Lecture 17 (3-21) Flashcards Preview

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1
Q

Cholesterol Biosynthesis: general early steps

A
  • control at the enzyme level
  • biosynthesis in the cytosol begins with two Claisen condensations
  • first step is a thiolase reaction
  • second step makes HMG-COA (3-hydroxy-3-methylglutaryl-CoA)
  • third step: HMG-CoA reductase - is the rate-limiting step in cholesterol biosynthesis (note: 2 NADPH reactions)
2
Q

Inhibiting Cholesterol Synthesis

A

Statins

  • Statins are cholesterol synthesis inhibitors - why? they hit the rate-limiting step in cholesterol biosynthesis
  • Lovastatin (mevinolin - ‘Mevacor’): is administered as an (inactive) lactone, blocks HMG-CoA reductase
  • In the body (after oral ingestion of it), the lactone is hydrolyzed to mevinolinic acid, competitive (TSA!) inhibitor of the reductase, K1=0.6 nM!
  • Mevinolinic acid is a transition-state analog of the tetrahedral intermediate formed in the HMG-CoA reductase reaction ( A TSA resembles the transition station the substrate molecule)

Other statins (anti cholesterol drugs) - HMG-CoA reductase inhibitors:

  • Lipitor, Zarator, Advicor, Crestor, Lescol, Zocor (Simvastatin), Atorvastatin (Lipitor), Fluvastatin (Lescol), Pravastatin (Pravachol), Cervastatin (Baycol)
  • Baycol muscle pain side effects in 1 in 10,000 people (the name for the condition thought of as Baycol muscle pain is called Rhabdomyolysis) –> Baycol was pulled from the market because of serious muscle problems
3
Q

Alternative anti cholesterol approach (+ an example of it)

A

Bile Acid Sequestrant:

  • sequester the bile acids so cholesterol can’t be absorbed (Welchol, Questran Light, Colestid)
  • Problematic: absorption of other lipids/vitamins!
  • Side effects: constipation, abdominal pain, bloating, vomiting, diarrhea, weight loss, flatulence

Colesevelam Hydrochloride: a bile acid sequestrant - has some GI side effects (because it throws off the process that gets FAs into system)

4
Q

Atherosclerosis: what is it, what does it cause

A
  • “Clogging…” or “hardening of the arteries”
  • Causes myocardial infarction (heart attack), stroke, peripheral vascular disease
  • main cause of death in NA and Europe
  • infiltration of vessel walls with lipids and formation of atherosclerotic plaques
  • Multifactorial: involvement of many genetic/environmental components

The problem:

  • a stable plaque scan cause BLOCKAGE - Myocardial Infarction!
  • unstable plaques lead to THROMBOSIS - stroke
5
Q

Lipid Transport and Lipoproteins: function + types

A
  • Lipoproteins are the carriers of the most lipids in the body
  • unesterified fatty acids bound to albumin/other proteins
  • phospholipids (PL), triacylglycerols (TAGs), cholesterol transported by lipoproteins

Types of lipoproteins:

  • high-density lipoproteins (HDL) – smallest amount of lipid and smallest size
  • low-density lipoproteins (LDL)
  • intermediate-density lipoproteins (IDL)
  • very low-density lipoproteins (VLDL)
  • chylomicrons (lowest protein:lipid ratio but largest size) – largest amount of lipid and largest size
6
Q

Properties of Major Lipoprotein Classes - origin of lipoproteins

A
  • HDL: liver (ER)
  • LDL: liver (synthesized from VLDL)
  • IDL: circulation (remnants from VLDL after FA’s delivered)
  • VLDL: liver (ER)
  • chylomicrons: liver
7
Q

Properties of Major Lipoprotein Classes - function of lipoproteins

A
  • HDL: returns excess cholesterol back to liver
  • LDL (from VLDL): main carrier of cholesterol and cholesterol esters
  • IDL: remnants from VLDL after FA’s delivered
  • VLDL: carry liver-SYNTHESIZED TAG to tissues
  • chylomicrons (Lowest protein:lipid ratio but largest size): carry DIETARY TAG and cholesterol from gut to tissues
8
Q

