Cholesterol biosynthesis Flashcards
explain the stucture of cholesterol:
sterol with an OH group
-weakly amphipathic
what is the single precursor of carbon atoms for cholesterol?
acetate
-every single carbon comes from acetate
what initiates cholesterol synthesis? where else is this seen?
cholesterol synthesis begins with the transport of acetyl coA out of the mitochondria
-this is the same as ketone synthesis
OAA -> citrate -> crosses IMM through citrate transporter (tricarboxylic acid transporter) and OMM through porins -> citrate lyase uses ATP to make acetyl coA and OAA
explain the 4 stages in the overall generation of cholesterol from acetate: where is this most likely occuring? what makes this energy intensive?
1) acetate -> mevalonate (6C)
2) mevalonate -> activated isoprene
3) activated isoprene -> squalene
4) squalene -> cholesterol
very energy intensive due to the breaking and forming of thioester bonds as well as needs for NADPH
-occuring in the liver
what is involved in stage 1 of cholesterol formation to make HMG-coA? what is formed? where does this occur? what enzymes are involed? how does this differ from ketogenesis?
1) linking 2 acetyl coA using a thiolase to generate acetoacetyl-coA (same as ketogenesis)
-HS-coA released
2) using coA, HMG-coA synthase enzyme generates HMG-coA
-HS-coA released
this occurs in the cytosol, in ketogenesis it occurs in the mitochondrial matrix
how many acetyl coA are used in phase 1 of cholesterol synthesis?
3 Acetyl coA
what is involved in genration of mevalonate? what is formed? where does this occur? what enzymes are involed? what is important about this step? how does this step differ from ketone synthesis?
1) conversion of HMG-coA to mevalonate using two NADPH and the HMG-coA reductase
-major regulatory step
-increases un repsonse to insulin
-decreases in response to glucagon
ketone synthesis uses HMG-CoA lyase instead of reductase
-producing acetoacetate instead of mevalonate
how is mevalonate converted into activated isoprene units?
mevalonate kinase transfers phosphate group from ATP to generate phosphomevalonate, which is then converted to isoprene through decarboxylation
in cholesterol synthesis, what happens to phosphomevalonate? what is used? why is this done?
converted into 5-pyrophosphomevalonate
-uses ATP and a kinase
-this makes it a good leaving group
in cholesterol synthesis, after the generation of 5-pyrophosphomevalonate, what occurs? what is used? what is generated?
1) decarboxylation
2) dephosphosphorylation
-uses ATP and releases CO2 and Pi
generates a C5 structure linked to pyrophosphate (there are 2 types, either IPP or DMAPP)
how many ATP are needed to convert mevalonate into 5C pyrophosphate molecules?
3 ATP
how is squalene formed? what drives this reaction? how many activated pyrophosphate molecules are needed to make squalene? why?
both 5C pyrophosphate molecules are linked together (using the energy from the release of PPi) to generate squalene
-uses 1 NADPH
6 pyrophosphate molecules are needed
-3 molecules generat a C15, 2 C15 molecules make squalene
what is involved in the conversion of squalene to cholesterol?
squalene monooxygenase is a regulated enzyme that uses NADPH and O2
-16 NADPH used and 2 NADH generated
how many C16 FA are needed to make cholesterol? explain.
3
what would you expect HMG-CoA to be involved in during the fasted vs fed state?
fasted: involved in ketone synthesis in the matrix
fed: involved in cholesterol synthesis in the cytosol /ER
-HMG-coA reductase is the first enzyme in cholesterol synthesis to be assiciated with the ER
how is HMG-CoA reductase regulated short term and long term?
rate of transcription and degradation is tightly controlled
long term: rate of gene transcription / enzyme degradation
-degradation increases when not in use
short term: covalent modification / statins
is HMG-coA reductase likely to be active phosphorylated?
no
-insulin will dephosphorylate HMG-coA reductase, activating it
how does AMPK impact HMG-coA reductase activity?
it will decrease activity of HMG-coA reuctase
-High AMP:ATP ratio activates AMPK -> phosphorylates HMG-coA reductase and ACC
->turns off pathways that use ATP
compare the difference in AMPK and protein kinase A. How does their response differ?
AMPK responds to energy levels within the cell ([AMP]) and PKA response to blood glucose levels outside of the cell
how does esterification affect cholesterol transport / storage? what enzyme catalyzes this?
esterification makes cholesterol completely hydrophobic to increase ability for transport and storage
-ACAT: takes a fatty acid from coA to cholesterol to generate cholesterol ester
what are lipids and cholesterol transported in?
lipoproteins
what 4 lipoproteins do chylomicrons contain? what do they do?
1) B48: allows us to get chylomicrons out of intestine into lymphatic system (cannot be taken up by LDL receptor)
2) C-II:activates LPL (hydrolizes TAGs to FFA + glycerol)
3) C-III: masks ApoE to inhibit uptake of CM into the liver (CIII falls off only after CM is hydrolized)
4) Apo-E: lipoprotein that can be taken up by the liver (signals formation of CM remnant)
what is the function of apolipoprotein B100? what other lipoprotein has the same function?
B100 is a ligand that allows the LP to be taken up by LDL receptors
-Apo E has the same function
what lipoprotwins have Apo A1? why?
HDL and Chylomicrons
-activates LCAT and interacts with ABC transporter
(LCAT forms cholesterol esters and helps with maturation of HDL particles)
what proportion of TG, Cholesterol and proteins would you expect in each type of lipoprotein?
