Exam 1 Flashcards
Fuel stores
+ what happens when we are fasting
After eating, dietary fuel that exceeds body’s immediate energy needs is stored:
- mainly as triacylglycerol in adipose tissue
- glycogen in muscle, liver + other cells
- protein in muscle
When fasting fuel is drawn from these stores + oxidized to provide energy
Metabolic roads to acetyl-CoA
in mitochondria:
Fatty acid (palmitate)
Ketone body (acetoacetate)
Pyruvate
Ethanol (also in cytosol)
Precursors of pyruvate
Glucose
Amino acid (alanine)
How is ATP produced
Respiration: Oxidation of fuels (glucose, amino acids, fats)
1) oxidized to acetyl CoA
2) Oxidized to CO2 in TCA cycle
3) Electrons lost during oxidation are transferred to O2 (final e- acceptor) in ETC
Structure of carbohydrates
Polysaccharide: starch
Disaccharides: sucrose, maltose, lactose
Monosaccharides: fructose, galactose, glucose
carbohydrates
(CH2O)n
n >/ 3
Structure of proteins
composed of amino acids joined together by peptide bonds
Structure of fats
composed of triacylglycerols
> 3 fatty acids esterified to one glycerol moiety
ester bonds are hydrolyzed by lipase (when digested)
Aerobic vs anaerobic metabolism
aerobic: glucose is oxidized completely to CO2 and H2O
anaerobic: glucose is oxidized to lactate
> NADH is oxidized to regenerate NAD+ so it can be used to synthesize pyruvate from glucose again
Which two parameters describe the kinetics of a catalyzed reaction:
Km
- Interaction of enzyme with substrate (binding)
- binding affinity of substrate for enzyme (affinity constant)
Km high = substrate affinity low
Km low = substrate affinity high
Km = Vmax/2
Vmax
- Conversion of substrate into product (catalysis)
- maximal rate of chemical conversion once substrate is in active site
What is a metabolic route
a consecutive series of enzymatic reactions:
product of one enzyme is the substrate for the next enzyme
Inhibition of enzyme reactions
competitive
(when product and inhibitor are the same: product inhibition)
- reversible
- Vmax is equal
- Km becomes higher (initial reaction rate + affinity down)
Irreversible inhibition
- Vmax decreases
- Km stays the same
Which pathways can glucose 6-p go into?
Glycolysis
Pentose phosphate pathway
Glycogen synthesis
Hexokinase
- what it does
- how is it regulated
glucose + ATP > glucose 6-P + ADP
regulation via negative feedback
> Hexokinase: inhibited by glucose-6-phosphate
Tissue specific isoenzyme
- glucokinase is for the liver + is not inhibited by its product
Glucose transport
Firstly passive transport occurs and then active transport to take up final amounts of glucose
Right after eating e.g 50mM in lumen: Glucose conc (~5mM in blood)
> Glucose moves from lumen to blood (from high to low) through the cells
> Passive transporter on apical and basolateral membrane (GLUT2)
What happens when equilibrium is reached: 8mM in lumen and in blood
> active transporters are necessary to take up final amounts of glucose from lumen into capillaries
> symport used (glucose and sodium are transported) (SGLT1)
> ATP required for active transport (indirectly in this case, uses Na+ K+ ATPase to create sodium conc to be used for the symport that transports the glucose)
Compare GLUT2 and SGLT1
GLUT2
uniporter
bidirectional transport
passive transport
SGLT1
symporter
unidirectional transport
secondary active transport
in the small intestine
Affinity of different GLUT isotypes
highest affinity to lowest:
GLUT1
all cells, red blood cells
Kt: 1 mM basal glucose uptake
requires glucose all the time so it has very low Km so it can take up glucose even when there is very little present
GLUT3
neurons, lymphocytes
1 mM basal glucose uptake
GLUT4
muscle cells, adipocytes
5 mM insulin-sensitive glucose uptake
GLUT5
small intestine
10 mM fructose transport
GLUT2
small intestine, liver, beta-cells
20 mM uptake dietary glucose
regulation insulin production (pancreas)
should only be active when we have just eaten, hence the high Km value
Regulation of enzymes: phosphorylation
Kinase adds phosphates to proteins which can produce a charge (e.g negative charges) which can bind a lot of water = switches enzyme from inactive to active or vice versa
Phosphatase can be used to remove phosphate to do opposite (in)activation
Serine, threonine, tyrosine all have hydroxyl groups which can be phosphorylated
Regulation of enzymes: allosteric
provide an example
Binding of allosteric activator shifts equilibrium between active and inactive enzyme conformation
e.g High concentration ADP (binds to allosteric site) signals that ATP formation is needed
> ADP-binding activates (regulation: e.g., of glycolysis)
> This results in accelerated ATP synthesis
What is a rate-limiting enzyme
the enzyme that can be regulated (i.e switched on or off)
What happens to excess glucose after a meal?
the liver stores it as glycogen
but if there is still excess it is converted to fat, namely palmitate
Homeostatic regulation in the fasted state
Energy Maintenance Mode: Balancing blood glucose levels.
Glycogenolysis: Liver breaks down glycogen to release glucose (which then gets transported to the brain, RBC,
Gluconeogenesis: Liver synthesizes glucose from non-carbohydrate sources (e.g., amino acids from muscle protein stores, and TG)
long-chain fatty acids are a major fuel for the liver: released from adipose tissue triacylglycerols > travel to liver asFA bound to albumin
Glucose metabolism in RBC
anaerobic, RBC do not have mitochondria = lactate is produced
Homeostatic regulation in the starved state
Glycogen stores are depleted
ketogenesis: liver produces ketone bodies as alternative fuel for the brain
Gibbs free energy
negative value: release of energy, reaction proceeds forward, exergonic/exothermic
positive value: endergonic, endothermic, backward reaction favoured
energy-requiring processes (deltaG > 0) are driven by energy-generating processes (deltaG < 0).
why mitochondria are so important for efficient ATP generation
Mitochondria are essential for efficient ATP generation as it is the site of oxidative phosphorylation.
Anaerobic glycolysis: in the cytoplasm
Produces a net yield of 2 ATP per glucose molecule.
Aerobic conditions: TCA cycle and ETC can successfully take place in the mitochondria, producing a net yield of 30-32 ATP per glucose molecule. Glucose is fully oxidized in aerobic respiration, generating significantly more energy than the partially broken down glucose in anaerobic respiration.
Co-enzyme A
activated carrier of carbons: strongly negative ΔG
> very large molecule
Which enzyme converts pyruvate to acetyl coA + what reaction takes place + structure + regulation
Pyruvate dehydrogenase
(oxidative decarboxylation)
controlled by NAD+ (gets reduced)
Turns C3 > C2 (the carbon is lost as CO2)
Structure of Pyruvate Dehydrogenase Complex (PDC)
E1 (Pyruvate Dehydrogenase) – Removes CO₂ from pyruvate.
E2 (Dihydrolipoyl Transacetylase) – Transfers the remaining two-carbon group to Coenzyme A (CoA).
E3 (Dihydrolipoyl Dehydrogenase) – Regenerates necessary cofactors.