General Structure of Lipoprotein

A
  • core of mobile TAGs and/or cholesterol ester
  • surface is a PL monolayer where polar head groups face outward - why not a bilyer?
  • —- If it had bilayer, it would have a hydrophobic shell but since this is a shuttle, you want polar heads outward to interact with solvent water (the phospholipids thus shield the hydrophobic lipids inside from the solvent water outside)
  • cholesterol and protein inserted into PL layer
  • Apoproteins: are the proteomic component of lipoprotein
9
Q

Lipoproteins in Circulation: general description of what’s happening + roles of each individual type

A

lipoproteins in circulation are progressively dilapidated/degraded by lipases (specifically Lipoprotein Lipase)
- as this happens, they lose TAG and get smaller (VLDL become IDL become LDL)

  • Chylomicrons’ main task is to carry dietary triglycerides from gut to peripheral tissues
  • VLDLs do same for TAG’s synthesized in the liver (carry lipids from liver)
  • Chylomicrons or VLDL’s anchored by LP lipase
  • LP lipase activated by apoC-II
  • LP lipase hydrolyzes TAGs
  • free FAs taken up by cell (they are unloading fat)
  • What remains? protein-rich REMNANTS!
  • LDL receptor removes LDL from circulation
10
Q

Lipoproteins in Circulation: LDL Receptor + associated condition

A
  • Removes LDL from circulation (Apo B-100 critical), get them into the cells

Familial Hypercholesteremia (FH):

  • genetic mutation in LDLR (LDL receptor)
  • heterozygous of mutant LDLR gene -> premature CVD between 30-40 (incidence 1:500)
  • homozygous: could lead to severe cardiovascular disease in childhood (pretty rare, incidence 1 in a million births)
11
Q

Lipoproteins in Circulation: what happens?

A

In the capillaries of muscle and adipose cells:

  • lipoprotein lipases hydrolyze triglycerides from lipoproteins
  • lipoproteins get smaller, raising their density (correlation with exercise)
  • VLDLs progressively converted to IDL and then LDL (they either return to the liver for reprocessing or are redirected to adipose tissues and adrenal glands)
12
Q

Cholesterol homeostasis: what’s happening + endogenous vs. dietary part

A
  • going through capillaries, unloading cargo (so tissues have fat-based source of energy) - particles built up, shrunk down, recycled, repeated

endogenous fat part: VLDLs
dietary fat part: chylomicrons

13
Q

Hypercholesteremia: causes

A
  • nurture (environmental) and - nature: the receptor - Familial defective apolipoprotein B-100 (mutation of apolipoprotein B that causes hyper cholesterolemia)
14
Q

What happens in atherosclerosis? (early vs. later)

A

“Lesions” and Plaque Formation

Early lesion:

  • ‘fatty streak’ is the first recognizable lesion
  • observed in autopsied youths from 10-14 yo

Intermediate lesion:
- layers of macrophages and smooth muscle cells

Advanced lesion:

  • fibrous plaques
  • covered by dense connective tissue cap with embedded smooth muscle cells and T lymphocytes overlaying lipid core and necrotic debris
15
Q

How are atherosclerotic plaques formed?

A
  • this is NOT just a “high cholesterol” problem
  • “Response to Injury” hypothesis: atherosclerotic plaques develop where vessel wall has been injured
  • source of injury: not entirely clear, but one source is oxidized LDL!!
16
Q

“Response to Injury” and Atherosclerosis: How is LDL oxidized? What’s the response to oxidized LDL?

A

How is LDL oxidized?

