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321 terms

Metabolism, Nutrition & Endocrinology (Part I)

Georgetown SMP '14 Molecular & Cellular Physiology
Adenine & Guanine
Cytosine, Thymine & Uracil
replaces adenine in ATP to make GTP
in DNA
in RNA
Major energy source in cell, but not sole source

**hydrolysis of phosphate bond releases heat → energy**

∙three phosphate groups with phosphoester bond b/w alpha phosphate and adenosine group
∙beta and gamma phosphates attached by phosphoanhydride bonds
→require lots of energy
→when broken, lots of energy released
Harnessing chemical energy
When bonds are broken, energy is released, either in form of heat or in terms of work done by system
Nucleophilic Attack of Phosphate
Nucleophilic attacks help drive reactions in one direction in cell, since enzymes aren't always picky about which direction rxn goes

Nucleophile attacks, leads to intermediate state, phosphate group then transferred and leaving group leaves → energy transferred to nucleophile
**Key: ΔG=ΔH-TΔS, rxn needs to be energetically favorable
∙just because phosphate linkage may not have enough energy to transfer phosphate doesn't mean rxn can't happen → if rxn can absorb enough heat it can happen, entropy helps too
ATP and nucleophilic attack
ATP phosphate can be attacked at any bond by nucleophiles

Possible products: Nucleophile bond to gamma phosphate, bound to gamma and beta phosphates, or bound to alpha phosphate and adenosine
electron carriers of cell

Oxidized: NAD⁺
Reduced: NADH

composed of AMP (lower ring with adenine) plus a nicotinamid base (upper ribose with red ring)

NADP⁺ is same molecule with extra phosphate group added to AMP

both can be reduced, but NAD⁺ reduced in catabolism, NADPH oxidized during anabolism
→done to keep high energy electrons separate

rxn that reduces NAD⁺ to NADH is limited by acidic conditions
Redox Reactions
Most energy is cell comes from high energy electrons being shuffled around so ATP can be more or work can be done, aided by oxidation and reduction
→burning of fuel takes lots of fuel, cells break down into smaller chunks via oxidation
→building of fuels (easier) done w/ reduction rxns

*often a H atom may be referred to as electron b/c it is typically donating its electron to another atom to fill shell (i.e. oxygen) → NAD⁺ to NADH
breakdown of more complex substances into simpler ones with release of energy
synthesis of more complex substances from simpler ones
Catabolism vs. Anabolism
Energy derived from flow of electrons to oxygen
Addition of hydrogen is actually a reduction

new molecule requires a lot of energy to remove hydrogen once added
Main fuel sources
Glucose (difficult to store) & saturated fatty acids (easy to store, preferred)

both can be catabolized for OxPhos, only glucose can be catabolized anaerobically (glycolysis)

both need to become acetyl-CoA to go through citric acid cycle, both will send electrons via NADH and FADH₂ to electron transport chain while acetyl group is completely oxidized
Most likely came from bacterial infection of eukaryote and eventually had symbiotic relationship

have own DNA/ribosomes, act like bacteria

outer membrane is permeable to everything, inner membrane is impermeable to everything
→important b/c electron transport chain proteins are on inner membrane and shuttle H⁺ ions out of matrix and into intermembrane space

matrix is where citric acid cycle takes place
Electron Transport Chain
Complex I → coenzyme Q (ubiquinone) → Complex III → soluble carrier protein cytochrome c → Complex IV → oxygen
* *10 protons total**

Complex II (succinate dehydrogenase) → coenzyme Q → complex III → cytochrome C → complex IV → oxygen
**6 protons total**
Alpha Amino Acids & Alpha Keto Acids
core backbone structure is identical, simple changes between side groups (via amination/deamination) switches b/w 2 forms
Routes of entry to ETC
depends on which electron carrier is used:
∙NADH → electrons to Complex I
∙Succinate → electrons to Complex II
Acceptor Of Reducing Equivalents from NADH in ETC
NADH hands off two electrons to FMN in complex I, makes NAD⁺ and H⁺

w/in complex I, electrons FMNH₂ transfer to iron-sulfur center
Flavin mononucleotide

cofactor that is covalently attached to one of amino acids making up complex I

triple ring structure

accepts 2 electrons to become FMNH₂
→electrons bound to FMN will have lower energy than thouse bound to NADH

when bound to adenylate nucleotide instead of amino acid, considered flavin adenine dinucleotide (FAD)
→both derived from riboflavin (vitamin B₂) while NAD is derived from niacin (vitamin B₃)
Iron-Sulfur Center
Iron is chelated by elemental or cysteine-bound sulfurs

Iron can accept electrons in Fe³⁺ state to become reduced Fe²⁺, can easily donate electrons to return to oxidized state
→electrons from FMNH₂ transfer to Fe-S center

abundance of Fe-S centers in inner membrane of mitochondria is actually what gives dark appearance to meat b/c tissue is slow twitch muscle, which uses fatty acids as major metabolic fuel
→fatty acids broken down by OxPhos in mitochondria
Coenzyme Q
lipid soluble cofactor that contains benzoquinone ring and hydrophobic isoprenoid chain
→side chain responsible for solubility w/in lipid membrane
→benzoquinone reduced upon accepting electrons from coenzyme Q

accepts electrons from Complex I's iron-sulfur center in ETC
Coenzyme Q transfers electrons to complex III in ETC
Complex III pumps out 4 more protons into inner mitochondrial space
Cytochrome C
Free protein in intermembrane space
→all contain heme - hold iron in place
**unlike hemoglobin, iron's role here is to hold electrons, i.e. in complex I

transfers electrons from complex III to complex IV in ETC
Energy from protons pumped by ETC is harnessed for ATP production
As protons are pumped across membrane, local gradient forms
→both a chemical potential (b/c cation accumulation results in low pH) and electrical potential (because of charge)
→potentials are local (not huge pH change occuring throughout entire matrix)

energy expended to create potentials → when proton returns to mitochondrial matrix, energy is released
→energy harnessed by F₁ ATPase to form phosphoanhydride linkage between ADP and inorganic phosphate
F-Type ATPase/ATP Synthase
F₀ domain - integral membrane protein
F₁ domain -catalytically active portion, peripheral membrane protein

Typically seen working in reverse direction → ATP hydrolyzed to provide energy to pump protons
→in inner mitochondrial membrane, however, proton gradient drives ATPase in opposite direction so ATP is produced
→loss of gradient = ATPase reversal, i.e. asphyxiation
Transport of ATP, ADP, Pi in ETC
Adenine nucleotide translocase (antiporter) exchanges ATP for ADP across inner membrane (brings in ADP for ATPase to use)

Phosphate translocase (symporter) uses proton gradient to bring phosphate into matrix from intermembrane space to provide phosphate for ATPase