+ ADP
– NADH and Acetyl CoA
Energy captured in TCA cycle
3NADH (two of them generated in the step when a carbon is lost as CO2, other NADH generated in OAA formation)
1 FAD(2H) (succinate to fumarate)
1 GTP (succinyl coA > succinate)
NADH carries energy as high-energy electrons
What are NAD+ and FAD
They are co-enzymes
FAD is a prosthetic group, part of a flavoprotein
NAD + is a soluble molecule
what happens to the flux through the TCA cycle if a cell has enough ATP
When there is sufficient ATP, flux through the TCA cycle decreases. Subsequent NADH accumulation inhibits various TCA cycle enzymes, such as isocitrate DH and alpha-ketoglutarate DH and malate DH. This results in the accumulation of various intermediates, such as citrate which is a product inhibitor of citrate synthase, reducing the entry of acetyl-CoA into the cycle
ALL NADH PRODUCING STEPS ARE INHIBITED
In the fed state when there is sufficient ATP, what happens to excess Acetyl CoA
excess acetyl-CoA is converted to fat
What stimulates isocitrate DH in the TCA cycle
ADP, signalling more ATP production is needed
What happens when OXPHOS stops recycling NADH?
TCA cycle will stop as well
OXPHOS coupling vs uncoupling
OXPHOS Coupling (Normal Process)
ETC + ATP synthesis are tightly linked.
e- from NADH & FADH₂ travel through the ETC, releasing energy.
This energy pumps protons (H⁺) across the inner mitochondrial membrane, creating a proton gradient.
Protons flow back into the mitochondrial matrix through ATP synthase, generating ATP.
Key Point: Energy from electrons is efficiently captured as ATP rather than lost as heat.
OXPHOS Uncoupling
Protons leak back into the matrix without passing through ATP synthase.
Energy from the ETC is released as heat instead of making ATP.
e.g for thermoregulation Brown fat in babies & hibernating animals uses uncoupling protein (UCP, thermogenin) to produce heat instead of ATP.
ETC
NADH and FAD2H go through ETC
1) Complex I (NADH Dehydrogenase)
Accepts electrons from NADH.
Transfers electrons to Coenzyme Q (Q10).
Pumps 4 protons (H⁺) into the intermembrane space.
- Complex II (Succinate Dehydrogenase)
Accepts electrons from FADH₂.
Transfers electrons to Coenzyme Q but does NOT pump protons - Complex III (Cytochrome bc₁ Complex)
Receives electrons from Coenzyme Q.
Transfers them to Cytochrome c.
Pumps 2 protons (H⁺) into the intermembrane space. - Complex IV (Cytochrome c Oxidase)
Accepts electrons from Cytochrome c.
Transfers them to oxygen (O₂), forming water (H₂O).
Pumps 4 protons (H⁺) into the intermembrane space. - ATP Synthase (Complex V)
Uses the proton gradient created by the ETC to synthesize ATP from ADP + Pi.
Q10 = Co-enzyme Q
- Ubiquinone (Q) – Oxidized Form
No extra electrons (fully oxidized).
Accepts electrons from Complex I (NADH dehydrogenase) or Complex II (Succinate dehydrogenase).
Converts into semiquinone (Q*⁻) after gaining one electron. - Semiquinone (Q*⁻) – Partially Reduced Form
Radical intermediate (unstable, carries one electron).
Important in the Q-cycle of Complex III.
Can either accept another electron to become ubiquinol (QH₂) or lose an electron to return to ubiquinone (Q). - Ubiquinol (QH₂) – Fully Reduced Form
Holds two electrons and two protons (H⁺).
Transfers electrons to Complex III (cytochrome bc₁ complex).
Releases protons (H⁺) into the intermembrane space, contributing to the proton gradient for ATP synthesis.
Converts back to ubiquinone (Q) after electron transfer.
ATP synthase
F₀ Subunit (Membrane-bound) → Acts as a proton channel (pore)
F₁ Subunit (Matrix-facing) → Catalyzes ATP synthesis (headpiece)
Mechanism of ATP Synthesis
Protons (H⁺) flow down their electrochemical gradient from the intermembrane space to the mitochondrial matrix through the F₀ subunit.
Proton movement rotates the F₀ subunit, transferring energy to the F₁ subunit.
The F₁ subunit undergoes conformational changes, converting ADP + Pi → ATP.
Newly formed ATP is released into the matrix for cellular use.
Complex II: link between ETC and TCA cycle
In TCA: catalyzes the conversion of succinate to fumarate, producing FADH₂
In ETC: The FADH₂ transfers its electrons to Coenzyme Q (ubiquinone), which continues electron flow to Complex III.
Gibbs free energy and oxidation reactions
2e- oxidation reactions
most energy to least
methane (-4) > hydroxyl (-2) > aldehyde / ketone (0) > carboxyl (+2) > carbon dioxide (+4)
H > OH > double bonded O > d.b O and another H turned into OH > 2 d.b O
Overview glycolysis
Two subsequent phases in glycolysis: preparative phase (Glucose > fructose 1,6-bisphosphate)
and
ATP-generating phase (fructose 1,6-bisphosphate > 2 triose phosphates > 2 pyruvate)
- Substrate-level phosphorylation
- Oxidation of glucose (C6) to two pyruvate (C3)
- Production of 2 ATP and 2 NADH
- Anaerobic glycolysis, with lactate formation
- Regulation of glycolytic flux (HK, PFK-1, PK)
First step of glycolysis
First step activates glucose.
glucose > glucose 6-P
- via hexokinase / glucokinase and ATP
- Glucose-6-phosphate (G6P) cannot pass the plasma membrane and leave the cell
- Phosphorylation traps and destabilizes the glucose molecule, facilitating its metabolism
- At high concentrations, G6P inhibits hexokinase activity (‘product inhibition’)
metabolic pathways emanating from glycolysis
Glucose 6-P > 5 carbon sugars
Pyruvate > alanine
3 phosphoglycerate > serine
How is NADH from glycolysis transported to OXPHOS
transported to mitochondrion via shuttle system
Thermodynamics of glycolysis
Regulation at HK, PFK-1, and PK, catalyzing irreversible reactions, with DeltaG «_space;0
even more negative for each regulation step
Spontaneous reactions:
the change in Gibbs free energy must be negative (DG < 0), releasing free
energy and allowing the reaction to proceed without external input
Irreversible reactions:
the change in Gibbs free energy is highly negative (DG «_space;0), proceeding in
only one direction, with little to no tendency for the reverse reaction to occur
Glycolysis regulation
ATP controls glycolytic rate
Activation step: phosphorylation of glucose to glucose 6-P by hexokinase which is product-inhibited.
Committed step: conversion of fructose 6-P to fructose 1,6-bisphosphate by PFK-1
allosteric regulation of PFK-1:
– for citrate and ATP
+ for AMP and fructose 2,6-bisp
Final regulatory step: conversion of PEP to pyruvate by pyruvate kinase. Pyruvate kinase
+ by fructose 1,6-bisp (feedforward activation)
– by ATP
Regulation of glycolysis in the muscle
In muscle, glycolysis is controlled by the energy status
PFK
– low pH, ATP, Citrate
+ AMP
Regulation of glycolysis in the liver
In the liver, glycolysis is controlled by fructose-2,6-bisphosphate (F-2,6-BP) (a metabolite)
> activates PFK 1
F-2,6-BP is produced by PFK2 and de-
phosphorylated by FBPase2, with two
catalytic domains on a single protein.