  • reactive O2 species (ROS) released by macrophages (and other cells) at the arterial wall
  • O2 radicals attack both protein and lipid components of LDL (LDL rich in polyunsaturated FA extremely susceptible!! - allylic oxidation)

Response to oxLDL:

  • increased adherence of macrophages and T lymphocytes to affected vascular area
  • macrophages migrate between endothelial cells and localize subendothelially
  • due to cholesterol accumulation, macrophages become “foam cells” combine with T cells and smooth muscle cells to become a ‘fatty streak’
17
Q

“Response to Injury” and Atherosclerosis: fatty streak

A

fatty streak:

  • creates environment for platelet adhesion
  • platelets release growth factors and cytokines, etc. (fibrous plaques)
18
Q

Unsaturated and Saturated dietary fats + atherosclerosis (include specific types)

A

Unsaturated (the food fats):

  • palmitoleic, oleic, linoleum, arachidonic, nervonic, etc.
  • create enhanced potential for vessel damage –> fibrous plaque formation

Saturated (the bad fats):

  • Lauric, mystic, palmitic, stearic, arachidic, etc.
  • Not all are bad: Palmitic is considered a ‘bad guy’ and it stimulates cholesterol synthesis (bad) BUT stearic may have a null effect on CVD and may actually be a good guy!

Atherosclerotic plaques: % contribution from diet only ~15% (so enjoy a good juicy burger every now and again!)

19
Q

Amino Acid Biosynthesis

A
  • Plants and microorganisms can make all 20 amino acids and all other needed N metabolites
  • In these organisms, glutamate is the source of N, via transamination (aminotransferase) reactions

Mammals:

  • in a sense we are inferior
  • we can make only 10 of the 20 aas
  • the others are classed as “essential” amino acids and must be obtained in the diet
20
Q

Non-essential amino acids

A
Alanine
Asparagine 
Aspartate
Cysteine
Glutamate
Glutamine
Glycine
Proline
Serine
Tyrosine
21
Q

Essential amino acids (and mg)

A
Arginine: mg unknown 
Histidine: 14
Isoleucine: 19
Leucine: 43
Lysine: 38
Methionine: 19
Phenylalanine: 33
Threonine: 20
Tryptophan: 5
Valine: 24
22
Q

Arg in Mammals

A

Arginine is considered an “essential amino acid”

  • actually semi-essential (derived from the diet, endogenous synthesis, and turnover of proteins)
  • dependent on: the developmental stage, health status
  • preterm infants can’t synthesize arg - adults can
  • surgery or other forms of trauma, sepsis and severe burns put an increased demand on the body for the synthesis of arg
23
Q

His in Mammals

A

Histidine is considered an “essential amino acid”

  • actually semi-essential
  • adults produce enough from other amino acids (in liver) to support the body’s daily needs
  • children obtain histidine through diet
  • essential, especially during infancy, for growth and development
24
Q

semi-essential amino acids

A

Arginine and Histidine

25
Q

Amino Acids: transaminations

A
  • Transaminations (often dependent on glutamate) are key for amino acid synthesis
  • Means of transfer of N between aa and kept acids - aa1 + alpha-veto acid2 –> aa2 + alpha-keto acid1

glutamate + alpha-keep acid ——-(pyridoxal phosphate dependent aminotransferase)—> alpha-KG + alpha-Amino acid

  • Aminotransferases (the enzymes that catalyze transamination reactions) exist for all amino acids except The and Lys.
  • Ex: Glutamate + oxaloacetate ——-glutamate-aspartate aminotransferase——> alpha-KG + Aspartate (an amino acid)

Reminder: PLP (pyridoxal phosphate) catalyzes 7 classes of reactions involving amino acids (reactions with bonds to alpha-carbon of the aa, bonds in side chain)

26
Q

Synthesis of Families of Amino Acids

A
  • all amino acids are grouped into families according to the intermediates from which they are made
  • alpha-Ketoglutarate - glutamate: Glu, Gln, Pro, Arg
  • Aspartate - aspartate: Asp, Asn, Lys, Met, The, Ile
  • Pyruvate - pyruvate: Ala, Val, Leu
  • 3-Phosphoglycerate - 3-phosphoglycerate: Glycolic, Lys, Ser
  • Aromatic - chorismate: Phe, tyr, trp
27
Q

