Metabolism Mark Shepard Flashcards
What 2 molecules feed the ETC with electrons? Where are these produced?
NADH and FADH2
Produced during glycolysis and via the krebs cycle`
Movement of electrons along the respiratory complexes produces ATP by what mechanism?
Oxidative phosphorylation
Give some types of electron carrier molecules, what they use to carry electrons and their names in different oxidative states?
1) Flavoproteins: Use flavo nucleotides
oxidised: FAD
reduced: FADH2
2) Ubipuinones: use a benzoquinone
Q = ubiquinone
QH
QH2 = ubiquinol
3) Cytochromes: use iron containing heme group
Fe2+ = ferrous
Fe3+ = ferric
4) Iron-sulfur proteins: iron complexed with sulfur
5) Copper: uses copper complexed to cysteine or heme
What are the structures of the 3 classes of cytochromes? (type a,b and c)
A) 2 Histadine amino acids and a isoprenoid chain
B) 1 Histadine AA
C) 1 Histadine AA and 2 Cysteine AA’s
Each class in distinguished by its absorption. What wavelengths does each absorb at?
A = 600nm B = 560 nm C = 550 nm
What are the 5 main structures involved in the ETC?
1) Complex I or NADH dehydrogenase
2) Complex II or Succinate dehydrogenase
3) Complex III or Cytochrome bc1 complex or cytochrome c oxireductase
4) Complex IV or cytochrome C oxidase
5) Coenzyme Q or Ubiquinone
Describe what occurs at complex I in the ETC
1) Complex I catalyses transfer of a hydride ion (2e-) from NADH to flavinmononucleotide (FMN) (NADH + H+ —> NAD+)
2) 2 electrons from FMN pass through a series of Fe-S (iron) centres to the Fe-S protein N-2
3) 2 electrons transfer from N-2 in the matrix arm of the complex to coenzyme Q to produce ubiquinol (QH2)
4) Process drives translocation of 4 protons from matrix into intermembrane space
Describe what occurs at complex II in the ETC
1) Succinate (from krebs cycle) is converted to fumarate and passes electrons through flavin (FAD) and several Fe-S centres in complex II on the way to coenzyme Q
2) Donation of electrons through FAD by glycerol-3-phosphate catalysed by G3P dehydrogenase
3) Acyl-CoA dehydrogenase also provides enzymes via electron transferring proteins (ETFs)
What are the 3 subunits of complex III?
Rieske Fe-S proteins (2Fe-2S) Cytochrome C1 - Interacts with cytochrome c for final step of electron transfer to complex IV Cytochrome b - ubiquinone binding site
What is the name for the process that occurs at complex III in the ETC and what are the steps?
Q cycle model - 2 molecules of QH2 are oxidised to Q - Each QH2 donates: > 1 electron to cytochrome c > 1 electron to a molecule of Q (1st step) or Q- (2nd step) regenerating 1 QH2 between them i.e 2 QH2 goes in, 1 QH2 comes out
What is the net reaction of QH2 oxidation within complex III?
- transfers 2 electrons to 2 different cytochrome c molecules
- 2H+ consumed in the matrix
- 4H+ translocated into intermembrane space
How are protons translocated at complex III? What is this called?
- Protons on QH2 are released into intermembrane space by cytochrome bc1
- Known as vectorial proton translocation
What are the 3 subunits of the functional core of complex IV?
1) 2 heme groups (a & a3) and CuB
Fe of a3 and CuB form a Fe-Cu centre
2) 2 Cu ions (CuA) that resembles a 2Fe-2S centre
3) Poor understanding
What occurs at complex IV in the ETC?
Cytochrome c (carrying e-) travels from complex III to complex IV Electron transfer from cytochrome c to CuA, to heme a, to heme a3-CuB centre and finally onto O2 to produce H20
What is the net production of 4 cytochrome c molecules passing 4 electrons through complex IV?
4 H+ used from the matrix to make 2H2O and 4 H+ pumped into the intermembrane space
How is energy provided to translocate protons across the inner mitochrondrial membrane?
ETC provides energy via thermodynamically favourable electron transfer reactions
What is produced by the movement of protons?
An electrochemical gradient
Electro - charge separation
Chemical - difference in pH
What is the electrochemical gradient of protons in the mitochrondria often referred to as?