Once ATP is in intermembrane space, it will freely cross outer membrane to cytosol
Stiochiometry of OxPhos
4 protons to make an ATP

# of protons pumped per pair of electrons:
∙starting from NADH: 10
∙starting from succinate: 6

Net yield: 2.5 ATP per NADH oxidized, 1.5 per succinate
Regulation of OxPhos
provides most of ATP produced in aerobic cells

complete oxidation of glucose (glycolysis, decarboxylation of pyruvate to acetyl CoA, TCA, electron delivery to ETC) yields 30-32 ATP molecules

Rate of respiration and OxPhos limited by level of ADP
→when not limited:brown fat

Hydrolysis of ATP by ATP synthase limited during ischemia by IF₁ inhibitor
ETC and Ischemia
In ischemia, ETC slows because there is no oxygen to accept electrons after complex IV
→would expect gradient to collapse w/o electron flow through ETC but IF₁ inhibitor prevents ATPase from functioning in reverse and hydrolyzing ATP

with loss of OxPhos, anaerobic glycolysis kicks in
→end products: pyruvate and lactic acid (negatively charged), resulting in decreased pH
→metal cations such as K⁺ or Na⁺ will exit mitosol in exchange for H⁺ along inner membrane (once in intermembrane space, freely cross outer membrane to cytosol, positive charge helps neutralize acidic pH)
→as mitosol loses cations, becomes acidic, activating IF₁, which binds to F₁ on ATPase to shut down function
Brown Fat
type of adipose tissue that is rich in mitochondria → dark appearance from iron

uncoupling protein (simple protein channel) present in cell's mitochondria
→allows protons to return back to mitosol but energy si released as heat instead of being captured in ATP
→proton gradient now uncoupled from ATP respiration
→some poisons take advantage of this mechanism by using proton ionophores, surround proton to allow it to cross inner membrane w/o passing through F-Type ATPase, which collapses gradient, energy from gradient released solely as heat
**does not block respiration, only productive use of gradient for ATP synthesis**
Cyanide Poisoning
Cyanide inhibits cytochrome oxidase (Complex IV) step of ETC

binds Fe³⁺ on one of hemes
→once bound, iron can no longer accept electrons, occluding electron flow

inhalation of amyl nitrate or IV infusion of NaNO₂ are treatments
→converts oxyhemoglobin to methemoglobin
→oxygen delivery not compromised b/c this affects only small portion of total hemoglobin
→small portion of methemoglobin provides an Fe³⁺ to cyanide which it will preferably bind to over iron in cytochrome A
process that metabolizes glucose so we can derive energy from it

anaerobic rxn

occurs in cytoplasm
→pyruvate decarboxylation, TCA, ETC all w/in mitochondria

also functions to produce intermediates that are important starting points in anabolic rxns w/in cell
→major mechanism en route to FA production (need pyruvate →acetyl-CoA→citrate)
→many enzymes also involved in gluconeogenesis (generation of glucose through non-carbohydrate sources)

2 phases: preparatory phase & payoff phase
Glucose is major fuel in skeletal muscle
especially in fast twitch

anaerobically converts glucose to pyruvate in cytosol, which is converted to lactic acid

after release from cytosol into blood stream, lactic acid returns to liver to be converted back to glucose

in slow twitch, w/ aerobic conditions, pyruvate instead sent into mitochondria matrix to be converted to acetyl CoA, continue citric acid cycle
Preparatory phase of glycolysis
Glucose is converted to two phosphorylated carbon molecules → glyceraldehyde-3-phosphate and dihydroxyacetone phosphate

ATP beta-gamma phosphate bond hydrolysis is coupled to glycolysis in order to form low energy phosphate compounds (G6P, G3P) → made into high energy phosphate compounds through pathway

in this phase, must be investment of ATP before eventual generation of ATP
Payoff phase of glycolysis
hydrolysis of high-energy phosphate bonds yields new ATP

get 2 ATP we invested back, plus 2 additional ATP

relatively small payoff

also get NADH
Key questions about glycolytic pathway and its role in fuel metabolism
In thermodynamic terms, what drives the forward rxn of glycolysis?

How are enzymes in pathway regulated to control the rate of glycolysis, and what purpose does this serve?

What is the chemical strategy used to harness energy from glucose, and what advantages and limitations does this pose physiologically?
Steps of Glycolysis
1. Hexokinase
2. Phosphohexose Isomerase
3. Phosphofructokinase-1 (PFK1)
4. Aldolase
5. Triose Phosphate Isomerase
6. Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH)
7. Phosphoglycerate Kinase
8. Phosphoglycerate Mutase
9. Enolase
10. Pyruvate Kinase
Glycolysis Net Yield
Glucose + 2 ADP + 2 Pi + 2 NAD⁺ → 2 pyruvate + 2 ATP + 2H⁺ + 2 H₂O
1st Rxn of Glycolysis: Hexokinase
[Glucose + ATP → glucose-6-phosphate + ADP]

first priming rxn

**hexokinase phosphorylates 6th position of glucose, forming G6P, ATP is hydrolyzed**

provides large negative ΔG that drives forward rxn (ΔG = -16.7 kJ/mol)


phosphorylation retains glucose in cell
→only glucose can pass through membrane transporters (GLTS), G6P will be retained in cell for fuel in glycolysis or made into glycogen as storage
2nd rxn of glycolysis: phosphohexose isomerase
[G6P ←→ Fructose-6-Phosphate (F6P)]

not very important step

has positive ΔG, therefore reversible, rxn direction is driven by concentration ratios
→subsequent rxn rapidly removes product, favoring forward rxn
Free Energy Level of Phosphorylated Compounds in Glycolysis
3rd step of glycolysis: PFK1
[F6P + ATP → Fructose-1,6-Bisphosphate (F1,6bP) + ADP]

F6P further phosphorylated by PFK1 to F1,6bP coupled to hydrolysis of beta-gamma phosphate bond of ATP

highly negative ΔG, irreversible

→once F6P gets phosphoryated to F1,6bP it has no choice but to go down glycolytic pathway and be metabolized

B/c this step is checkpoint for glycolytic pathway, activity of PFK1 is heavily regulated
4th step of glycolysis: Aldolase
[F1,6bP ←→ dihydroxyacetone phosphate (DHAP) + glyceraldhehyde-3-phosphate (GAP)]

F1,6bP cleaved into two triose sugars
→only GAP moves forward in glycolysis
→DHAP can be converted to GAP in next rxn, or used as intermediate in FA synthesis

Rxn has very positive ΔG, readily reversible
→main driving force is low product concentration b/c product is rapidly depleted by subsequent rxns
5th step of glycolysis: Triose Phosphate Isomerase

DHAP isomerized to GAP to continue in glycolysis

last step in preparatory phase of glycolysis
6th step of glycolysis: glyceraldehyde-3-phosphate dehydrogenase (GAPDH)
[GAP + Pi + NAD⁺ ←→ 1,3-Bisphosphoglycerate (1,3bPG) + NADH + H⁺]