When do RBC and muscle cells produce lactate from glucose
RBC: always bc no mitochondria
Muscle: when O2 levels are low
Cory/Cori cycle
active during intense muscle contraction (anaerobic glycolysis)
a hepatic gluconeogenesis that consumes lactate as its substrate
muscle cells produce lactate, which liver cells convert back to glucose
Anaerobic glycolysis (enzymes)
Lactate dehydrogenase reversibly converts pyruvate into lactate (which is then secreted)
lactic acidosis
when lactic acid production exceeds lactic acid clearance
can be caused by
> decreased oxidation of NADH and FAD2H in ETC = pyruvate converted into lactate
> inhibition of TCA cycle enzymes
Pentose phosphate pathway
glucose 6-P > ribose 5-P
- cytosolic pathway active in all cells
- For production of NADPH and ribose-5-phosphate (C5 sugars) for RNA and DNA synthesis
oxidative phase and non-oxidative phase
Ox phase is irreversible
non-ox phase has reversible reactions
Uses of NADPH
FA synthesis
Glutathione reduction
Cholesterol synthesis
Nucleotide biosynthesis
Oxidative phase of the Pentose phosphate pathway
oxidation and decarboxylation of glucose 6-phosphate
6C > 5C
via 6-Phosphogluconate DH
3 glucose 6-P > (via 6 NADPH, 3 CO2) > 3 ribulose 5-P
regulated by NADP+
Non-oxidative phase of the pentose phosphate pathway
reversible rearrangement and transfer reactions
transketolase is the enzyme catalyzing xylulose 5-P > glyceraldehyde 3-P
transaldolase catalyses
glyceraldehyde 3-P > fructose 6-P
transketolase 1)
C5 + C5 >< C3 + C7
transaldolase
C3 + C7 >< C6 + C4
transketolase 2)
C4 + C5 >< C6 + C3
Galactose metabolism
galactose > glucose 6-P
then either
> glucose (in the liver)
OR
> glycolysis (in other tissues)
Pentose Phosphate Pathway in erythrocytes
hexose-monophosphate shunt
the ribose 5-P is not used to synthesize nucleotides and instead feeds back into glycolysis
+ produces NADPH
Fructose metabolism
Dietary fructose is primarily taken up by the liver
fructose > (fructokinase) > fructose 1-P > glyceraldehyde or dihydroxyacetone -P > glyceraldehyde 3-P > rest of glycolysis
Glutathione
major anti-oxidant in the cell
has a thiol group (SH)
Glutathione metabolism
* Reduced glutathione (GSH) is essential to prevent oxidative damage to proteins
2 GSH + peroxide > (glutathione peroxidase) > GSGG + H2O + ROH
thiol group, forms a disulfide bond
Glutathione reductase reduces oxidized GSSG back to two reduced GSH
The electrons to reduce GSSG are provided by NADPH
ROS in RBC
Reactive oxygen species (ROS) can cause hemolysis
glucose 6-phosphate dehydrogenase deficiency would inhibit NADPH recycling = glutathione reductase behavior inhibited ?
RBCs rely on glutathione (GSH) to neutralize ROS.
Glutathione peroxidase converts H₂O₂ into water (H₂O) using reduced glutathione (GSH).
Glutathione reductase regenerates GSH from its oxidized form GS-SG, using NADPH.
NADPH is produced in the pentose phosphate pathway by glucose-6-phosphate dehydrogenase (G6PD).
If G6PD is deficient, less NADPH is available, reducing the ability to regenerate GSH.
Without enough GSH, H₂O₂ accumulates, leading to increased ROS levels.
Reduction of O2 to water
Successive one-electron reductions of molecular oxygen (O2) yield:
superoxide (O2−.), hydrogen peroxide (H2O2), hydroxyl radical (.OH), and water (H2O)
* Collectively, these three intermediates are called reactive oxygen species (ROS)
* The hydroxyl radical (.OH) is among the most reactive free radical known: powerful oxidizing agent and initiates the oxidative destruction of all types of biomolecules
lactate dehydrogenase
Reaction mechanism: lactate + NAD+ > pyruvate + NADH + H+
catalyzes the transfer of a hydrogen ion (H+) from C2 oxygen and a hydride ion (H–) from C2 to co-enzyme NAD+
Hydrogen ion (H+) to active site His-195 and a hydride ion (H–) to NAD+
function of fatty acids
fatty acyl coA can be converted to:
energy via beta-oxidation ketogenesis
storage as triacylglycerols
membrane lipids as phospholipids and sphingolipids
beta-oxidation
breakdown of fatty acids: b-carbon of the fatty acid is oxidized
fatty acids > acetyl coA
Fatty acids are activated by fatty acyl-CoA synthetase via coupling with CoA
= fatty acyl-CoA (using ATP)
Carnitine Shuttle System
CPT I catalyzes the exchange of CoA with carnitine (a zwitterion), allowing the fatty acyl-carnitine to traverse the mitochondrial membrane
Once inside matrix, CPT-II converts the fatty acyl-carnitine back into fatty acyl-CoA, ready for beta-oxidation
Beta-Oxidation
Cycle repeats until the entire fatty acid is converted into acetyl-CoA
Acetyl CoA then either:
- Enters TCA cycle for ATP production
OR
- Used for ketone body synthesis in fasting states
CoA in beta oxidation
activated carrier of acyl groups
ATP reacts w/ fatty acid > fatty acyl CoA synthetase catalyzes CoA binding to fatty acyl AMP > fatty acyl CoA synthetase catalyzes removal of AMP = fatty acyl coA (bound by thio-ester bond)
What regulated flow of fatty acids
Carnitine antiporter (a translocase) located in the inner mitochondrial membrane
Acyl-CoA cannot pass the mitochondrial membrane
uses CPT1 to change molecule to transfer it from cytosol into matrix
beta-oxidation spiral
1) oxidation (fatty acyl CoA > trans fatty enoyl CoA)
2) Hydration (trans fatty enoyl CoA > hydroxyl acyl CoA)
3) oxidation (hydroxyl acyl CoA > keto acyl CoA)
4) thiolytic cleavage (keto acyl CoA > fatty acyl coA + acetyl CoA)
Go from Cn to Cn-2 per cycle
b-Oxidation spiral of palmitate (C16)
Breakdown of palmitoyl-CoA in b-oxidation spiral
palmitoyl-CoA (C16) > C14 + acetyl CoA (C2)
6 repetitions of b-ox spiral (in total 4 cycles) = generated 8 acetyl CoA (or 7?)
Breakdown of uneven-carbon fatty acids
Propionyl-CoA (C3-CoA) of uneven-carbon fatty acids is converted to succinyl-CoA (C4-CoA) (which enters the
TCA-cycle as an intermediate)
Beta-oxidation and the brain
Brain/neuronal cells have no beta-oxidation because they cannot absorb saturated longchain fatty acids (myelin!).