The proton motive force
What is the chemiosmotic model?
That the proton motive force drives ATP synthesis
Give some similarities between ATP synthesis by photophosphorylation (PP) and oxidative phosphorylation (OP)
1) Reaction centres, electron carriers and ATP forming enzymes are located in an intact membrane that is impermeable to H+ (unless transported)
2) Can be uncoupled from electron transport mechanisms by reagents that promote H+ passage through the membrane
3) ATP synthesis is catalysed by ATP synthases that both have FoF1 sub-structures
4) Both inhibited by venturicidin
What type of ATPase is ATP synthase?
F-type
What are the main 2 structures of ATP synthase and where is each found?
Fo portion found in the inner mitochondrial membrane
F1 portion found in the matrix
What subunits does F1 contain?
3 alpha 3 beta 1 gamma 1 lower case delta 1 epsilon
What subunits does Fo contain?
a,b and c in proportions ab2c10-12
What reaction does ATP synthase catalyse and how does it achieve the energy for this?
ADP + Pi —> ATP + H2O
Uses energy via flow of H+ from P side (intermembrane) to N side (matrix)
What are the 3 conformations of the alpha-beta pairs in the F1 portion of ATP synthase?
ADP bound, ATP bound and empty
What is the energy from the proton gradient used for?
Used to release bound ATP
Formation of ATP uses very little energy (reversible reaction)
Describe the relationship between Fo and F1 and what occurs when protons flow through the membrane
- b2 and delta of Fo associates with an alpha-beta pair in F1, holding them in a fixed position relative to the membrane
- The Fo cyclinder of c subunits is attached to the F1 gamma and epsilon
- As the H+ flows through the membrane, the cyclinder and shaft rotate and the beta subunits of F1 change conformation as the interactions with the gamma subunit changes
How was the rotation of Fo shown experimentally?
F1 was bound to a microscope slide
C subunit was biotinylated (covalent binding of biotin)
Avidin was bound to biotin
Fluorescent actin was attached to avidin
ATP provided as substrate, rotation of actin observed
What occurs upon each 120 degree turn of the gamma subunit?
1) Converts beta-ATP to beta-empty and the ATP dissociates
2) This promotes the condensation of ADP + Pi to form ATP
3) beta-empty site becomes beta-ADP and loosely binds ADP + Pi
How many protons does ATP synthase use to make one ATP?
3
What is the net amount of protons that move per ATP produced and where do the rest come from?
4
1 proton is transferred by the phosphate translocase (symporter) along with dihydrogen phosphate (H2PO4-) (phosphate needed for ATP synthesis)
What is the P/O ratio?
Phosphate/Oxygen ratio
Amount of ATP produced from the movement of two electrons through a defined electron transport chain
What was the equation originally assumed in order to determine the amount of ATP produced from oxidative phosphorylation?
xADP + xPi + 1/2O2 + H+ + NADH —> xATP + H20 + NAD+
where x equals P/O ratio as an integer
What were the assumed P/O ratios of NADH and Succinate?
NADH - 3
Succinate - 2
Why is it difficult to measure P/O ratios experimentally?
1) Mitochondria consume ATP in many other reactions in the matrix
2) Mitochondria consume oxygen for purposes other than oxidative phosphorylation
What must be considered to gain the true P/O ratio?
1) How many protons are pumped out during oxidation of NADH
2) How many protons leak back across the membrane
3) How many protons must pass through ATP synthase to synthesise ATP
What are the true P/O values for NADH and succinate and how are they determined?
- Each NADH will pump 10H+ into the mitochondiral intermembrane space
- Each succinate (via FADH2) will pump 6H+
- 4H+ are used to fully synthesise 1 ATP
10/4 = 2.5 for NADH
6/4 = 1.5 for succinate
Why must NADH rely of alternative pathways to deliver electrons to the ETC?
Because it cannot freely diffuse from the cytosol to the matrix
What are the names of the 2 pathways NADH uses to move to the matrix?