CM: TG> cholesterol > Pro
VLDL: TG> cholesterol> pro
LDL: cholesterol > pro > TG
HDL: pro ~ cholesterol > TG
explain the exogenous pathway of lipoprotein and lipid transport:
1) TG from diet are incoporated into CM (made in the intestine)
2) CM deliver TG to peripheral tissues (through the action of CII which activates LPL)
3) A CM remnant is formed and CIII falls off
4) CM remnant is taken up by the liver by LDL receptors which respond to ApoE
explain the exogenous pathway of lipoprotein and lipid transport:
1) VLDL takes up cholesterol and TG made in the liver
2) released directly into the bloodstream and VLDL particles exposed to LPL, releasing FFA + glycerol
3) loss of TG and protein through the activity of LPL converts VLDL into IDL
4) IDL particles get converted into LDL particles which are taken up by other tissue to deposit cholesterol
5) LDL can be taken back up to the liver by B100
what happens with a LPL deficiency? What does this result in?
TAGs will not be broken down -> Apo CIII will not be released from the chylomicron -> Apo E will not be exposed -> chylomicrons will not be taken up by the liver and will stay in the bloodstream
-high levels of CM in blood
-high levels of VLDL (can still be taken up by liver)
how is LDL,VLDL, CM remnants and IDL removed from the blood?
1) clathrin coated pits take up particles through binding of receptors (interacts w Apo E / B100)
2) particle in the cell is acted on by lysosomes which release cholesterol, FA and AA
3) increased cholesterol levels inside the cell stimulate ACAT which converts free cholesterol to cholesterol esters
4) high levels of cholesterol at the ER decrease HMG-coA reductase synthesis and LDL receptor synthesis
explain how the LDL recpetors function. explain the function of 3 domains.
there are a number of domains responsible for internilization of ApoE/B100 lipoproteins
1) binding domain
2) domain responsible for pH-dependent conformation change
3) domain for receptor clustering and internalization
how does HDL remove cholesterol from peripheral tissues? what transporter is needed?
Apo A1 interacts with ABCA1/CERP
-uses ATP to move cholesterol from inside the cell to HDL particles
why are we more likely to get plaque formation in humans?
because LDL receptor and ABCA1 (CERP) deficiences are dominant
-only one mutation is needed to cause change so there is a higher chance
what is the purpose of LCAT? how does this differ from ACAT? where are they found? what do they do? what is the fatty acid source for each?
LCAT: enzyme found in the bloodstream that converts cholesterol in HDL to cholesterol esters
-aids in maturation of HDL particles and transport of cholesterol from peripheral tissues to liver
-FA coming from glycerophospholipid (lecithin)
ACAT: cytosolic enzyme that converts cholesterol to cholesterol esters for storage
-prevents cholesterol accumulation in cells
-FA coming from fatty acyl-coA
other than a cholesterol ester, what is produced using LCAT? explain the structure
CE and a lysophospholipid (lysolecithin)
-glycerol backbone, FA at C1, phosphate at C3
why is cholesterol esterified?
converts it to its most hydrophobic form to be sucked into interior of HDL for storage / transport
What 3 proteins are critical for HDL formation?
Apo A1
ABCA1/CERP
LCAT
what is the function of SR? what 2 reasons make this a beneficial receptor for HDL?
Scavenger receptor (SR)
-HDL binds to these receptors to release cholesterol (to liver or steroid hormone producing tissues)
-both tissues which have high needs for cholesterol
1) not subject to down regulation from high cholesterol levels
2) doesnt have to take full HDL disk into the cell to remove cholesterol which allows for better recycling of HDL
-can be removed from blood without LDLr
what is the indirect pathway that allows us to get cholesterol to peripheral tissues?
Cholesterol ester transfer protein (CETP) enzyme takes TAGs from LDL to HDL and takes cholesterol from HDL to LDL
-allows us to redistrubute cholesterol to tissues
what is arteiosclerosis? how/ why does plaque build up? what is the affect of HDL?
thickening of arteries from build up of fat and cholesterol
-causing chronic inflammation
It is a response to chronic minimal injury to endothelium
-Monocytes,lipoproteins,platelets,etc. rush to help with endothelial injury
-macrophages and foam cells come to help and end up worsening the issue
HDL:
-inhibits oxidation
-inhibits inflammation
-decreases activation of endothelium
-decreases coagulation / platelet aggregation
how does insulin impact cholesterol synthesis?
insulin promotes synthesis by activating HMG-CoA reductase
what is the master regulator of cholesterol metabolism? explain the mechanism when cholesterol levels are high vs low.
the sterol regulatory element-binding protein (SREBP) is the master regulator
High cholesterol:
1) binding of cholesterol to SCAP (SCREBP cleavage-activating protein) allows SCAP/SREBP complex to associate with INSIG (insulin-induced gene)
2) association with INSIG prevents it from dissociating to ER membrane
-preventing it from transporting to golgi
Low cholesterol
1) cholesterol doesn’t bind
2) SREBP /SCAP dissociates from complex into the golgi
3) in the golgi SCAP allows S1P to work, which allows S2P to cleave SREBP
4) HLH taken to nucleus and affects transcription genes for:
-HMG-CoA reductase
-gene for LDL receptor
-gene that produces ACAT
why is SREBP the master cholesterol regulator?
Its cleavage by S2P from SCAP allow it to produce HLH which is taken into the nucleus, affecting:
1) HMG-CoA reductase
2) LDLr
3) ACAT producing gene