The brain relies on glucose, the only backup is ketone bodies
ketone body synthesis: ketogenesis
Glucose shortage triggers ketone-body production: Because of low oxaloacetate levels, acetyl-CoA is diverted to ketogenesis
occurs in the liver during fasting/starvation
Brain, muscle, and other tissues can use ketone bodies produced by the liver
2 acetyl coA > (via thiolase) acetoacetyl CoA > (via HMG CoA synthase) HMG CoA > (via HMG CoA lyase, releasing Acetyl CoA) acetoacetate > D-beta-hydroxybutyrate OR (spontaneously) acetone
which are the ketone bodies produced
acetoacetate, D-beta-hydroxybutyrate, acetone
ketolysis
breakdown of ketone bodies
D-beta-hydroxybutyrate > (via D-beta-hydroxybutyrate DH) acetoacetate > acetoacetyl CoA > 2 acetyl CoA
Transport of ketone bodies into the blood
acetoacetate and β-hydroxybutyrate are synthesized in the liver and transported into the blood circulation together with H+ ions by a co-transporter
In the hunger state of healthy individuals: starvation
ketosis
Increased concentrations of ketone bodies in the blood; not dangerous
In untreated Diabetes mellitus Type 1:
ketoacidosis
Transporting such high concentrations of ketone bodies in the blood that the blood acidifies, increasing the risk of coma
fatty acid synthesis
occurs in the liver in the fed state
The 2C building block for fatty acid synthesis is malonyl-CoA in the cytosol
The elongation of fatty acid synthesis starts with the formation of acetyl-ACP and malonyl-ACP, through binding to acyl-carrier protein.
* Fatty acids are synthesized by cycles of the following reactions:
– condensation (to a beta-keto) (C2 > C4)
– reduction (to a beta-hydroxy) (C4 >
– dehydration (to a enoyl bond)
– reduction
(C4 > C6)
triacylglycerol synthesis
occurs in the liver in the fed state
Excess glucose stored as fat (triglycerides)
Precursors: fatty acids or glucose 3-P
Is fatty-acid breakdown simply is a reversal of fatty-acid synthesis
Fatty acid breakdown is not simply a reversal of fatty acid synthesis.
beta-oxidation: in the mitochondrial matrix
FAD and NAD+ are reduced to FAD(2H) and NADH.
Fatty acid synthesis: in the cytosol oxidizes NADPH to NADP+.
Breakdown:
Oxidation, hydration, oxidation, cleaving
Activation step: acyl CoA dehydrogenase step
Acyl carrier: CoA
C2 unit product: acetyl CoA
Synthesis:
Condensation, reduction, dehydration, reduction
Activation step: creation of malonyl CoA
Acyl carrier: ACP Acyl carrier protein and is hooked up to a protein
C2 unit donor: malonyl CoA
both acyl carriers contain vitamin B5
Elongation or desaturation of fatty acids
The endoplasmic reticulum uses other enzymes to convert palmitate to the
required fatty acids: longer, unsaturated
fatty acid synthase
a dimer
Dimerization is essential for the activity of fatty acid synthase, producing palmitate from malonyl-CoA
FA essential nutrients
Omega-3 and omega-6
high-energy state (citrate levels?)
Citrate production is boosted
Use of citrate
Citrate transports acetyl-CoA groups
Citrate transport is coupled to the conversion of NADH to NADPH needed for fatty acid synthesis.
For every molecule of acetyl-CoA that is transported from the mitochondrion to the cytoplasm one molecule of NADPH is generated.
How is a futile cycle of simultaneous breakdown and synthesis is prevented in fatty-acid metabolism
Never simultaneous breakdown and synthesis
Malonyl-CoA (synthesis) blocks entry of fatty acids into the mitochondrion
by inhibiting the carnitine antiporter (CPT1)
Acetyl CoA carboxylase is activated during fatty acid synthesis, which results in high levels of malonyl CoA. Malonyl CoA inhibits CPT I, an enzyme which aids fatty acid oxidation by transporting long-chain fatty acids into the mitochondrion, so that fatty acid oxidation cannot continue. Consequently, fatty acid breakdown does not occur whilst fatty acid synthesis is taking place.
Regulation of acetyl-CoA carboxylase
Activity of malonyl production is controlled at two levels (produced by acetyl-CoA carboxylase):
* by hormonal control: demand of the body, insulin activates phosphatase (causes acetyl-CoA carboxylase to go from inactive form to active form)
* by AMP and citrate: energy status of the cell itself (activates acetyl-CoA carboxylase)
Allosteric regulation of acetyl-CoA carboxylase
+ citrate
– palmitoyl-CoA (Palmitate will inhibit synthesis unless further processed)
Acetyl-CoA carboxylase active only as a polymer
storage of triglycerides from VLDL
stored in muscle and adipocytes
Essential amino acids
Essential: Histidine, Isoleucine, Leucine, Lysine, Methionine, Phenylalanine, Threonine, Tryptophan, valine
Pool of amino acids in the body
blood amino acids can come from dietary protein, endogenous protein, urea, dietary glucose
Blood amino acids can go to synthesis of new proteins and nucleotide, NT, and hormone synthesis
What to do with a surplus of amino acids?
The body does not store amino acids
* Surplus of amino acids is used for:
- energy production (ATP synthesis)
- fat synthesis
- ketone body synthesis (ketogenic amino acids)
- glucose synthesis (glucogenic amino acids)
amino acid breakdown very general view
broken down into carbon skeleton and amino group
From amino acid to energy or fat
alanine reacts with an alpha-keto acid giving another amino acid and pyruvate
- done by transamination: transfer of an amino-group from an a-amino acid to an a-keto acid
Transamination is reversible and the concentration of amino acids determines its fate
Synthesis of non-essential a.a
occurs through transamination
e.g
Some amino acids are linked to metabolites of glycolysis or the TCA cycle
pyruvate <> alanine
oxaloacetate <> aspartate
alpha-ketoglutarate <> glutamate
Liver detoxification of amino acids
a-Amino group of amino acids removed by trans-amination and de-amination (liver)
Surplus of many amino acids lead to:
Step 1: Formation of glutamate intermediate
* The removal of the a-amino group often starts with a transamination reaction in which the a-amino group is transferred to a-ketoglutarate (C5) to form glutamate (C5)
Step 2: Removal of amino-group from glutamate
* Oxidative deamination of glutamate by the enzyme glutamate DH
* Reaction products are: a-ketoglutarate and NH4+
Step 3: Urea production
* Overall reaction with glutamate leads to a carbon skeleton and NH4+
* In humans, NH4+ is converted mainly to urea in the urea cycle
Glutamate DH
predominantly present in the liver + is localized in mitochondria together with some of the other enzymes of the urea cycle
- This compartmentalization ensures that the toxic ammonium ion (NH4+) is contained
- High concentrations of NH4+ are harmful to the brain, but the exact cause of the neurotoxic action of NH4+ is largely unknown.
Which a.a. lead to pyruvate
3C a.a
The amino groups of serine (and threonine) can be directly converted to NH4+.