1) Malate-Aspartate shuttle
2) Glycerol-3-phosphate shuttle
Describe how the malate-aspartate shuttle allows cytosolic NADH to pass its electrons to the ETC
1) Reducing equivalents (can be lone electron or hydrogen atom) from cytosolic NADH are transferred to oxaloacetate to yield malate, catalysed by malate dehydrogenase (NADH converted to NAD+) in intermembrane space
2) Malate passes through inner membrane via malate alpha-ketoglutarate transporter
3) Reducing equivalents passed to NAD+ by matrix malate dehydrogenase to form matrix NADH (malate converted back to oxaloacetate)
4) NADH passes electrons to respiratory chain
5) Oxaloacetate converted to aspartate in matrix and transported into intermembrane space by glutamate-aspartate transporter
6) Aspartate converted to oxaloacetate, ready to be converted to malate
Where in the body is the Malate-Aspartate shuttle mostly used?
Liver, kidney and heart mitochondria
How does the glycerol-3-phosphate shuttle differ from the malate-aspartate shuttle?
1) Produces FADH2 from FAD on the inner membrane instead of NADH from NAD+ in the matrix
2) Delivers electrons to coenzyme Q (ubiquinone) and thus into complex III instead of complex I
Where in the body is the Glycerol-3-phosphate shuttle mostly used?
Skeletal muscle and brain mitochondria
What effect do ADP + Pi and NADH have on mitochondrial energy metabolism control?
ADP + Pi: accelerate the citric acid cycle and oxidative phosphorylation NADH inhibits: - pyruvate kinase - pyruvate dehydrogenase - citrate synthase - isocitrate dehydrogenase - alpha-ketoglutarate dehydrogenase (ATP also inhibits many of these)
Why are mitochondria susceptible to their own set of diseases?
Because they have their own genome
What is Leber’s hereditary optic neuropathy (LHON) and what is it caused by?
- Affects CNS and causes vision loss in adulthood
- Single ND4 base change causes Arg –> His mutation in complex 1
- Mitochondrial defect in electron transfer from NADH to complex 1
- Also seen with single base change in cytochrome b gene
What is Myoclonic Epilepsy & Ragged-Red Fibre Disease (MERRF) and what is it caused by??
- Causes uncontrolled muscle jerks
- Mutation of mitochondrial gene that encodes tRNA specific for lysine
- Defective production of several proteins
- Skeletal muscle fibres have abnormally shaped mitochondria
What 2 factors cause mitochondrial heterogeneity among daughter cells?
1) Not all mitochondria in a cell will encode the disease causing mutation
2) Unequal separation of mitochondria during cell division
How are mitochondria diseases inherited?
Maternally inherited
- Males carry their mothers mtDNA but only females pass it on to their offspring
What are the 4 main disease results of inherited metabolic diseases?
1) Accumulation of substrate
2) Accumulation of a normally minor metabolite
3) Deficiency of product
4) Secondary metabolic phenomena
Give an example of a metabolic disease that is due to accumulation of substrate
Phenylketonuria
What is phenylketonuria and what is it caused by?
- Mutation in human phenylalanine hydroxylase (PAH) gene
- Body cannot break down phenylalanine so it accumulates in blood, brain and urine
Give an example of a disease caused by the accumulation of a normally minor metabolite
Cataracts in patients with galactosemia results from accumulation of the sugar alcohol galacticol
What are the 4 main modes of inheritance of metabolic disorders?
1) Autosomal recessive
2) Autosomal dominant
3) X linked
4) Mitochondrial inheritance
Give some examples of autosomal recessive metabolic disorders
- Tay Sachs disease
- Phenylketonuria
What is the name of disorders that have variable symptom severity?
Variable penetrance
Give an example of autosomal dominant disorder that shows variable penetrance
Variegate Porphyria
Give an example of an x linked metabolic disorder
Hunters Disease
Give some examples of important molecules that amino acids are precursors of
Hormones Coenzymes Alkaloids Cell wall polymers Antibiotics Pigments Neurotransmitters
Give some specific examples of where amino acids are involved in synthesis of important molecules
1) Glycine used to create haem
2) Arginine, glycine and methionine are required for synthesis of phosphocreatine
3) Phenylalanine and Tyrosine are used to synthesise alkaloids: found in cayenne pepper, vanilla, cloves
4) Nitric oxide (bacteriocidal) made from Arginine
What are the 3 examples (that we need to know) of AA that produce neurotransmitters? Which neurotransmitter do they produce?