The alpha-Ketoglu Family and the Urea Cycle

A
  • Glu, Gln, Pro, Arg, and sometimes Lys

Glutamate synthesized from:

  • alpha ketoglutarate, histidine, ornithine, arginine, proline, or glutamine
  • note: since biosynthesis, these are reversible

Glutamate is the key precursor to: hangover

  • GABA, Gln –> brain
  • Pro, HO-Pro –> collagen
  • Asp, Asn –> metabolism

Note the importance of ornithine:

  • precursor to Arg
  • intermediate in urea cycle
  • intermediate in Arg degradation

Biosynthesis of Arg gives us an N- recycling tool in the Urea cycle
- Arginine has a guanidino group

28
Q

Amino Transferases (+ 2 examples)

A
  • the most common compounds involved as a donor/acceptor pair in transamination reactions are glutamate and alpha-ketoglutarate (these participate in reactions with many different aminotrasnferases)
  • some clinically-relevant serum aminotransferases: serum glutamate-oxaloacetate-aminotransferase (SGOT) (aka aspartate amino transferase or AST), Alanine Transaminase ALT
29
Q

ALT + clinical application

A

(Alanine Transaminase) - a serum aminotransferase

  • The ALT test detects liver damage but also: viral hepatitis, congestive heart failure, bile duct problems, infectious mononucleosis or myopathy
  • ALT values are usually compared to the levels of alkaline phosphatase (ALP) and aspartate aminotransferase (AST), to help narrow which form of liver disease may be involved
30
Q

Arg Synthesis and the Urea Cycle

A
  • The guanidino group of Arg: N and C come from NH4+, HCO3- (Carbamoyl-P), the alpha-NH2 from Glu and Asp
  • Breakdown of Arg in the urea cycle releases two N’s and one C as urea
  • Important in N-excretion and N-balance (occurs in the livers of terrestrial vertebrates)
  • Urea cycle is linked to TCA by fumarate
31
Q

Urea Cycle - importance, location

A

Important for excreting excess systemic N resulting from excess aa intake (i.e. protein diet)
- consume ~100g protein per day - need to excrete 1 mol of excess N daily

Important because main form is NH3 - toxic to the central nervous system (coma-inducing)

  • Encephalopathy
  • Inhibits excitatory neurotransmitters
  • Probably effective at the K+ pumping level

Confined to the liver (no surprise there!) and involves cytosolic and mitochondrial processes working in concert
- Liver dysfunction certain to have neurological ramifications, e.g., in chronic cirrhosis

32
Q

Urea Cycle Disorders (UCDs): overview

A
  • inborn erros of metabolism
  • A genetic disorder caused by a deficiency of one of the enzymes in the urea cycle
  • Nitrogen accumulates in the form of ammonia, a highly toxic substance, and is not removed from the body resulting in HYPERAMMONEMIA (ammonia reaches the brain through the blood, where it causes irreversible brain damage, coma and/or death)
  • Many cases of urea cycle disorders go undiagnosed and/or infants born with the disorders die without a definitive diagnosis - exact incidence of cases is underestimated
  • Possible that up to 20% of SID syndrome cases may be attributed to an undiagnosed inborn error of metabolism
  • In April 2000, research experts at the Urea Cycle Consensus Conference estimated the incidence of the disorders at 1 in 10,000 births
33
Q

Urea Cycle Disorders: names of specific hyperammonemias

A

Hyperammonemias:

  • Arginosuccinic Aciduria: Arginiosuccinate Lyase Deficiency
  • Hyperargininemia: Arginase Deficiency
  • Citrullinemia: Arginiosuccinate Synthetase (ASS) Deficiency
  • Carbamoyl Phosphate Synthetase-I (CPS-I) Deficiency
  • Ornithine Aminotransferase Deficiency
  • Ornithine Transcarbamylase (OTC) Deficiency
  • N-Acetylglutamate Synthetase Deficiency Deficiency
34
Q