* This deamination reaction is catalyzed by a dehydratases (because dehydration precedes deamination).
Serine > pyruvate + NH4+
Which a.a. lead to oxaloacetate
4C a.a
e.g
asparagine > (uses H2O, releases NH4+) aspartate > ( oxaloacetate > aspartate > (uses ATP and glutamine, generates AMP and glutamate) asparagine
Which a.a lead to a-ketoglutarate
5C a.a
uses NADP > NADPH + NH3
NH3+ amino group turns into double bonded O (ketone)
Degradation of branched-chain amino acids
The three branched-chain amino acids (BCAA) (valine, isoleucine, and leucine) are degraded via a similar metabolic pathway
Valine and Isoleucine
> propionyl CoA > succinyl CoA (GLUCONEOGENIC)
Leucine and Isoleucine (KETOGENIC)
Isoleucine is gluconeogenic and ketogenic
Destination of the a.a. carbon skeletons
The carbon skeletons of the 20 proteinogenic amino acids are converted into 7 major metabolic intermediates, which enter glycolysis or the TCA cycle
Acetoacetyl CoA (C4)
Acetyl CoA (C2)
Pyruvate (C3)
OAA (C4)
Fumarate (C4)
Succinyl CoA (C4)
alpha-ketoglutarate (C5)
What are the benefits of dividing the urea cycle over two different compartments of the liver cell?
urea synthesis starts inside the matrix of the mitochondria as that is where the toxic NH4+ is so it is accessed faster and the mitochondria has 2 membranes = is it sequestered in the mitochondria so that the dangerous ammonium does not get out into the blood. The rest happens in the cytosol as urea needs to be excreted and cross the plasma membrane. Substrate of urea cycle is in mitochondria but the end product is near the transporter
How many ATP does the synthesis of urea cost
3 ATP are used but net costs 4 ATP because it generates an AMP which costs an ATP to convert it to ADP so that it can enter the ETC
Urea cycle
Urea synthesis is a cyclic process
that occurs only in the liver
* in mitochondrion and cytosol
* Nitrogen atoms of urea are
derived from NH4+ and aspartate
* Carbon atom is derived from
hydrogen carbonate (HCO3–,
hydrated CO2)
* Oxygen atom is derived from
water
Kinetics ATP, ADP, AMP
Conversion of ATP to AMP releases
more energy
Recycling of AMP to ATP costs
one additional ATP
Gluconeogenesis: precursors, process, organs, energy
Precursors:
FA do not get converted to glucose
Lactate, Glycerol (can also be from triacylglycerol breakdown), Amino acids
Not a reversal of glycolysis because pyruvate > PEP is irreversible
Lactate > pyruvate > OAA > PEP > reverse of rest of glycolysis
Glycerol > glycerol 3-P > triose-P > reverse of rest of glycolysis
Amino acids
C3 Alanine > pyruvate > OAA > etc etc
C4 or C5 a.a. > C4 or C5 TCA intermediate > OAA > etc etc
Liver + to a lesser extent the kidneys are responsible for maintaining glucose levels in the blood circulation during fasting through gluconeogenesis
Energy required for the gluconeogenesis is supplied by fatty acids (through b-oxidation)
Reciprocal regulation of gluconeogenesis and glycolysis in the liver
F-2,6-BP is a metabolite that
regulates the balance of
glycolysis/gluconeogenesis in
the liver
* F-2,6-BP is produced by PFK2
and dephosphorylated by
FBPase2
* A bifunctional protein: Two
catalytic domains in one protein
– gluconeogenesis
+ glycolysis
Highly regulated step for gluconeogenesis
Pyruvate to OAA is catalysed by pyruvate carboxyalse (Acetyl-CoA from b-oxidation activates pyruvate carboxylase)
> this is why energy req for gluconeogenesis comes from FA
when is Glucose-alanine cycle active?
glucose-alanine cycle is active during prolonged exercise and starvation
different physiological roles of the amino acids that are present
abundantly in the blood circulation during fasting
Glutamine goes to kidney and gut and is further oxidized (used to synthesize alanine so that energy sources can be maintained by synthesizing glucose and ketone bodies)
Alanine precursor for gluconeogenesis
Why both?
> glutamine has 2 amino groups (alanine does not), gut and kidney get rid of one (to be used for the organs own metabolism), leaving glutamine with 1 which then makes alanine which goes to the liver
glutamine is used to transfer nitrogen through the blood
> in the muscle glutamate reacts with NH4+ giving glutamine which then goes to kidney and gut and used as Fuel (energy source) and Nitrogen donor for purine biosynthesis
DNA process
Dna replication > transcription into RNA (start point is after the promoter) > translation into proteins
Glutamine and glutamate deamination
The kidney can remove the amino-groups of glutamine and glutamate and produce ammonium (NH4+), not urea, to maintain pH of urine
Nucleotides
NMP, NDP, NTP
> mono, di, tri depending on amount of phosphate groups attached to nucleoside (ribose + base)
DNA strand structure
2’-deoxy
one phosphate group is bound to the 5’ end of a ribose nucletide and to the 3’ end of a different nucleotide ribose carbon via phosphodiester bonds = phosphate backbone
base pairing
A - - T
C - - - G
Name the Purine and pyrimidine bases
Purine: A, G (more carbons)
Pyrimidine: C, T
Deoxyribonucleoside triphosphates (dNTPs)
serve as substrates for DNA synthesis
> 4 diff ones for each base
RNA structure
ribose
single stranded, not double helix, has a hydroxyl group and phosphodiester bonds
Converting NTPs into dNTPs
via ribonucleotide reductase (reduction reaction, gaining e- coming from NADPH which comes from the pentose phosphate pathway)
OH reduced to H at the 2’ carbon
which building blocks are purine bases and pyrimidine bases synthesized?