1) Tryptophan —> Serotonin (Depression)
2) Glutamate —> GABA (gamma-aminobutyrate)
3) Histidine —> Histamine (Allergies)
What type of neurotrasmitter is GABA and what is underproduction associated with?
- Inhibitory neurotransmitter
- Underproduction associated with epileptic seizures
What type of drug is used to treat epilepsy and give an example
GABA analogues
Diazepam (Valium) - binds to GABA receptor, but at a site distinct from endogenous GABA
What enzyme converts glutamate to GABA?
Glutamate decarboxylase
What enzyme converts histidine to histamine?
Histidine decarboxylase
How is aspartame synthesised?
Dipeptide of phenylalanine and aspartate
What is phenylketonuria?
Rare congenital disease in which phenylalanine cannot be metabolised due to a deficiency in the enzyme phenylalanine hydroxylase (PAH)
Phenylalanine and phenylpyruvate builds up in blood and brain
What is the consequence of an accumulation of phenylalanine?
Prevents transport of amino acids across the blood brain barrier, inhibiting synthesis of key neurotransmitters and disrupting protein synthesis in the brain Can lead to: Learning difficulties Behavioral problems Epilepsy and seizures
What 2 things must PKU sufferers limit in their diet?
1) Aspartame (can be broken down into phenylalanine)
2) Protein
How are people tested for PKU?
Phenylalanine detected in blood samples using tandem mass spectometry
The conversion of phenylalanine to tyrosine by phenylalanine hydroxylase utilises what other process?
Conversion of tetrahydrobiopterin (BH4) to dihydrobiopterin (BH2)
Other than PAH deficiency, what other metabolic deficiency can cause PKU?
Deficiencies in BH4 regeneration
Tyrosine and BH4 are required for the synthesis of which 3 neurotransmitters?
Serotonin, epinephrine and dopamine
What can be supplemented to treat the neurotransmitter deficiencies?
1) 5-hydroxytryptophan (precursor of serotonin)
2) L-DOPA (precursor of dopamine. Dopamine –> norepinephrine –> epinephrine)
Name 4 uses for nucleotides and give examples
1) Precursors for DNA and RNA
2) Carriers of energy - ATP, GTP
3) Cofactors - FAD, NAD, CoA
4) Second messangers - cAMP, cGMP
What are the 2 nucleotide synthesis pathways and describe each?
1) De novo pathway
- synthesised from metabolic precursors: amino acids, ribose-5-phosphate, CO2 and NH3
2) Salvage pathway
- recycle free bases and nuclosides released from DNA + RNA breakdown
What are the 2 main products of pyrimidine metabolism and how are each metabolised?
1) NH4+ - converted to urea via urea cycle
2) End product is Methylmalonyl-semialdehyde - can be converted to succinyl-CoA vi the valine catabolic pathway
Describe the degeneration of AMP (purine)
1) AMP loses phosphate through action of 5’-nucleotidase
2) Deamination by adenosine deaminase yields inosine
3) Cleavage of ribose by hydrolysis to hypoxanthine
4) Oxidation to xanthine
5) Further oxidation to uric acid
Describe the degeneration of GMP (purine)
1) GMP loses phosphate through 5’-nucleotidase
2) Cleavage of ribose by hydrolysis to yield free guanine
3) Deamination to yield xanthine
4) Oxidation to uric acid
What is gout and what is it caused by?
- Joints become painful and arthritic due to deposition of sodium urate crystals
- caused by excess uric acid
- kidneys also affected
Who is commonly affected by gout and what genetic factor can cause it?
Common in males, usually results from under excretion of uric acid
Can be caused by genetic defect in an ABC transporter involved in urine excretion
What are the 2 main methods of controlling gout symptoms?
1) Dietary changes e.g reducing intake of meat and seafood, consuming vitamin C, limiting alcohol and fructose consumption and avoiding obesity
2) Non-steroidal anti-inflammatory drugs and pain killers
What are the symptoms of gout?
- severe pain in joints
- joints feeling hot and tender
- swelling of affected joint
- red, shiny skin over the affected joint
What is the main treatment of gout?