Carbamoyl Phosphate Synthetase I (CPS-I) Deficiency

A

a hyperammonemia (Urea Cycle disorder)

  • doctor found this in a woman who 10 hours after delivery of child became disoriented, agitated –> coma –> seizures
  • died 3 days after delivery
  • found to have CPS I deficiency
  • she had been on little or no meat or dairy products
35
Q

Overview of Nucleotide Metabolism

A
  • sizable portion of the metabolic map
  • the immense importance of these compounds is obvious
  • —– AMP (–> ATP)
  • —– GMP (–> GATP)
  • —– other dNMPs (–> dNTP)
  • First discussion on making ATP/GTP for use as building blocks
36
Q

Nucleotide Biosynthesis

A
  • Nearly all organisms synthesize purines and pyrimidines “de. novo” (important - thesis are the building blocks of genetic code)
  • Many organisms also “salvage” purines and pyrimidines from diet and degradative pathways (ribose can be degraded to generate energy but purine and pyrimidine rings cannot)
  • Nucleotide synthesis pathways are good targets for anti-cancer/antibacterial strategies*****
37
Q

Biosynthesis of Purines: sources of atoms of purine ring

A

(the Frankenstein of Biomolecules)

John Buchanan “traced” the sources of all nine atoms of purine ring:

  • N-1: aspartic acid
  • N-3, N-9: glutamine
  • C-4, C-5, N-7: glycine
  • C-6: Co2
  • C-2, C-8: THF - one carbon units
38
Q

Inosine-5’-P Biosynthesis: overview

A
  • 11 steps
  • the purine ring is built on a ribose-5-P foundation
  • goal: inosine-5’-P biosynthesis (inosine-5’-monophsophate/IMP) which is a purine nucleotide
  • the riobose-5-P is from PPP
39
Q

Inosine-5’-P Biosynthesis: Step 1

A

Ribose-5-P pyrophosphokinase:

  • Ribose-5-P activated by Rib-5-P pyrophosphokinase (this step REGULATED)
  • PRPP: is limiting substance for purine syntheiss
  • But….PRPP is also a branch point - what can you deduce?
  • –PRPP serves additional metabolic needs. therefore, the NEXT reaction (not this one) is actually the committed step in the pathway

(The purine ring is built on a ribose-5-P foundation)

40
Q

Inosine-5’-P Biosynthesis: Step 2

A

GLutamine PRPP amidotransferase:

  • Gln PRPP amidotransferase (KEY in regulation)
  • changes C-1 configuration (alpha) to the beta position (need beta-glycosides)
  • this N becomes N-9
  • G- and A-nucleotides inhibit this step but at distinct guanine- and adenine-specific allosteric sites
  • Note- glutamine-dependence
41
Q

Gln PRPP amidotransferase

A

Site of action for Azaserine
- Antibiotic and anti-tumor agent

Glutamine analog

  • Covalently binds the active site of Gln-dependent enzymes
  • inhibitior/anti-tumor agent
  • anti-fungal agent
42
Q

Inosine-5’-P Biosynthesis: Step 3

A

Glycinamide ribonucl. synthetase:

  • a. Glycine carboxyl activated by -P from ATP
  • b. Amine attacks glycine carboxyl (–> Glycine carboxyl condenses with amine)
43
Q

Inosine-5’-P Biosynthesis: Step 4

A

Glycinamide ribonucl. transformylase:

  • First of two THF-dependent rxns
  • Formyl group of N10-formyl-THF is transferred to free amino group of GAR (—> All atoms for 1st ring present now)
44
Q

Inosine-5’-P Biosynthesis: Step 5

A

FGAM synthase:

  • C-4 carbonyl forms a P-ester (from ATP) and active NH3 attacks C-4 to form imine
  • TRANSAMINATION
  • like reaction 2 - irreversibly inhibited by azaserine
45
Q