Amino acids contribute nitrogens and carbons (not all the carbons in the nucleic acids are from amino acids though)
Purine bases:
glycine, glutamine, and aspartate
CO2, and N10-formyl-FH4
Pyrimidine bases: a free base > into nucleotides
Aspartate, carbamoyl phosphate (CO2 + Glutamine)
Ribose 5-phosphate is NOT a building block but plays an important role in the synthesis
Ribose 5-phosphate in nucleotide synthesis
Ribose 5-phosphate (from PPP)
5-Phosphoribosyl-1-pyrophosphate (PRPP) synthase (+ ATP co-enzyme) catalyzes the conversion of Ribose 5-phosphate into PRPP
PRPP is used for purine and pyrimidine synthesis and in salvage pathways
One carbon metabolism
e.g Tetrahydrofolate (FH 4 ) carries a single one-carbon group in one-carbon metabolism (+ contributes it to purine synthesis and pyrimidine)
Uses NADPH as co-enzyme
Folate is reduced via dihydrofolate reductase > FH2 > reduced again with same enzyme to FH4 which can now accept 1 carbon from an a.a
De novo purine synthesis
base is synthesized on the activated ribose, PRPP
Activation step: phosphorylation of ribose 5-P into PRPP (via PRPP synthase + 2 ATP)
> committed step: PRPP (+ glutamine + H2O + glutamine phosphoribosyl amidotransferase) > 5-phosphoribosyl 1-amine (had an amine group now added)
addition of glycine ontop of amino group
addition of all the a.a. + CO2 + FH4 (in diff steps)
forms IMP: the branch point for adenine and guanine nucleotide biosynthesis
If there is a lot of GTP, aspartate is used to convert it to AMP > ADP > RNA
or ADP > RR > dADP > DNA
If there is a lot of ATP, glutamine is used to convert it to GMP > GDP > RNA
or GDP > RR > dGDP > DNA
GTP conversion to AMP in purine de novo synthesis
aspartate is used and fumarate is removed, asparate-to-fumarate
conversion = donation of an
amino group, as in the urea cycle
fumarate is an intermediate of the TCA cycle
Purine de novo synthesis regulation
Regulation by negative feedback regulation
Activation step: ADP or GDP inhibits PRPP synthase
Committed step: GMP > GDP > GTP or AMP > ADP > ATP inhibits glutamine phosphoribosyl aminotransferase
Lots of GMP ofc inhibits the conversion step of IMP to GMP
Lots of AMP inhibits IMP > AMP
De novo pyrimidine synthesis
the base is synthesized first, followed by attachment of the activated ribose, PRPP
Activation step: formation of PRPP but not committed step as it can also go to purine synthesis
Committed step: (glutamine + CO2 + 2ATP > carbamoyl phosphate) via Carbamoyl phosphate synthase II
Glutamine delivers the ammonia (NH3)
conversion of the uracil base (RNA)
to the thymine base (DNA) is done by one-carbon metabolism (reductive methylation reaction) using FH4:
- one carbon is added to UMP to make TMP
- i.e difference between U and T is a methyl group
UMP in de novo pyrimidine synthesis
UMP is an intermediate
UMP > UDP and then either UTP or dUDP
Anti-cancer therapies
Thymidylate synthase (conversion of dUMP RNA to dTMP DNA) and dihydrofolate reductase (for FH4) are drug targets in cancer chemotherapy
Methotrexate is a folate analog and a competitive inhibitor of dihydrofolate reductase
5-Fluorouracil inhibits thymidylate synthase via competitive inhibition
* Like methotrexate, 5-fluorouracil inhibits the conversion of the uracil base (RNA) to the thymine base (DNA)
Purine salvage pathways
nucleotides can be converted to nucleosides and free bases
recycling of the following into one another instead of de novo synthesis:
Adenine, AMP, Adenosine, Inosine, Hypoxanthine, IMP, Guanine, GMP, Guanosine
adenosine deaminase (ADA)
in purine salvage pathway
adenosine deaminase (ADA) breaks down dATP
> conversion of adenosine into inosine
SCID (immunodeficiency)
caused by defective adenosine deaminase (ADA), necessary for the breakdown of purines
Lack of ADA = accumulation of dATP
> Accumulation of dATP will inhibit the activity of ribonucleotide reductase (the enzyme that reduces ribonucleotides to generate deoxyribonucleotides for DNA synthesis)
* The effectiveness of the immune system depends upon lymphocyte proliferation and hence dNTP synthesis.
Without active ribonucleotide reductase, DNA synthesis in
lymphocytes is inhibited and the immune system is compromised
Free base generation (purine)
purine nucleoside phosphorylase serparates molecule into ribose 1-P and free purine base
Purine degradation
nucleotide to uric acid to urine
AMP > IMP (via adenosine deaminase)
GMP and AMP are broken down separately and eventually both form xanthine which is broken down into uric acid via xanthine oxidase
Gout + nucleotide metabolism
Gout results from excess uric acid, which forms crystals and causes inflammation. Uric acid is produced in the purine degradation pathway when hypoxanthine and xanthine are converted by xanthine oxidase. Overproduction of purines or reduced uric acid excretion can contribute to gout. Allopurinol inhibits xanthine oxidase, reducing uric acid production and leading to the excretion of hypoxanthine and xanthine instead. Its active form, oxypurinol, competes with xanthine for the enzyme’s binding site.
Addition of nucleotides to infant formulae
nucleotides are non-essential, we can synthesize them on our own = they are not necessary in infant formula
Digestion of dietary carbohydrates
entry of starch, lactose, sucrose
> digestion starts in the mouth w/ salivary alpha-amylase
> broken down into alpha-dextrins
> pancreas secretes alpha-amylase + HCO3-
> broken down into tri- and oligosaccharides, maltose, and isomaltose in the small intestine
Maltase + isomaltase are broken down and move into into intestinal epithelial cells as glucose
sucrose > glucose and fructose via sucrase
Lactose > glucose and galactose via lactase
fiber cannot be digested via the enzymes, it is broken down via bacteria in the colon
Explain why the digestive enzymes do not digest the pancreas
because the enzymes are secreted, stored, and transported in their inactive form (as zymogens)
> only activated in the small intestine
> pancrease also secretes HCO3- which helps keep them inactive as enzymes optimal pH is quite low so bicarbonate helps this optimal pH be avoided
> stomach acid HCl helps activate the enzymes
No zymogens in carbohydrate digestion due to specificity to 1-4 glycosidic bonds
Lipases:
> lipases are activated on the micelles
> lipase is inhibited by bile salts
> colipase kicks out bile salts allowing enzymes to become active
After digestion of dietary carbohydrates what happens
monosaccharides are absorbed
Action of salivary and pancreatic α-amylases
breaks bonds of starch into tri- and oligosaccharides, maltose (alpha 1,4 glycosidic bond), isomaltose (alpha 1,6 glycosidic bond) and alpha-dextrins
> 1 enzyme can break one bond = several enzymes needed
dietary fiber
soluble polysaccharides that cannot be hydrolyzed by human digestive enzymes,
may be hydrolyzed and converted by bacterial enzymes in the colon, forming hydrogen (H2), CO2 or methane CH4 gas, or the short-chain fatty acids: acetate (C2), propionate (C3), and butyrate (C4), or lactate
Maltase and isomaltase activities
Maltase:
cleaves the alpha 1-4 bond of maltose and maltotriose
Isomaltase:
cleaves the alpha 1-6 bond of isomaltose
Sucrase-isomaltase complex
It is a transmembrane protein
> in the intestinal lumen, the sucrase-isomaltase complex is cleaved into its two catalytic domains:
sucrase: hydrolyzes sucrose > glucose and fructose
Isomaltase: hydrolyzes isomaltose
Glucose transporters in intestinal cells
Glucose transport from the intestinal lumen via passive facilitative transport by GLUT2
and via secondary (indirect) active transport by the SGLT1 transporter
GLUT 5 for fructose
Movement of GLUT through the body
GLUT2 transports from pancrease beta cells and the gut, using GK, to the portal vein
> GLUT 2 + GK > liver
from liver to
> GLUT 3 + HK to brain
> GLUT4 + HK to adipose tissue or muscle
> GLUT 1 + HK to RBC
Digestion of dietary proteins
Digestion starts in the stomach
> HCl denatures dietary proteins and pepsin breaks down proteins into peptides
> Pancreas secretes HCO3- and zymogens and proenzymes
> aminopeptidases breakdown proteins further into di- and tri-peptides and amino acids
which are then transported into intestinal epithelial cells > go to the blood
Following carbohydrate digestion, free amino acids and di- and tripeptides are absorbed
On-site activation of dangerous proteases
Gastric and pancreatic proteases are synthesized as inactive precursor proteins (proenzymes/zymogens)
- in the stomach H+ activates pepsinogen > pepsin
- enteropeptidase activates trypsinogen into trypsin
they are activated ONSITE !!!