- Allopurinol
- Xanthine oxidase inhibitor
- Xanthine oxidase converts hypoxanthine to xanthine and xanthine to uric acid
- prevents uric acid production
What evolutionary loss has made gout common in humans?
Loss of urate oxidase
- Urate oxidase converts uric acid into allantoin
How are free purine bases processed?
- Processed to produce their nucleoside monophosphate e.g for adenine:
> Adenine + PRPP —-> AMP + PPi
> enzyme for that conversion is adenosine phosphoribosyltransferase - Guanine and hypoxanthine (adenine catabolites) are both salvaged by hypoxanthine-guanine phosphoribosyltransferase
What is Lesch Nyhan Syndrome?
- Lack of hypoxanthine-guanine phosphoribosyltranserase activity
- Mental retardation, hostility
- Brain is especially dependant on salvage pathway
- Purine levels rise via novo synthesis, leading to excess uric acid, producing gout like tissue damage
How are dietary proteins, excess amino groups and excess NH4+ dealt dealt with?
1) Dietary protein is degraded to AA’s in the GI tract
2) Excess amino groups may be tranferred to pyruvate to produce alanine
3) Excess NH4 in most other tissues is converted to sidechain nitrogen of glutamine
What are the 2 most abundant amino acids in most tissue?
Glutamine and glutamate
Once amino acids from dietary proteins have been transported to the liver, what is the first step in amino acid catabolism?
- Removal of an alpha-amino group to produce an alpha-keto acid, catalysed by an aminotransferse
- Aminotransferase utilises pyridoxal phosphate (PLP) as cofactor
- Amino group transferred to alpha-ketoglutarate, converting it to glutamate
- Known as transamination
What then occurs to the produced glutamate?
- Deamination of glutamate back to alpha-ketoglutararte by L-glutamate dehydrogenase
- Uses either NAD+ or NADP+ as electron acceptor
- Releases ammonia in the liver
What occurs to free ammonia generated in various tissues?
- Excess free ammonia is incorporated into glutamine by glutamine synthetase
- Glutamine transported in the bloodstream to the liver for subsequent processing into glutamate by glutaminase (Glutamine to glutamate releases 1 ammonia)
Why is it important that excess ammonia is removed from tissue?
Ammonia is toxic to animal cells
What is ammonia ultimately converted into for excretion?
Urea
Why is urea a good candidate for nitrogen excretion?
- soluble
- non toxic
- energetically inexpensive
In what 2 forms can ammonia enter the urea cycle?
1) Aspartate
Glutamate + Oxaloacetate (linked to krebs cycle) a-keto-glutarate + aspartate (using aminotransferase)
3) Ammonia delivered via portal vein is incorporated into Carbamoyl Phosphate by Carbamoyl Phosphate Synthetase I which enters cycle
What are the 4 enzymatic steps of the Urea Cycle?
1) Ornithine transcarbamoylase
- converts ornithine and carbamoyl phosphate to citrulline
2) Argininosuccinate sunthetase
- converts citrulline and aspartate to argininosuccinate
3) Argininosuccinase
- converts argininosuccinate to arginine
- reaction releases fumarate - linked to krebs cycle
4) Arginase
- Regeneration of ornithine and formation of urea
What are the 2 N-acquiring reactions in the urea cycle?
1) Reaction catalysed by carbamoyl phosphate synthetase I, the first nitrogen enters from ammonia
2) Reaction catalysed by argininosuccinate synthetase, the second nitrogen enters from aspartate
What occurs to the flux of nitrogen through the urea cycle during a high protein diet and during starvation?
- During high protein diet, C-skeletons are used for fuel and high levels of urea are produced from amino groups
- During starvation, muscle protein is broken down and C-skeletons are used for fuel and high levels of urea are produced from amino groups
How is the urea cycle regulated for a high protein diet or starvation? (2 ways)
1) 4 urea cycle enzymes and carbamoyl phosphate synthetase I are synthesised at higher rates
2) On a shorter timescale, N-acetylglutamate allosterically activates carbamoyl phosphate synthetase I
How can specific defects in urea cycle metabolism be detected?
By identification of accumulated metabolites
What kind of diet can’t be tolerated by people with genetic defects in urea synthesis?
Protein rich diet
Why is a protein free diet not a valid treatment option?