1st step in payoff phase b/c formation of high energy phosphate compound (1,3bPG)
→acyl phosphate bond very unstable, has high phosphoryl group transfer potential (can be readily transferred to ADP)

Pi comes from inorganic phosphate, NOT ATP

NAD⁺ reduced as it accepts electron GAP
→must be regenerated for rxn to occur again


accumulation of lactic acid and H⁺, decrease in pH from physical exertion makes pathway less effective, favors reverse rxn
7th step of glycolysis: Phosphoglycerate Kinase
[1,3bPG + ADP ←→ 3-phosphoglycerate (3-PG) + ATP]

high energy acyl phosphate bond in 1,3-bPG broken down to form 3-PG
→phosphoryl group transferred to ADP to form ATP


reversible, despite highly negative ΔG
→reversed in gluconeogenesis

enzyme called kinase for adding phosphate group in reverse rxn
enzymes that catalyze transfers of electrons (hydride ions or H atoms)
enzymes that catalyze group-transfer reactions (.ie. methyl group or phosphoryl group)
enzymes that catalyze transfer of functional groups to water
enzymes that catalyze addition of groups to double bonds, or formation of double bonds by group removal
enzymes that catalyze transfer of groups within molecules to yield isomeric forms (from one part of a molecule to another part of same molecule, basically just rearranging)
enzymes that catalyze formation of C-C, C-S, C-O and C-N bonds by condensation reactions coupled to ATP cleavage (energy dependent reaction, usually uses ATP, but sometimes uses other high energy cofactors)
phosphorylate proteins with ATP as phosphate donor
enzymes responsible for removing phosphoryl group
8th step of glycolysis: phosphoglycerate mutase
[3-PG ←→ 2-phosphoglycerate (2-PG)]


phosphate group moved from 3rd carbon to 2nd

reversible, positive ΔG
9th step of glycolysis: enolase
[2-PG ←→ phosphoenolpyruvate (PEP) + H₂O]

forms another high energy phosphate compound, PEP, w/ elimination rxn forming double bond, releasing water
10th step of glycolysis: pyruvate kinase
[PEP + ADP → pyruvate + ATP]

last step

very important b/c high energy phosphate in PEP is transferred to ADP

irreversible, highly negative ΔG
→bypassed in gluconeogenesis
Regeneration of NAD⁺
Aerobic regeneration: occurs in mitochondria, produces high yield ATP, but NADH cannot pass through inner membrane of mitochondria and requires electron carrier for transport, making process slow

Anaerobic regeneration: occurs in cytosol, where pyruvate converted to lactate by lactate dehydrogenase (LDH), NAD⁺ rapidly generated, can be fed back into GAPDH step of glycolysis
→problem is that lactate accumulation can create oxygen debt, acidosis
→lactate reconverted to pyruvate in liver
Regulation of PFK1
if surplus of energy or products in cytosol, want to slow down glycolysis

if we have shortage of energy indicated by high levels, of AMP, ADP, or high concentrations of upstream metabolites (f2,6bP) we want to speed up glycolysis
allosteric stimulator of PFK1

phosphofructokinase-2 (PFK2) constitutively on in muscles, converts small amount of F1,6bP into F2,6bP

F2,6bP is indicator of F1,6bP, indicating PFK1 needs to increase rate
Pentose Phosphate Pathway (PPP)
we are not responsible for names of enzymes/intermediates, but know significant in metabolism and major end products

Oxidative phase & non-oxidative phase

Purpose of pathway:
∙reduction of NADP⁺
∙synthesis of ribose-5-phosphate
synthesized in PPP, used for making nucleotides in DNA/RNA synthesis and cofactors (coenzyme A)
Oxidative Phase of PPP
main event is reduction of NADP⁺ to NADPH while carbon from G6P is oxidized to CO₂

NADPH has important role in anabolic rxns (cholesterol, FA and lipid synthesis), also protects cells by scavenging free radicals (preventing oxidative damage)
→particularly important in RBCs (susceptible to oxygen free radical damage)
→eliminates radicals through glutathione system

Important end product: Ribose-5-phosphate used for making nucleotides in DNA/RNA synthesis and cofactors such as coenzyme A
tripeptide w/ cysteine in middle capable of forming intermolecular disulfide bonds b/w 2 glutathione molecules when oxidized

converted to oxidized form (GSSG) after removal of oxygen free radicals

reduced form GSH has to be regenerated using NADPH

PPP serves as way to regenerate NADPH so we can regenerate reduced form of glutathione
Non-oxidative phase of PPP
involves 5-carbon sugars (ribose-5-phosphate) exchanging carbons, leading to formation/regeneration of G6P

GAP also a product of these rxns
Pyruvate dehydrogenase complex (PDC)
contains 3 enzymes (E1, E2, E3) and requires 5 cofactors (TPP, FAD, NAD⁺, Coenzyme A, lopoic acid)

pyruvate converted to acetyl-Coa by PDC in mitochondria, where it enters TCA

attachment of CoA activates acetyl group, allows it to be transferred to oxaloacetate in TCA
thiamine pyrophosphate, vitamin B₁₂ derivative
E1 in PDC
pyruvate dehydrogenase

pyruvate decarboxylated, acetyl group attaches to TPP in hydroxylethyl form, acetyl group then transferred from TPP to oxidized lipoic acid, which is attached to Lys residue on E2

→transfer reduces lipoic acid, attached group is again acetyl form

** rxn is irreversible because CO₂ release is highly exergonic, CO₂ sequestered, unavailable after release**
E2 in PDC
dihydrolipoyl transacetylase

CoA attacks acetyl group, releasing it from lipoic acid, generating one molecule of acetyl-CoA
E3 in PDC
dihydrolipoyl dehydrogenase

rxn in E2 leaves lipoyllysine in reduced state, needs to be reozidized in order for PDC to continue functioning

→electrons first transferred to FAD to form FADH₂, then to NAD⁺ to form NADH and H⁺
made up of 3'-phosphoadenosine, diphosphate, pantothenic acid and β-mercapto ethylamine

β-mercapto ethylamine contains thiol group (-SH) that forms thioester bond with acetyl group
Regulation of PDC
regulation occurs at E1 b/c step is irreversible

product inhibition occurs when acetyl-CoA and NADH compete with NAD⁺ and CoA for binding sites, which drives rxn backwards
→makes sense b/c end products in high concentrations should shut down forward rxn

phosphorylation of E1 by pyruvate dehydrogenase kinase inactivates enzyme →covalent modification
Citric Acid Cycle
Citrate Is Krebs' Starting Substrate For Making Oxaloacetate

occurs in mitochondrial matrix, electrons harnessed delivered to respiratory chain in inner membrane