> Enteropeptidase is bound to the membrane of the intestinal brush border
Secretion of zymogens by the pancreas
- Exocrine cells of the pancreas secrete many different types of digestive enzymes, such as amylases, lipases,
and proteases - Digestive enzymes are secreted as inactive precursors, so-called zymogens or pro-enzymes
- They are secreted via the pancreatic duct into the lumen of the small intestine (duodenum)
Digestion of dietary proteins: Enzyme regulation
Complete ON-OFF regulation: on-site activation of dangerous pancreatic pro-proteases in the duodenum by limited proteolysis, initiated by membrane-bound enteropeptidase
Hydrolysis of peptide bonds in proteins
R–C=O, bonded to N-R
C-N bond is hydrolyzed
R-C bound to O-
H3N+ -R
Differentiation: there is substrate specificity of the different proteases
Amino acid transport through intestinal cells
All via secondary (indirect) active transport of the zwitterion (as the amino acid, after being hydrolyzed has a negatively charged O and a positively charged amino group)
> a.a enter blood stream to form the blood pool of a.a + then taken up by organs
Interorgan amino acid exchange after an overnight fast
done via glutamine and alanine
Renal glutamine metabolism
Glutamine contains two amino groups, which can be removed by subsequent deamination in the kidney, generating two ammonium ions
- Excretion of NH 4+ helps buffer acidemia (acidic blood)
Enterocytes: what do they use for fuel?
they are intestinal epithelial cells
> do not use glucose or fatty acids
as a fuel
> Use amino acids and can use ketone bodies
Glutamate can make citrulline and ornithine (part of urea cycle)
Digestion of dietary fat
TG enters body
> bile salts, HCO3-, lipase, and colipase are released
bile salts aggregate onto the TG
> colipase kicks these off so lipase can break down the triacylglycerol into 2-MG and FA
products combine to form micelles and bile salts aggregate onto them
pancreatic lipase breaks down TG in the micelles (cleaves TG at carbons 1 and 3) into 2-MG and free FA
bile salts can then leave and be recycled
FA and 2-MG are absorbed into intestinal epithelial cells where they form nascent chylomicrons which transport them to the blood
What is absorbed after triacylglycerol digestion
2-monoacylglycerols (2-MG) and
free fatty acids (FA)
> resynthesized into triacylglycerols
Bile salts
strong amphipathic molecules
- Cholesterol is a very hydrophobic molecule, with a hydrophilic hydroxyl group
- The liver produces bile salts from cholesterol
- Like detergents, bile salts emulsify dietary fat to form micelles
Turns into cholate which develops a very hydrophilic (due to its OH and COO- groups) face and other hydrophobic face (amphipathic)
> Bile salts intercalate, hydrophobic part is in triglycerides and hydrophilic part faces the outside (forming an outer barrier kinda)
Critical micelle concentration
- Detergents form micelles above the critical micelle concentration (CMC)
- The critical micelle concentration of bile salts is 5−15 mM
Emulsification of triacylglycerols
Bile salts, but not bile acids, act as emulsifiers
> Due to the emulsifying action of bile salts, large lipid droplets from food are broken down into many lipid micelles
Conjugation of bile salts
Conjugation lowers the pKa of the bile salts (The more ionised the bile salt the higher its solubility), making them better detergents
cholic acid pKa~6
glycocolic acid pKa~4
taurocholic acid pKa~2
Cholic acid−Cholate
Cholic acid is a bile acid (protonated, uncharged)
Cholate is a bile salt (deprotonated, anion, strongly amphipathic) and functions as a detergent
Bile acid <> Bile salt + H +
R−COOH <> R−COO − + H +
COO- increases as you go up pH so if pKa is smaller the peak of COO- 100% conc is reached at an earlier (lower) pH
Active site of pancreatic lipase
Active site of pancreatic lipase is shielded from water by a lid structure
Co-lipase binds pancreatic lipase at the water-lipid interphase, opening up its active site and allowing triacylglycerols to enter for digestion
Pancreatic cholesterol esterase and PLA 2
Pancreatic cholesterol esterase cleaves cholesterol esters (product is cholesterol)
Pancreatic phospholipase A 2 (PLA 2 ) cleaves phospholipids
> PLA 2 acts at the lipid-water interface
Fat metabolism in the fed state
fat from the gut > chylomicrons > lymphatics > TAG > muscle or adipose tissue
fat from liver > TAG > VLDL > TAG > muscle or adipose tissue
Bile salt recycling
recycled in the enterohepatic circulation
Bile salts are synthesized in the liver, stored in the gallbladder, secreted in the duodenum, reabsorbed in the terminal ileum, and returned to the liver by portal blood (safely bound to albumin)
Chylomicron formation and secretion
Intestinal cells absorb free fatty acids and 2-monoacylglycerols, resynthesize them to triacylglycerols (in the smooth ER), and package them as chylomicrons (using apoprotein B-48)
VLDL particles contain apolipoprotein B-100, whereas chylomicrons contain a
truncated version of this protein, apolipoprotein B-48
Nascent chylomicrons are secreted via exocytosis into the chyle (milky fluid) of the lymphatic system
Nascent chylomicrons mature in the blood circulation: liver-produced HDL transfers the accessory proteins ApoE and ApoCII (Activator of lipoprotein lipase (LPL)), forming mature chylomicrons
Fate of chylomicrons
Muscle and adipose tissue secrete lipoprotein lipase (LPL), which is activated by ApoCII to digest triacylglycerols of chylomicrons to free fatty acids and glycerol
The muscle (including the heart muscle) can obtain fatty acids from chylomicrons or VLDL, even if the concentration of lipoprotein particles is low
Lipoprotein lipase (LPL) isozymes
Km of muscle LPL: low
Km of adipose LPL: high
Insulin stimulates synthesis and secretion of adipose LPL
The liver contains apoE receptors on its surface, allowing chylomicron remnants to bind and be taken up by receptor-mediated endocytosis
Cholesterol functions
precursor for steroid hormones, bile acids, vitamin D
component of membranes controlling fluidity
> provides rigidity in plasma membrane by disrupting fluidity
> provides fluidity by disrupting tightly packed membrane
Artherosclerosis
a vascular disease caused by LDL cholesterol, forming plaques in arteries between the endothelial cells and muscle cells = obstruction of coronary artery = myocardial infarction (heart attack) and cerebral infarction (stroke)
Multi-factorial disease
Environment: smoking, diet, lack of exercise
Risk factors: high cholesterol (LDL), low HDL (somewhat disputed), hypertension, diabetes, gender (being male)
Genetic components:
Familial Hypercholesterolemia (FH)
‘genetic predisposition’, indicating multiple different genes
LDL passes endothelial cells (via LDL-receptor, receptor-mediated endocytosis) + is modified into mLDL = dangerous
> modification is on the apolipoprotein
> macrophages identify this as a foreign agent = take this up via phagocytosis = oxidized LDL accumulates in these macrophages (now called foam cells)
Cholesterol characteristics
C27
Largely hydrophobic (tail) due to rings
> hydrophilic polar head group, Hydroxyl group: both H bond acceptor and donor, polar group
How do muscle cells get energy
their main energy is provided by the breakdown of lipids > TCA cycle > OXPHOS
Cholesterol transport
there is a cholesterol transporter that is crucial for cholesterol absorption in the intestine
> Dietary cholesterol is taken up by enterocytes
inhibitors of this transporter can be used to decrease cholesterol absorption
Committed step of cholesterol biosynthesis + why it is an irreversible step
conversion of HMG-CoA to mevalonate, catalyzed by the enzyme HMG-CoA reductase and NADPH as the co-enzyme
> uses 2 NADPH = 2 reduction reactions occur, first reduction = becomes an alcohol, second reduction = becomes an aldehyde (namely mevalonate)
Irreversible because reaction is highly exergonic and has a significantly negative Gibbs free energy value
Where does the energy for cholesterol synthesis come from?
