydrolipoyl dehydrogenase E3 cofactor

FAD, NAD+

what are the 3 enzyme components of PDH

E1 : pyruvate dehydrogenase (Decarboxylase)

E2: dihydrolipoyl transacetylase

E3: dihydrolipoyl dehydrogenase

PDH complex structural points

- 3 diff. enzymes

- multienzyme ( >100 subunits) complex: reaction intermediates pass from enzyme to enzyme... assembly line

- essential cofactors participate in catalytic reactions

PDH mechanism summary

1. CO₂ removed

2. Acetyl transferred to CoA

3. NADH produced

PDH mechanism- step 1

CO₂ removed

pyruvate decarboxylase + cofactor thiamine pyrophosphate (E1) removes carbon

PDH mechanism- step 2

acetyl transferred to CoA

dihydrolipoyl transacetylase + cofactor lipoic acid (E2) transfers acetyl

PDH mechanism- step 3

NADH produced

dihydrolipoyl dehydrogenase (E3) regenerates oxidized lipoic acid and FAD

pyruvate fate if TCA is not required (ex. gluconeogenesis active)

high [acetyl CoA] activates pyruvate carboxylase

blocks pyruvate → TCA

pyruvate is diverted away from oxidation and instead carboxylated into intermediates that support glucose synthesis:

- conversion via pyruvate carboxylase into oxaloacetate (OAA) within the mitochondria

pyruvate fate if TCA is required (ex. for fat synthesis)

low [acetyl CoA] activates PDH complex in matrix

CoA → acetyl CoA → helps synthesize fatty acids

what happens to PDH regulation when energy is high/cell has adequate energy

adequate energy:

↑ ATP

↑ NADH

↑ acetyl-CoA

activates PDH kinase → phosphorylates PDH → deactivates PDH

what does pyruvate dehydrogenase kinase (PDK) do

phosphorylates pyruvate dehydrogenase/decarboxylase

if kinase is active → acetyl CoA production ↓

______ turns on pyruvate decarboxylase (E1)

dephosphorylation

what happens when energy is low in cell

low energy:

- low ATP/ADP

- low acetyl CoA/CoA

- low NADH/NAD+

↑ pyruvate, ↑ ADP

activates PDH phosphatase → dephosphorylates → pryuvate dehyrogenase (E1) on

pyruvate dehydrogenase phosphatase (PDP)

activates pyruvate decarboxylase/ dehydrogenase (E1) in low energy

if kinase off and PPase on → acetyl CoA production increased

what does insulin do to PDH

activates pyruvate dehydrogenase phosphatase (PDP)

turns PDH on → promotes energy storage

**activates the pyruvate dehydrogenase (PDH) complex by removing inhibitory phosphate groups

PDH summary

- irreversible

- large multi- enzyme complex w cofactors

- rate limiting/ key regulatory step

- feedback inhibited by NADH & acetyl- CoA

- regulated by phosphorylation

- activated by insulin via dephosphorylation

CoA → Acetyl-CoA

NAD+ → NADH + H⁺

pyruvate → CO₂

pyruvate carboxylase vs PDHC

pryuvate carboxylase:

- pyruvate + CO₂ + ATP → oxaloacetate + ADP + Pi

- activated by high energy state

pyruvate dehydrogenase complex:

- pyruvate + CoA + NAD → acetyl-CoA + CO₂ + NADH

- inhibited by ATP, acetyl-CoA, and NADH (high energy)

other names for tricarboxylic acid (TCA) cycle

- citric acid

- krebs

what is TCA

metabolic pathway in aerobic respiration that oxidizes acetyl-CoA to produce CO₂, ATP/GTP, and reduced cofactors (NADH, FADH₂)

in the mitochondrial matrix of eukaryotes

fueling the ETC to generate the majority of cellular energy

what enters the TCA

acetyl-CoA (2 C)

combines with oxaloacetate (4C) → citrate (6C)

what happens to carbons in TCA

2 carbons enter

2 carbons leave as CO₂

energy yield per acetyl-CoA

3 NADH

1 FADH₂

1 GTP (by substrate level phosphorylation... equivalent to ATP)

what are the 3 irreversible enzymes

1. citrate synthase

2. isocitrate dehydrogenase

3. α-ketoglutarate dehydrogenase

what inhibits TCA

isocitrate dehydrogenase:

- ATP

- NADH

α-ketoglutarate dehydrogenase:

- succinyl-CoA

- NADH

what activates TCA

isocitrate dehydrogenase:

- ADP

- Ca²⁺

α-ketoglutarate dehydrogenase:

- Ca²⁺

if there is high energy (ATP and reducing power NADH), turn ____ citric acid cycle

off

**prevents unnecessary production of energy when the cell already has adequate supply**

enzymes that transfer potential to energy carriers

- isocitrate dehydrogenase

- α-ketoglutarate dehydrogenase:

- succinyl CoA synthetase

- succinate dehydrogenase

- malate dehydrogenase

how is α-ketoglutarate dehydrogenase similar to PDH

same cofactors:

- thiamine pyrophosphate (vit B1)

- lipoate

- coenzyme A

- FAD

- NAD+

produces NADH

releases CO₂

irreversible

rate limiting

key differences between α-ketoglutarate dehydrogenase and PDH

α:

- not regulated by phosphorylation

- regulated by Ca²⁺ (secondary messenger)

α-ketoglutarate dehydrogenase complex

1. irreversible

2. large multi enzyme complex with 5 cofactors

3. rate limiting

4. feedback inhibited by succinyl CoA and NADH

5. NOT regulated by phosphorylation

6. regulated by Ca2+

CoA → succinyl CoA

NAD+ → NADH + H⁺

α-ketoglutarate (more carbons) → CO₂

location of TCA

eukaryotes = matrix of mitochondria

prokaryotes = cytoplasm

memory aids for intermediates

Citrate Is Krebs Starting Substrate For Making Ocaloacetate

Oh! Can I Keep Some Succinate For Myself

visual of irreversible steps

- pryuvate dehydrogenase (PDH) complex

- citrate synthase

- isocitrate dehydrogenase

- a-ketoglutarate dehydrogenase

enzymes

carrier molecules in TCA

acetyl-CoA

NADH

FADH₂

GTP

each turn of cycle → 3 NADH and 1 FADH₂

GTP = produced directly through substrate-level phosphorylation when succinyl-CoA is converted to succinate

total yield from 1 glucose

glycolysis (cytosol: 2 pyruvate + 2 ATP + 2 NADPH

pryuvate to acetyl-CoA in mitochondria (PDH): 2 acetyl-CoA +2 NADH

TCA (mitochondria): 6 NADH + 2 FADH₂ + 2 GTP

glycolysis and the TCA cycle produce energy ____ and ____ _____

carriers, building blocks

what is anaplerosis

replenishing TCA intermediates that have been extracted for biosynthesis

retain homeostasis of cellular metabolism

what are anaplerotic reactions

chemical reactions that form intermediates of a metabolic pathway

ex. pyruvate → oxaloacetate via pyruvate carboxylase

why are shuttles needed

NADH cannot cross inner membrane

malate/aspartate:

- reduced NADH → oxidized NAD+

-reduced NADH ← oxidized NAD+

shuttles transfer ____ equivalents from cytosol to mitochondria

reducing

two main shuttles

malate-aspartate shuttle:

- electrons from cytosolic NADH to oxaloacetate, forming malate, which crosses into the mitochondrial matrix, is re-oxidized back to oxaloacetate, and generates mitochondrial NADH

- high energy tissues (heart, liver)

- produces NADH

glycerol-3-phosphate shuttle

- muscle

- FADH₂

Malate-aspartate is active in high-energy tissues (liver/heart), while glycerol phosphate works efficiently in muscle.

purpose of shuttles is to transfer _____ from NADH produced in the _____ (during glycolysis) across the impermeable mitochondrial membrane. This allows the electrons to enter the electron transport chain (ETC) to produce _____, while regenerating to keep glycolysis running... they are reversible

electrons, cytosol, ATP

how is NAD+ and FAD regenerate

oxidative phosphorylation

main purpose of TCA

not ATP directly

producing NADH/FADH₂ for ETC

where is lactate dehydrogenase LDH located in cell

cytosol

how many NTP molecules were produced by catabolism of one glucose molecules by the end of the TCA cycle

4 (2 ATP glycolyis, 2 GTP TCA)

where is pyruvate carboxylase/dehydrogenase located in the cell

mitochondria

which TCA cycle step made GTP

conversion of succinyl CoA to succinate