Overall rxn: Acetyl-CoA + 3NAD⁺ + FAD + GDP + Pi →2CO₂ + CoA + 3NADH + 3H⁺ + FADH₂ + GTP

circular pathway, rate of acetyl-CoA oxidation determined by amount of intermediates available
→more intermediates, faster it is oxidized
→increase [acetyl-CoA] will never promote net increase in # of intermediates
→pyruvate can promote net increase
TCA intermediates feed many anabolic rxns
Oxaloacetate can be used as starting material for glucose synthesis
→ATP from oxidation of acetyl-CoA through TCA can fuel gluconeogenesis
→if oxaloacetate depleted during glucose synthesis, TCA will slow down, so will production of useable energy

α-ketoacids → oxaloacetate and α-ketoglutarate form carbon skeletons of aspartate and glutamate, respectively
→amination and reduction of either will form corresponding amino acids, will play key role in nitrogen regulation in cell
Steps of TCA
1. citrate synthase
2. aconitase
3. isocitrate dehydrogenase
4. α-ketoglutarate dehydrogenase
5. succinyl-CoA synthetase
6. succinate dehydrogenase
7. fumarase
8. malate dehydrogenase
1st step of TCA: citrate synthase

addition of acetyl-CoA (2C) to oxaloacetate (4C) to create citrate (6C)

large negative ΔG needed to drive rxn forward b/c there is little starting material
2nd step of TCA: aconitase

rearrangement rxn to form isocitrate from citrate
→involves elimination and addition of water to change location of hydroxyl group
3rd step of TCA: isocitrate dehydrognase
irreversible, 1st oxidation step

involves decarboxylation and oxidation to form α-ketoglutarate

has large negative ΔG

either NAD⁺ or NADP⁺ can act as electron acceptor, specific isoform of enzymes determines which is used
→one of two isoforms used, never both
4th step of TCA: α-ketoglutarate dehydrogenase

analogous to PDC
→same cofactors, same structure
→instead of CoA being attached to acetyl group, it is attached to succinyl group → forms succinyl-CoA

large negative ΔG
5th step of TCA: succinyl-CoA synthetase

thioester linkage formed with CoA carries large amount of free energy, harnessed and used to form phosphoanhydride bond in synthesis of GTP

modest ΔG because reaction coupled to GTP synthesis
6th step of TCA: succinate dehydrogenase
readily reversible
→ΔG = 0
succinate dehydrogenase is Complex II in ETC respiratory chain, embedded in mitochondrial inner membrane
→succinate oxidized, electrons transferred to FAD, producing fumarate and FADH₂
rxn (along with next 2 steps) analogous to 1st three steps of β-oxidation of fatty acids
7th step of TCA: fumarase

addition of water to double bond

comparable to 2nd step of β-oxidation of fatty acids
8th step of TCA: malate dehydrogenase

oxidation rxn, producing oxaloacetate, NADH, H⁺

proceeds in forward direction despite very large positive ΔG b/c oxaloacetate is present at such low concentrations and citrate synthase has large negative ΔG

comparable to 3rd step of β-oxidation
Regulation of TCA
logical → high-energy demand stimulates TCA, resting/fed state inhibits

Allosteric regulation and substrate availability are 2 mechanisms that influence rate of TCA on day-to-day basis

cannot increase TCA intermediates through general means such as adding acetyl-COA, specific rxns required

Starving cell (go): low ATP, high ADP/AMP, low NADH, high NAD⁺, low citrate/succinyl CoA, high calcium levels

Satiated cell (stop): high ATP, low ADP/AMP, high NADH, low NAD⁺, high citrate and succinyl-CoA
Regulation of TCA via substrate availability
levels of NAD⁺, oxaloacetate, acetyl-CoA are limiting factors
Regulation of TCA via allosteric regulation
important enzymes: citrate synthase, alpha-ketoglutarate, isocitrate dehydrogenase (also PDC)

ATP inhibits, ADP/AMP stimulates
Pathology associated with TCA
inhibition of PDC will affect every cell
clinical presentations will be associated with most metabolically active tissues: neurons
blocking by limiting thiamin availability (vitamin B₁) causes Wernicke encephalopathy and BeriBeri
→associated with decreased thiamin intake
→Wernicke seen in lots of alcoholics due to poor nutrition, but constant presence of EtOH in CNS doesn't help
PDC deficiency (PDCD) manifests with similar neurological disturbances seen in thiamin deficiency
→patients survive because PDC only links glycolysis, glycolysis can function on its own
Malate-Aspartate cycle
used to oxidize NADH back to NAD⁺, reaches to mytosol where NADH can deal some electrons to OxPhos chain

important in liver, kidney, heart where tissue has steady and constant demand for energy

only reason we do this is b/c inner membrane of mitochondria is highly selective and does not allow oxaloacetate or NADH/NAD⁺ to move freely

conversion b/w malate and oxaloacetate in either direction is mediated by MDH
∙malate→oxaloacetate has high positive ΔG that is only overcome in mytosol due to relative concentrations
∙in cytosol, reverse rxn occurs

relation b/w asparate and oxaloacetate
→Asp aminotransferase mediates shuffling of oxaloacetate into/out of mitochondria
Glycerol-3-P Pathway
for demanding cells (skeletal muscle, neurons) that break down lots of glucose, need faster more efficient way of restoring NAD⁺ levels

can use G3P shuttle to deliver electrons directly from ETC
1. use cytosolic G3P DH to convert DHAP to G3P, oxidizing NADH in process
2. G3P diffuses toward inner mitochondrial membrane and is converted back to DHPA by mitochondrial G3P DH
→during this step electrons delivered directly to coenzyme Q using FADH₂/FAD molecule
→entering ETC at coenzyme Q and skipping proteins 1 and 2 produces less of proton gradient across mitochondrial membrane (2 ATP short as opposed to malate-aspartate shuttle)
Delivery of Fatty Acids to mitochondria
to absorb, must break down TAGs to free fatty acids and glycerol backbone

hydrolytic lipases produced by pancrease are hydrophilic, have tough time reaching hydrophobic fat globules in intestine

gall bladder releases biles salts into small intestine, primarily molecules of cholesterol linked to amino acids, giving them amphipathic qualities, can emulsify fat globules and provide aqueous lipases easy access

once broken down, free fatty acids and glycerol are taken up by small intestine, re-esterified and linked with various apolipoproteins (ApoC II)
→once fat globules assembled they are deemed chylomicrons, which roam around body, dropping off fat wherever apolipoproteins are recognized by receptors on capillary wall until they reach liver
→TAGs must be broken down again to get into some peripheral cells, only to be re-esterified

if liver wants to export fats, does so via VLDL
→deposits fat to extra-hepatic tissues, becomes more dense (LDL)
Mobilizing Fat
in adipocytes, fat stored as TAGs located in globules encased by perilipin
→for hormone sensitive lipase to access TAGs, must get through perilipin coat
→PKA is stimulated to phosphorylate both perilipin and HSL to do so, phosphorylated perilipin will dossociate from globule, activated HSL will acess tag
→→→free FAs released, carried into blood stream by albumin

fat insides adipocytes constantly in flux, cyclical fashion
Free Fatty Acids
Free FAs must be activated with terminal CoA

fatty acyl-CoA synthase responsible for replacing carboxylic head group of FFA with a CoA
→includes priming with AMP before Acyl-CoA added

for activated FFA to gain access to mitcohndria, must be linked with carnitine
→carnitine acyl transferase esterifies FFA with carnitine molecules, gets FFA past outer mitochondrial membrane
Fatty acid oxidation that produces NADH, FADH₂, acetyl-CoA
→NADH, FADH₂ donate electrons to ETC

enzymes that help complete will proceed through cycle that generates pair of electrons and acetyl-CoA for every removal of two carbons