Because humans cannot synthesise 9 of the 20 AAs
What are the 4 treatments for defects in urea cycle enzymes?
1) Limiting protein intake
2) Increase renal excretion of ammonia
3) Use of biochemical tricks to remove ammonia
- Using benzoate (administered in diet)
- glycine is replenished y liver enzyme glycine synthase, which consumes ammonia
4) Specific therapies to correct particular enzyme deficiencies
Give some examples of specific therapies to correct particular enzyme deficiencies
1) Deficiency in N-acetylglutamate synthae results in the loss of the normal activator of carbamoyl phosphate synthetase I
- Treated by administering carbamoyl glutamate (analogue of N-acetylglutamate synthase)
2) Supplementing diet with arginine useful for treating deficiencies of ornithine transcarbamoylase (OCTD), argininosuccinate synthetase and argininosuccinase
3) Rare cases of arginase deficiency, arginine must be excluded from diet
There is a relationship between high levels of cholesterol in the blood and what disease?
Cardiovascular disease
What are the main roles of cholesterol?
Cell membranes
Precursor of steroid hormones and bile acids
What are the 2 ways in which humans acquire cholesterol?
Diet and biosynthesis
What does high cholesterol in the blood cause?
Accumulation of cholesterol in the blood vessels (atheromas) causing hardening and narrowing of the vessels (atherosclerosis)
Atheromas in which part of the body are the leading cause of death in the UK?
Atheromas causing blockage in the coronary arteries that supply the heart with blood
(coronary heart disease)
Where are all 27 carbons found in cholesterol supplied from? (C27H46O)
Simple precursor: Acetate
Cholesterol synthesis involves what type of intermediates?
Isoprene intermediates
What are the 4 stages of cholesterol synthesis?
1) Synthesis of mevalonate from acetate
2) Conversion of mevalonate to 2 activated isoprenes
3) Condensation of 6 isoprene units to form squalene
4) Cyclisation of squalene to the 4-ring steroid nucleus
Describe the synthesis of mevalonate from acetate
1) Acetate used to synthesise acetyl-CoA
2) 3 molecules of acetyl-CoA condense to form HMG-CoA catalysed by HMG-CoA synthase
3) HMG-CoA is reduced to form mevalonate by HMG-CoA reductase
Describe the conversion into isoprene units
1) 3 phosphate groups transferred to mevalonate from 3ATP
2) Pi and carboxyl groups leave producing the activated intermediate ^delta3-isopentenyl phyrophosphate
3) Isomerises to yield second intermediate dimethylallyl pyrophosphate
Describe the condensation of 6 isoprene units to form squalene
1) isopentenyl phyrophosphate and dimethylallyl pyrophosphate undergo head to tail condensation
2) Another head to tail condensation yields farnesyl pyrophosphate (3 isoprenes have now been used)
3) Head to Head condensation of 2 farnesyl pyrophosphates and loss of 2PPi results in squalene
Describe the cyclisation of squalene to the 4-ring steroid nucleus
1) Oxidation and reduction via squalene monooxygenase produces an epoxide intermediate
2) Double bond structure of epoxide intermediate allows cyclase enzyme to convert linear molecule to cyclic structure
What intermediate is formed in animals and how is this converted to cholesterol?
In animals, forms intermediate Lanosterol
Converted to cholesterol via 20 reactions
Where does the majority of cholesterol synthesis take place?
In the liver
What are the 3 main forms in which cholesterol is exported from the liver?
1) Biliary cholesterol
2) Bile acids; hydrophobic derivatives that aid in lipid
digestion
3) Cholesteryl esters
What occurs to cholesterol for storage and transport?
Esterification to even more hydrophobic form
Cholesterol is the precursor to which vitamin?
D
How is cholesterol transported in the plasma?
Attached to plasma lipoproteins e.g low density lipoprotein
What is the composition of LDL?
1,500 molecules of cholesteryl esters 500 molecules of cholesterol 800 phopholipids Single Apolipoprotein B-100 (Each type of lipoprotein has a different composition)
What are the 4 main types of lipoproteins from low to high density?
Chylomicrons
VLDL
LDL
HDL
What is the correlation between HDL and arterial disease and LDL and arterial disease?