NADPH generated from the PPP
cholesterol synthesis reactions
Acetyl CoA (C2 + C2) > via cytosolic thiolase, acetoacetyl CoA (C4) > via cytosolic HMG-CoA synthase and another C2, HMG CoA (C6) > via HMG-CoA reductase, mevalonate > then phosphate groups added in each consecutive reaction > CO2 cleaved to give isoprene (C5) (equilibrium established) > isoprene added to another molecule > C30 cholesterol > cleaving of CO2 until C27 cholesterol is formed
Mechanisms for Regulation of cholesterol synthesis
Transcription regulation
SREBP is a transcription factor anchored in the ER membrane.
It is bound to SCAP, which acts as a sensor for cholesterol levels.
- High cholesterol levels: SCAP retains SREBP in the ER, preventing cholesterol synthesis.
- Low cholesterol levels drop: SCAP transports SREBP out the ER
Transcription factor SREBP binds
the SRE and increases transcription of the HMG-CoA reductase gene
Sterol regulation
sterols activate HMG CoA reductase proteolysis/degradation
AMP and insulin regulation
AMP and insulin have opposing effects on the activity of HMG-CoA reductase
> A lot of AMP = too little ATP to make cholesterol = AMP kinase, deactivates the enzyme
> Insulin suggests a lot of energy = activates phosphatase = activation of HMG CoA reductase
Low cholesterol or deficient LDL receptor lead to increased synthesis of cholesterol via activation of SREBP
What aspect of cholesterol metabolism makes it difficult to achieve optimal homeostasis?
there is no cholesterol breakdown process = one you have cholesterol it accumulates
cholesterol into cholesterol ester
done via ACAT (acyl-Coenzyme A
cholesterol acyltransferase)
> turns into a neutral lipid droplet
Bile salt secretion and re-uptake
Secretion: Bile salts are formed from cholesterol and transported via bile to the intestine.
Re-uptake: Bile salts solubilize lipids from food and are re-absorbed (95%) by the terminal ileum.
Inability to reabsorb 5% of bile salts allows limited removal of cholesterol from the body, from liver to feces
Bile acid biosynthesis
Cholesterol > cholic acid
bile acids inhibit this pathway
plasma lipoproteins
insoluble
phospholipids form the outside layer (hydrophilic head outside, hydrophobic tails inside)
cholesterol esters + TAG inside them
cholesterol integrated into phospholipid outer layer
Which organ synthesizes cholesterol, what can happen with the cholesterol after?
Liver is main synthesizer of cholesterol
Where does cholesterol go?
It can either make lipoproteins, or store it
Liver stores cholesterol in lipid droplets in adipocytes
Lipoprotein sizes
chylomicrons > VDLDL > LDL > HDL
chylomicrons vs LDL
chylomicrons have more TG than chol
LDL have more chol than TG
Reverse lipoprotein transfer
HDL bringing cholesterol back to liver
> meantime in the blood it converts cholesterol to cholesterol esters via LCAT
After eating, Triacylglycerols in chylomicrons
chylomicrons excreted from intestinal epithelial cells > lymph > blood > LPL breaks down chylomicrons = FA can be distributed to organs
muscle (including heart muscle) can obtain fatty acids from chylomicrons or VLDL, even if the concentration of lipoprotein particles is low
Lipoprotein lipase (LPL) isozymes
K m of muscle LPL: low
K m of adipose LPL: high
Insulin stimulates synthesis and secretion of adipose LPL
Critical micelle concentration (CMC)
the lowest concentration of surfactants in a solution at which micelles start to form
> micelles form so hydrophobic parts can be hidden from water
FA and cholesterol Between meals or fat-free diet
fatty acids and cholesterol are synthesized from carbohydrates and protein via VLDL
LPL lipoprotein lipase
breaks down: Triacylglycerol + 3 H2O > 3FA + glycerol
Triglyceride content of lipoprotein is depleted, thus lipoprotein becomes smaller
Same process as chylomicrons > chylomicron remnants
- Activated by insulin
- requires cofactor apo C-II
- present in endothelium of capillaries
Localization: outside of plasma membrane of endothelial cells that cover the capillaries
NB: LPL is NOT produced by endothelial cell
Forward Cholesterol Transport vs. Reverse Cholesterol Transport
FCT
- liver to peripheral cells
- delivers cholesterol to peripheral cells
- VLDL + LPL gives off MG and FA > IDL + LPL gives off MG and FA > LDL
RCT
- peripheral cells to liver
- Removes excess cholesterol via HDL to prevent buildup
- (pre-)HDL from liver and intestine takes up excess cholesterol
in peripheral tissues and transports it to the liver
- cholesterol esters can be sent to liver via CETP
- SRB1 takes cholesterol into liver
Peripheral Cells:
take up LDL via LDL receptors, release cholesterol
cholesterol can be converted to cholesterol esters via ACAT for storage and membrane protection
Export excess cholesterol via HDL in RCT
- cholesterol goes to HDL via transporter ABCA1 transporter
Liver Cells:
Produce and send out cholesterol via LDL in FCT
Take in excess cholesterol via HDL for excretion in bile (RCT)
also have LDL receptors (making the liver a cholesterol sensor)
Familial Hypercholesterolemia (FH)
results from defects in LDL receptor = accumulation of VLDL, IDL, LDL
Statins competitively inhibit the committed step of cholesterol synthesis: can be dosed for individual patients
> prevents binding of substrate HMG-CoA
CETP
transfers CE from HDL to vLDL
Increase HDL (-cholesterol) by inhibiting CETP??
Excretion of cholesterol
Only way to excrete cholesterol is via bile salts, but retained via the enterohepatic circulation
Malonyl CoA
product of acetyl CoA carboxylase reaction
provides carbons for palmitate synthesis
inhibits CPTI
levels are elevated in fed state