Entire process repeats until we have cleaved 16-carbon fatty acid into 8 acetyl-CoA units
Steps of β-Oxidation
1. Acyl-CoA dehydrogenase
2. Enoyl-CoA dehydrogenase
3. β-Hydroxyacyl-CoA dehydrogenase
4. Thiolase (acyl-CoA acetyltransferase)
Step 1 of β-Oxidation: Acyl-CoA dehydrogenase
similar to succinate dehydrogenase of TCA

involves oxidation of alpha and beta carbons

palmitoyl-CoA will be converted to trans-Δ2-enoyl-CoA, takes place on immer mitochondrial membrane where enzyme is located
→enzymes released harnessed by FAD, which ultimately ferries electrons to coenzyme Q

**acyl-CoA dehydrogenase is also analogous to Complex II in ETC**
Step 2 of β-Oxidation: Enoyl-CoA dehydrogenase
similar to fumerase

hydrate double bond just formed b/w alpha and beta carbons
→beta-carbon receives hydroxyl

trans-Δ2-enoyl-CoA → L-β-hydroxyl-acyl-Coa

sets stage to remove acetyl-CoA, first need to convert β-hydroxy to β-keto
Step 3 in β-Oxidation: β-Hydroxyacyl-CoA-dehydrogenase
similar to MDH

oxidized β-hydroxy carbon into keto, allows us to reduce NAD⁺ into NADH

L-β-hydroxy-acyl-CoA → β-Ketoacyl-CoA
Step 4 in β-Oxidation: Thiolase
aka acyl-CoA acetyltransferase

simple nucleophilic attack by sulfur of CoA-SH on β-carbon, creating new acyl-CoA molecule and generating Acetyl-CoA
Path of Electrons from Acyl-CoA dehydrogenase
Acyl-CoA DH is bound to inner mitochondrial membrane

ETF - electron-transferring flavoprotein
Oxidation of Mono-unsaturated Fatty Acids
β-oxidation will proceed as normal until it encounters double bond in Δ³-Δ² position

to continue, but convert cis-double bond and move its position one carbon over
→enzyme Δ³,Δ²-enoyl-CoA isomerase performs this function, allows us to skip acyl-CoA DH

good example: oleic acid
Oxidation of Odd-Numbered Fatty Acid Chains
only made in plants, but must be able to digest them once ingested

only difference from normal β-oxidation is what we do once we get to last three carbons

ex: Propionyl-CoA → need to add another carbon to split resulting 4-chain chain into last 2 acetyl-CoA molecules

2 steps
Step 1 of Oxidation of Odd-Numbered FA chains: Propionyl-CoA carboylase
addition of carboxyl group converting propionyl group to D-methlmalonyl-CoA

necessary cofactors: HCO₃⁻, ATP, Biotin
Step 2 of Oxidation of Odd-Numbered FA chains: methylmalonyl-CoA epimerase & methylmalonyl-CoA mutase
need L, not D-stereoisomer

epimerase converts starting point into L-methylmalonyl-CoA

from here, need to create linear four carbon chain
→function carried out by mutase, forms succinyl-CoA, requires coenzyme B₁₂ to move carboxyl group
→now ready to product 2 acetyl-CoAs
Benefits and Limitations of Fatty Acid Oxidation
*breakdown of FAs into acetyl-CoA units cannot help generate TCA cycle intermediates*
→one exceptionL propionyl-CoA conversion to succinyl-CoA
→small amount of succinyl-CoA produced during breakdown of odd-numbered FAs can contribute to generation of TCA cycle intermediates

FAs are incredibly great stores of energy
→1 palmitate molecule (16C) can result in 108 ATPS
→hydrophobic, can be stored in great quantities
Ketone Bodies
produced in liver cell mitochondria when we have excess of acetyl-CoA, which only occurs when we have excess β-oxidation

acetoacetate and D-β-hydroxybutyrate are ketone bodies that serve as sources of fuel for extra-hepatic tissues, usually during periods of starvation
→brain can use ketone bodies for 33% of energy needs during starvation, reducing need for glucose

acetone also ketone body but not for fuel, produced in small amounts, exhaled

will be released by liver, distributed to various extra-hepatic tissues that have cells with mitochondria capable of converting them back to acetyl-CoA units
→acetoacetate may also be converted to acetone via an enzyme
→rxn may also occur via spontaneous decarboxylation w/o enzyme
→acetone produced because of dangerous excess of acetyl-CoA and other ketone bodies, not as fuel, wants to be exhaled to detox blood
synthesized from acetyl-CoA produced by β-oxidation

begins with condensation of 2 acetyl-CoA catalyzed by thiolase (reversal of last step in β-oxidation)

3 steps
Step 1 of Ketogenesis: Thiolase
reverse of last step in β-oxidation

combine 2 acetyl-CoA units to produce acetyoacetyl-CoA

one CoA-SH released too
Step 2 of Ketogenesis: HMG-CoA synthase
condensation rxn combining acetoacetyl-CoA with another acetyl-CoA

rxn requires water, releases another CoA-SH group

product is β-hydroxy-β-methylglutaryl-Coa (aka HMG-CoA)
→also produced during cholesterol synthesis but happens in smooth ER, in ketogenesis forms in mitochondrial matrix
Step 3 of Ketogenesis: HMG-CoA LYase
HMG-CoA lyase cleaves acetyl-CoA unit producing acetoacetate
→acetyl-CoA added in previous step just as primer for this step
Formation of D-β-hydroxybutyrate
acetoacetate is 1st ketone body, but predominant form in blood is reduced version, D-β-hydroxybutyrate
→conversion carried out by enzyme D-β-hydroxybutyrate dehydrogenase, requires oxidation of NADH
Ketone Body Breakdown
once these emergency energy compounds ahave been distributed to mitochondria of extra-hepatic tissues, two steps needed to convert them back to acetyl-CoA
→forst, any D-β-hydroxybutrate must be oxidized back to acetoacetate by D-β-hydroxybutyrate dehydrogenase, requires NAD⁺