-ve correlation between HDL levels and arterial disease
+ve correlation between LDL levels and arterial disease
What is the name for HDL removing cholesterol from cells and what interaction occurs?
Reverse cholesterol transport
- involves interaction of ApoA-I with ABC1 protein in cholesterol rich cells
How does HDL process excess cholesterol?
- Lecithin-cholesterol acyl transferase (LCAT) on surface of HDL converts cholesterol to cholesteryl esters
- Cholesterol rich HDL then returns to the liver
How is cholesterol taken up into cells?
- Receptor mediated endocytosis
- LDL has ApoB-100 that is recognised by LDL receptor on cell surface - LDL taken into endosome
- Lysosomes containing enzymes fuse with endosome and hydrolyse cholesteryl esters, releasing cholesterol and fatty acids
- Cholesterol may then be incorporated into membranes or re-esterified for storage
How is cholesterol biosynthesis regulated?
- In mammals; controlled by intracellular concentrations of cholesterol and by insulin and glucagon
- Mediated by transcriptional regulation pf HMG-CoA reductase gene and other genes induced in synthesis of cholesterol
What is the role of sterol regulatory element-binding proteins (SREBP)?
- Bound to SREBP cleavage activating protein (SCAP) and are inactive when cholesterol levels are high
- When cholesterol is low the active domains of STEBPs are cleaved and migrate to the nucleus to initiate trancription of target genes
Describe how insulin and glucagon are involved in cholesterol control
- Covalent modification of HMG-CoA reductase by phosphorylation
- Glucagon stimulates phosphorylation - INACTIVE
- Insulin stimulates dephosphorylation - ACTIVE
What are the other main 2 methods for cholesterol regulation?
1) High cholesterol activates Acyl-CoA-cholesterol acyl transferase (ACAT): makes cholesterol esters
2) LDL receptor: high levels of cholesterol diminishes transcription of LDL receptor gene, reducing uptake of cholesterol from the blood
What is Familial Hypercholesterolemia?
- Genetic disorder
- Mutations in LDLR gene - defective LDL receptors: very high LDL in blood
- Atherosclerosis develops in childhood
What is Familial HDL deficiency and Tangier disease?
- Low HDL levels from defective ABC1 protein: HDL cannot pick up cholesterol from cells and are rapidly degraded
What group of drugs are used to treat high cholesterol? How do they work? Give specific examples
- Statins
- Competitive inhibitors of the enzyme HMG-CoA reductase in the mevalonate pathway
- Prevent the production of mevalonate from HMG-CoA in cholesterol synthesis
- Lovastatin, Compactin
Give some examples of steroid hormones
- Progesterone
- Testosterone
- Cortisol
- Aldosterone
How are steroid hormones synthesised from cholesterol?
- Requires the removal of the carbons in the sidechain of the cholesterol D-ring
- Also involves oxygenation and hydroxylation
What is oxygenation and hydroxylation?
Oxygenation - addition of oxygen
Hydroxylation - addition of a hydroxyl group (-OH)
Who discovered vitamins in 1929?
Hopkins and Eijkman
Why is it essential to obtain vitamins through diet?
Some cannot be synthesised or are made in such small amounts
Which 4 vitamins are synthesised from isoprene units?
A,D,E and K
What are the only 2 vitamins that can be synthesised in the body?
D and K
What is vitamin A?
An umbrella term describing several components including retinol and retinal
What do deficiencies in Vitamin A cause?
Night blindness
Name some sources of vitamin A
- Liver
- Brocoli
- Sweet potato
- Carrot (Beta-carotene from plants can be converted to vitamin A)
What does vitamin A associate with to produce rhodopsin and where is this found?
Associates with opsin
Found in membrane discs of rod cells
What type of receptor is rhodopsin?
G protein coupled receptor
How is cis-retinal produced from B-carotene?