Folate: clinical significance, involvement in pathway

A
  • THF (folic acid) dependence in two steps (4 and 10) of Inosine-5’-P Biosynthesis provides susceptibility to folic acid analogs
  • Folic acid may play a role in preventing cancer in selected tissues
  • Diet rich in folic acid, high in methionine and low in alcohol reduce the risk of colon cancer
  • Folic acid is protective against breast cancer
  • Folic acid may also improve cognitive function (slow senility/Alzheimer’s)
46
Q

Inosine-5’-P Biosynthesis: Step 6

A

Steps 6-8: Closing the first ring (carboxylation and attack by aspartate)

Step 6:
Aminoimidizole ribonucleotide synthetase
- closing the ring- similar in some ways to step 5
- ATP activates the formyl group by phosphorylation, facilitating attack by N

47
Q

Inosine-5’-P Biosynthesis: Step 7

A

Steps 6-8: Closing the first ring (carboxylation and attack by aspartate)

Step 7:
Aminoimidizole ribonucleotide carboxylase
- Carboxylation results from CO2 addition at C-4 position

48
Q

Inosine-5’-P Biosynthesis: Step 8

A

Steps 6-8: Closing the first ring (carboxylation and attack by aspartate)

Step 8:
Succinyliaminoimidizole-4-carboxamide synthetase
- Attack by the amino group of aspartate links this amino acid with the carboxyl group

49
Q

Inosine-5’-P Biosynthesis: Step 9

A

Steps 9-11: loss of fumarate, another 1-C unit and the second ring closure

Step 9: Adenylosuccinate lyase
- 4 C’s of Asp removed as fumarate (impt. in muscle)

50
Q

Inosine-5’-P Biosynthesis: Step 10

A

Steps 9-11: loss of fumarate, another 1-C unit and the second ring closure

Step 10: AICAR formylase
- Another 1-C addition catalyzed by THF (second THF-dependent reaction)

51
Q

Inosine-5’-P Biosynthesis: Step 11

A

Steps 9-11: loss of fumarate, another 1-C unit and the second ring closure

Step 11: IMP Synthase
- Amino group attacks formyl group to close the second ring

52
Q

Anaplerotic vs. Cataplerotic

A

Anaplerotic: make TCA intermediates

Cataplerotic: utilize TCA intermediates

53
Q

Inosine-5’-P Biosynthesis: Overview + Analogs

A
  • THF (Folic acid) dependence in two steps (4 & 10) provides susceptibility to folic acid analogs
  • Analogs block purine synthesis (DNA replication, cell growth) by INHIBITING THE SYNTHESIS OF THF!

Sulfonamides: compete with PABA

Methotrexate: binds DHF reductase avidly (irreversible inhibitor)

54
Q

Apoproteins (definition + examples)

A

the proteomic component of lipoprotein

Apo A-1: main protein in HDL, activates LCAT
Apo B-100: main protein in LDL, binds to LDL receptor (largest molecular weight)
Apo C-II: important in composition of chylomicrons and VLDL; acts ages Lipoprotein Lipase (smallest molecular weight)
Apo E: important in Chylomicrons, VLDL, and IDL, allowing the binding of these lipoproteins to the hepatocytes

55
Q

Inosine-5’-P Biosynthesis: why 7 ATP?

A

6 steps use ATP (one each at steps 1.3.5.6.7.8) but accounting lists as 7
- 7 high energy phosphate bonds (7 ATP equivalents) are consumed because alpha-PRPP formation in reaction 1 followed by PPi release in reaction 2 represents the loss of 2 ATP equivalents

56
Q

AST + clinical application

A

an aminotransferase

serum glutamate-oxaloacetate-aminotransferase (SGOT) (aka aspartate amino transferase or AST)

Blood level of AST may be elevated because:

  • liver damage caused by: infection (viral hepatitis or mono), gallbladder disease, toxins (such as alcohol), cancer
  • muscle damage caused by: muscle disease (polymyosin), progressive muscular dystrophy, injury (a fall, auto accident)
  • kidney, heart, or liver injury
  • heart failure
  • kidney failure
  • pancreases inflammation
  • and others