*under starvation conditions, FAs are broken down to acetyl-CoA units, but w/o glucose we cannot generate TCA cycle intermediates needed to metabolize excess acetyl-CoA → means we can't actually accelerate rates of acetyl-CoA use and therefore our liver converts them to ketone bodies*
Step 1 of Ketone Body Breakdown: β-ketoacyl-CoA transferase
converts ketone body into acetoacetyl-CoA

uses succinyl-CoA to attach CoA-S- to product

*β-ketoacyl-CoA transferase not present in brain under normal conditions but is activated during starvation to allow use of ketone bodies*

not catalyzed in liver
Step 2 of ketone body breakdown: Thiolase
thiolase splits 4-carbon compounds into 2 acetyl-CoA units

also adds another CoA-S
Fatty Acid Synthesis
site of synthesis: cytoplasm

starting material for synthesis: acetyl-CoA

source of acetyl-CoA units: mitochondria

problem: acetyl-CoA cannot be transported across mitochondrial inner membrane
Citrate shuttle
site of FA metabolism is cytoplasm but starting compound for FA synthesis is acetyl-CoA, which comes from mitochondria

acetyl-CoA cannot cross inner mitochondrial membrane, so attached to carrier → citrate

acetyl-CoA units in matrix attached to oxaloacetate by enzyme citrate synthase (from TCA), citrate can then move through transporter and into cytosol

in cytosol, citrate lyase frees acetyl-CoA unit in energy-dependent fashion using ATP

*REMEMBER - CoA-SH removed by synthase, added by lyase*

how do we get NADPH here:
→oxaloacetate converted to malate via MDH and then malate is decarboxylated to pyruvate by malic enzyme
→malic enzyme delivers reducing equivalents to NADP⁺ forming NADPH (also produced in PPP)
Step 1 in FA synthesis: Acetyl-CoA carboxylase
Acetyl-CoA carboxylase enzyme will until produce malonyl-CoA, which inhibits CAT1

know that we are using bicarbonate to attach carboxyl group to an acetyl-CoA
→often utilizes biotin as cofactor
→specifics not important
Step 2 in FA synthesis: Fatty Acid Synthase
conglomerate of enzymes in one polypeptide that helps assemble FAs

acyl carrier protein (ACP) is in center w/position that can attach malonyl-CoA
→before attached, want to attach acetyl-CoA to KS subunit of FA synthase

Condensation: once synthase charged with malonyl and acetyl group, have nucleophilic attack by malonyl group on acetyl group and decarboxylation

Reduction: now 4-carbon chain attached to ACP, NADPH used to reduce β-keto group (opposite of β-oxidation)

Dehydration: loss of water to form enoyl group

Reduction: NADPH used to reduce enoyl group to acyl group, results in acyl chain lengthened by 2 carbons, longer chain then transferred to KS subunit and process starts over again
Storage lipids

Membrane lipids


Glycerol backbone

2 FA side chains

Phosphoester that can be further modified with different groups
Ester-linked glycerophospholipids
Plasmalogen & Platelet-activating factor
ester-linked glycerophospholipid
→ether-linked alkene at position 1, resistant to phospholipases

don't know what function is

abundant in cardiac tissue

most common group is choline

elevated levels seen in metastatic cancers (possible biomarker?)
Platelet-activating factor
ester-linked glycerophospholipid
→ether-linked alkyl group at position 1 AND acetyl ester at position 2

slightly more soluble than plasmalogen → able to circulate in blood as molecular signal

when bound to receptors, triggers events → mediates release of granules from platelets, i.e. serotonin → causes platelet aggregation

inactivated by cleavage of acetyl ester
sphingosine backbone

FA at position 2

various groups at position 3
Glycerophospholipid Synthesis
Glycerol-3-phosphate precursor for TAGs and glycerophospholipids
→synthesis of TAGs and glycerophospholipids share common origin

Phospholipids are result of condensation of phosphoric acid with 2 alcohols

Two strategies for head group attachment: activated diacylglycerol & activated head groups

Eukaryotic synthesis of anionic phospholipids PG, cardiolipin and PI is similar to that of bacteria via CDP-diacylglycerol
Activated diacylglycerol vs. Activated head group in glycerophospholipid synthesis
Activated diacylglycerol:
∙energy activated form of diacylglycerol with CDP, can react with head group
∙bacteria employ this strategy
→can be anionic, zwitterionic, pathway changes

Activated head group:
∙energy activated hydroxyl group on head group with CDP to react with diacylglycerol
involves expansion and contraction of lung alveoli

lungs have surfactant on alveoli
→difficult to separate alveoli w/o surfatant → force must be greater to overcome surface tension
→surfactant made up of mainly DPPC (Dipalmitoylphosphatidylcholine) and unsaturated PC (phostphatidylcholine), also have phosphatidylglycerol, cholesterol, FAs, proteins and other lipids
Fetal Lung Surfactant Production
L/S ratio is usually >2.0

in premature birth, 40% chance of RDS (L/S ratio is 1.5-1.9)

if L/S <1.5, 70% RDS
Sphingolipid biosynthesis
4 stages:
∙synthesis of sphinganine, an 18-carbon amino alcohol
∙attachment of fatty acid via amide bond
∙desaturation of alkane chain of acylsphinganine (results in ceramide)
∙head group attachment
→picture deceiving, because phosphocholine is attached here, not just head group

previously thought that sphingomyelin was synthesized from N-acylsphingosine and CDP-choline, now known to be minor pathway

most prevalent acyl groups in SM are 16:0 and 18:0

synthesis occurs in Golgi, unlike ceramide and other phospholipids that are synthesized in sER
Cerebroside Biosynthesis
begins with ceramide, one sugar added

glucocerebroside - minor lipid in non-neural tissue, however, is precursor of golobosides and gangliosides

galactocerebroside - common lipid in brain, often referred to as cerebroside

both are linked by β-linkage at position 1 of hexose
Sulfatide Biosynthesis
sulfatides = 15% of brain white matter lipids

PAPS (3'-phosphoadenosine-5'-phosphosulfate) serves as source for activated sulfate group

sulfate transferred almost exclusively to 3-OH of galactose

w/ addition of sulfate, monosaccharide has negative charge, so sulfatides are not neutral sphingolipids
Ganglioside and Globoside Biosynthesis
addition of more sugars that cerebrosides

additions carried out by series of glycosyl transferases (in Golgi)

addition of 2nd NANA to Gm₃ leads to Gd series

addition of GlcNAc to lacosyl ceramide bore NANA leads to Ga₂

overall, more than 60 known gangliosides

when one or more sugars added are sialic acid, generating gangliosides
→large, diverse class of sphingolipids
→allows different sphingolipids to serve as molecular tags or markers
Simple Ganglioside Nomenclature
G = ganglioside
→M = 1 sialic acid
→D = 2
→T = 3
→Q = 4
→also have other letters based on branching