1) Cleavage of B-carotene to vitamin A1 (retinol)
2) Oxidation of alcohol to aldehyde produces cis-retinal
Describe the formation and breakdown of rhodopsin
1) Cis-retinal forms a covalent bond with a lysine sidechain of opsin
2) Absorbs a ‘green/blue’ photon: light induced isomerisation to trans-retinal results in formation of Metarhodopsin II, and dissociation of opsin and trans-retinal
3) Elicits signalling cascade via interaction with transducin, a heterotrimeric G-protein
Describe the visual cycle
1) Cis-retinal associates with opsin to form rhodopsin
2) Light induced isomerisation to trans-retinal initiates light perception and subsequent dissociation of opsin and pigment
3) Trans retinal (in rod cell) converted to trans-retinol by retinal dehydrogenasewhich moves into retinal epithelial cell
4) Trans-retinol converted to cis-retinal by retinol isomerase, which then moves back into the rod cell ready to bind to another opsin
Describe the biochemical mechanism of sight
1) Light induces conformational change and dissociation
2) Transducin binds GTP (activated)
3) Diminishes cGMP by elevating phosphodiesterase activity
4) Closes Na+ channels, resulting in hyperpolarisation of the photoreceptor cells
5) Diminishes release of glutamate into the synapse: This depolaries the inter-connecting (bipolar) neuron, which stimulates the ganglionic neuron
How do most animals synthesise vitamin C?
From glucose
Why can’t some animals (such as humans, gorillas, guinea pigs) synthesise vitamin C from glucose?
Because they have lost the last enzyme in the pathway
How do humans obtain vitamin C?
Must consume plant matter (e.g orange 53mg/100g)
What are the roles of vitamin c in human?
- Required for collagen synthesis
- Antioxidant
- Metal chelator: facilitates iron absorption from the intestine
What does a lack of vitamin C cause?
Scurvy - disease of connective tissue due to lack of collagen
What are the symptoms of scurvy?
- swollen gums, vulnerable to bleeding
- pain in limbs, particularly legs
- reddish bruise coloured spots
- severe pain and bleeding inside joints
- new wounds may fail to heal
What can occur if scurvy is left untreated?
Can enlarge the heart muscle causing cardiac bleeding, can result in death
What 2 molecules are included in the term vitamin D?
Vitamin D2 (ergocalcifero) and Vitamin D3 (cholecalciferol)
What occurs in the skin to produce vitamin C?
- 7-dehydrocholesterol is converted to cholecalciferol (vitamin D3) using UV light - inactive form
- Vitamin D3 converted to 1,25(OH)2 vitamin D3 - active form
What foods contain vitamin D?
Oily fish, red meat and eggs
What are the 2 major effects of vitamin D?
1) Uptake of calcium in the intestine
2) Release of calcium from bones
What is rickets? What is it caused by?
- Disease in children characterised by impeded growth and deformity of long bones
- Caused by deficiency in vitamin D or/and calcium
- Can be caused by genetic defects as in pseudovitamin D deficiency
What is rickets known as in adults? What are the symptoms?
- Osteomalacia is the milder adult condition
- Diffuse body pains, muscle weakness and fragility of bones
What is the treatment for Rickets?
- Administration of Vitamin D2 or D3
- D2 commercial product via UV irradiation of cryosterol from yeast
What are the 2 vitamin k’s and where do humans get each from?
Vitamin K1 - from green plants
Vitamin K2 - from gut microorganims
Why are newborns in some countries given injections of vitamin k?
- Vitamin K involved in synthesis of blood clotting proteins
- Newborn clotting factors far less than adults; vulnerable to vitamin K deficiency bleeding
- Clotting factors lower in babies due to sterility of gut and under synthesis of precursor proteins
What are the symptoms of vitamin K deficiency?
- Localisation of blood (haematomas)
- Red or purple skin spots (petechiae)
- Stomach pains
- Risk of massive uncontrolled bleeding
- Cartilage calcification
What is Warfarin and what is it used for?
- Main oral anticoagulant used in the UK
- Works by inhibiting an enzyme in the synthesis of coagulation factors from vitamin K - inhibits formation of active prothrombin
- Potent rodenticide
- Valuble drug for treating diseases linked to blood clotting e.g coronary thrombosis (causing heart attack), stroke, deep vein thrombosis
What type of drugs cause an increase demand for vitamin B, C and E?
- Oral contraceptives
- Tobacco
What can large excesses of Vitamin C and Vitamin A cause?
C: gastrointestinal problems
A: foetal malformations (retinoic acid is a signalling molecule that guides development of the embryo)