# = 5- moieties (excluding SA)
Regulation of Sphingolipid Synthesis & Degradation
carefully controlled

normally, constant turnover of sphingolipids, which occurs in lysosomes

most often in glycolipid storage diseases, biosynthetic rate remains normal but mutation exists in structural gene for hydrolase
∙impaired degradation of sphingolipids results in accumulation of membrane lipids = severe clincal mutations
→accumulation of membrane lipids in lysosome damages lysosome functions
∙must be able to degrade sequentially just as synthesized
Lipid storage diseases
Generalized gangliosidosis

Tay-Sachs disease

Sandhoff's disease

Fabry's disease

Gaucher's disease

Niemann-Pick disease
Symptoms of lipid storage diseases
→enlargement of liver, spleen


corneal opacity
→classic sign

macular spots

ataxia, dystonia, hypotonia
→due to effect on neuronal tissue

Lipid Functions
vast bulk of cellular lipids serve as bilayer structural components

can serve as modulators of activity of both soluble and membrane-bound proteins
→especially acidic phospholipids

phosphatidylionsitol serves as part of tether of GPI-anchored membrane proteins

ether-linked glycerophospholipids, such as platelet-activating factor, can act as molecular signals

derivatives of phosphatidylinositol act as intracellular second messengers in signal transduction pathways involved with many aspects of cellular regulation

FA metabolites, the eicosanoids, act as potent, short-lived hormone-like molecules
Phosphatidylinosital derivatives serve as intracellular signals
generates diacylglycerol, which activates protein kinase C, leads to regulation of other enzymes by protein phosphorylation

head group leads to release of intracellular calcium, leads to regulation of other enzymes by calcium
Eicosanoids are derived from arachidonic acid released from phospholipids
phospholipids → arachidonate → prostaglandins, thromboxanes, leukotrienes, lipoxins, hepoxilins, epi-llipoxins

arachidonic acid has 4 unsaturated bonds on it
→gives rises to 2 and 4 series
→if it had 3 unsaturated bonds, it would give rise to 1 and 3 series
→5 unsaturated bonds → 3 and 5 series
Linoleate & α-linoleate are essential dietary fatty acids for mammals
elongation steps, carried out in sER and mitochondria, involve addition of acetyl groups from coenzyme A

only plants have ability to desaturate linoleate to α-linoleate, humans desaturate to γ-linoleate
α-linoleate vs. γ-linoleate
α-linoleate is ω-3 fatty acid
→has unsaturated bond that is only 3 positions in from ω-carbon
→flax seed, plants, seafood (animals that eat lots of it) are rich in it

γ-linoleate is ω-6 fatty acid
→has unsaturated bond that is 6 positions in from ω-carbon

study showed that ω-3 FA reduces risk of sudden cardiac death, also lowers TAG levels
Desaturation of palmitate and stearate
Desaturation of palmitate (16:0) and stearate (18:0) is carried out by fatty acyl-CoA desaturase is mixed-function oxidase, w/ both acyl chain and NADPH being oxidized
Plant acyl desaturases
Located in ER, convert oleate (by far most often at sn₂ position) of PC into linoleate and linolenate w/ sequential introduction of unsaturated bonds at carbons 12 and 15
desaturases, which may also act upon free FAs, are located in chloroplasts
Phospholipases cleave glycerophospholipids at different sites
named for where they cleave
Phospholipase A
in response to hormonal and other signals, releases arachidonate from sn₂ position of glycerophospholipids

glucocorticoids have inflammatory effects by blocking this A₂ activity
Arachidonate conversion
In sER, arachidonate convered to PGH₂ by bifunctional enzyme, cyclooxygenase (COX)

conversion is 2 step process
1. molecular oxygen is introduced to produce PCG₂
2. peroxidase activity produces PGH₂

PGH₂ is precursor for production of prostaglandins and thromboxanes

action of COX blocked by aspirin and other NSAIDS
non steroidal anti-inflammatory drug
Aspirin as NSAID
irreversible inhibition of COX
Model of Membrane-associated COX
wanted to see if there was drug that blocked COX2 but not COX1

COX 1 - involved with normal housekeeping functions, has isoleucine

COX2 - more for inflammation, pain, fever, has valine instead of isoleucine
→targeted with molecule that can fit by Val but not Ile
Second Generation of NSAIDS that can target COX2 but not COX1

COX Pathway gives rise to prostaglandins and thromboxanes
Thromboxane has oxane structure, originally found in thrombocytes
Prophylactic baby aspirin
goal is to prevent clotting when we don't want it

prostacyclin → saying don't aggregate
thromboxane → saying aggregate

w/ thrombosis, balance is thrown out of whack

baby aspirin inhibits step that is common to both
→binds irreversibly, bot COX can recover by making new COX
→vascular endothelial cells can recover, platelets cannot, prostacyclin wins
Linear pathway from arachidonic leukotrines
instead of COX acting on it, lipoxygenases act on arachidonate
Leukotrine biosynthesis begins with action of 5-lipoxygenase
LTB₄ produces inflammation, implicated in chronic inflammation associated with atherosclerosis

LTC₄, LTD₄, and LTE₄ (cysteinyl-leukotrines) are important mediators of immune-mediated inflammatory rxns of anaphylaxis

treatments for asthma include antileukotriene agents that inhibit 5-lipoxygenase or binding of activator protein
Lipoxin biosynthesis begins with action of 15-lipoxygenase
lipoxins having opposing action to LTC₄, inhibit bronchial spasms, have anti-inflammatory properties
New class of lipoxins: epi-lipoxins
differ in stereochemistry

synthesis triggered by aspirin

also have potent anti-inflammatory actions, further explaining efficacy of aspirin
Newest Players: Resolvins and Protectins
potent anti-inflammatory action

derived from 20:5 and 22:6 ω-3 FAs by pathway involving aspirin acetylated COX2 followed by 5-lipoxygenase
→in absence of aspirin-triggered COX2, EPA and DHA are acted on by 15-lipoxygenase to generate other anti-inflammatory agents

has been suggested that ω-3 FAs, w/ aspirin, may reduce clinical symptoms of several disease states, including inflammatory disorders (arthritis), cardiovascular disease, asthma, and certain cancers
functions directly as NT
→acts to block impulses traveling in spinal cord to stimulate skeletal muscle

conjugated to cholesterol derivative to form glycocholic acid (bile salt)

incorporated directly during de novo synthesis of purines

condenses with succinyl-CoA in first step of heme biosynthesis
Neurotransmitters (simplistic def.)
molecule that is synthesized and stored in synaptic vesicles in neural cells

release is triggered by action potential

bound and recognized by target cell

activity can be regulated
Porphyrin biosynthesis
glycine can lead to heme w/in porphyrin molecule
along with glutamine, predominant circulating AA

transports amino groups from muscle to liver (transamination from muscle pyruvate) in process called glucose-alanine cycle
→takes amine groups to liver to be excrete in urea
source of 1-carbon fragments (folate derivatives) used in biosynthesis
side chain gets added on to folate derivatives
→serves as donor in many rxns
transporter of amino groups

serves as form of activated ammonium ion
→source of NH₄⁺ in kidney

amino group donor in purine biosynthesis

amino group donor in biosynthesis of amino sugars
Amino Sugar Biosynthesis
sugars that have amino groups of saccharide

key rxn:
F6P + glutamine → glucosamine-6-P + glutamate

all other amino sugars derived from glucosamine-6-P
neurotransmitter → primary NT in CNS
→acts on both ion channels and G-coupled receptors

source of γ-amino butyrate (GABA) via decarboxylation of α-carboxyl
→GABA is major inhibitory transmitter in brain
→GABA receptor is target of benzodiazepines/barbiturates
→synthesis uses cofactor (PLP)
→→→decarboxylation step

Participant in transaminations → source of amino groups for most other amino acids

Reactant in ammonia fixation, which results in production of glutamine
Ammonia Fixation
glutamate dehydrogenase

right to left pathway is minor pathway for glutamate synthesis and ammonium fixation
activated by rxn w/ ATP to form S-adenosyl methionine (SAM)
→source of methl groups for most methylation rxns

SAM can be decarboxylated to leave propylamine reisude attached to sulfur
→precursor of spermine and spermidine (polyamines)
multiple amine groups
immediate precursors of urea production via action of arginase in urea cycle

source of NO, a 2nd messenger
NO Synthesis
cofactors include flavin, tetrahydrobiopterin

NO synthase found in many cell types
→macrophages, vascular endothelial cells, neurons, hepatocytes, etc

NO is short lived compound that acts near site of synthesis

activates guanylyl cyclase production of cGMP in target cells
Glycine, arginine and methionine all contribute to synthesis of creatine and phosphocreatine
Create and phosphocreatine are components of an energy buffering (ATP regenerating system) in skeletal muscle cells
decarboxylation produces histamine

major stimulant of acid release in stomach and in systemic rxns to allergens
Phenylalanine and Tyrosine
under normal conditions, Phe is dietary essential amino acid, but Tyr is not since it is produced from Phe

Tyr is precursor of several important moecules
→dopa (dihydroxyphenylalanine), dopamine, epinephrine, norepinephrine

Tyr is precursor of thyroxine, T3

melanin is derived from oxidation of dopa, catalyzed by Tyr
→lack of enzyme: albinism
Biopterin Cofactor
provides electrons for reduction of oxygen in hydroxylation of phenylalanine
→regeneration requires specific reductase

deficiency of phenylalanine hydroxylase (phenylketonuria, PKU) is usually screened for at birth (blood sample measuring for elevated Phe levels)
→controllable by specialized diet with limited Phe, Tyr supplementation
→less required by puberty

deficiency of biopterin reductase is more severe, cannot be controlled by dietary means
intermediate in synthesis and epinephrine/norepinephrine as well as NT involved in control of voluntary movement (parkinson's)

formed by decarboxylation of dopa

both epi and norepi are NTs and signaling molecules
→fight-or-flight, sympathic neres
Thyroid Hormones
active form is tri-iodothryonine (T3)
→T4 is thyroxine

formed by iodination and condensation of tyrosine residues in thyroglobulin followed by proteolysis of portein and hormone release
hydroxylation (5th-position) followed by decarboxylation yields serotonin (NT)
→hydroxylation step requires biopterin cofactor
→decarboxylation step requires PLP cofactor

some Trp can be degraded to nicotinic acid (precursor for NAD⁺) but this is usually insufficienct to eliminate dietary need
Serotonin Biosynthesis
Glycogen Storage Diseases
Type Ia, von Gierke's disease

Andersen's disease

McArdle's disease
Type Ia, von Gierke's disease
Glucose-6-phosphatase deficiency

results in hypoglycemia, lack of glycogenolysis induced by epinephrine and glucagon

Hormones tend to increase output of glucose from liver but in disease liver does not recognize these hormones
Andersen's disease
Defect in glycogen branching enzyme

causes cirrhosis and abnormal glycogen, diminished hyperglycemic response to epinephrine
McArdle's disease
muscle glycogen phosphorylase deficiency

high muscle glycogen, reduction in blood lactate and pyruvate after exercise, no post-exercise drop in pH, normal hyperglycemic response to epinephrine
Metabolic Fuels, Advantages & Disadvantages
Glucose & Carbohydrates
→need minimal level for brain or animal will die
→can cross blood/brain barrier very quickly

Fatty Acids & TAGs
→Very hydrophobic, will be stored in compact form in adipose tissue
→can only be catabolized aerobically
→cannot cross blood/brain varrier quickly enough to be viable fuel

Ketone bodies
→can only be catabolized aerobically
→can also cross blood/brain barrier very quickly, under starvation conditions ~33% of energy
→Problem: negatively charged, causes pH issues

Amino acids & protein
→play key role in providing carbon units necessary for metabolic functions
Relevant properties of glucose
Highly polar

Attract water to blood
→don't want too much glucose despite the fact that we need baseline level

B/c of -OH groups, glucose is very reactive
→can be problem because it can become active nucleophile and attack compounds like collagen (making it very brittle)
Structure of glycogen
Polymers of glucose where molecules are linked together with 1-4 bonds
→counts as single molecule, osmotic pressure goes down as compared to glucose

Branch points
→i.e. 1→6 linkages

Fuel storage
→maximally 1-2% in muscle, 10% in liver
Role of branching

amylopectin branched every 24-30 residues

Glycogen branched every 8-10 residues
→more compact, takes up less space


Non-reducing end vs. reducing end
→non-reducing end is reactive end → branching increases # of non-reducing ends, allows for explosive increase in mobilization
Glycogen as storage form
∙rapid mobilization of glucose
∙glucose metabolized aerobically and anaerobically
∙glucose units released can directly maintain essential blood levels of glucose

∙hygroscopic property, binding 3-4 times its weight in water, limits storage
∙short term storage (liver depleted of glycogen after 12 hour fasting)
Glycogen Mobilization
Enzyme that mobilizes is called glycogen phosphorylase

wants to break down 1-4 glycositic linkage b/c it is high energy, makes use of energy to phosphorylate one unit of glucose, making glucose-1-phosphate and a new non-reducing end
Interconversion between G6P and G1P
we want glucose-6-phosphate, not glucose-1-phosphate

use phosphoglucomutase

by doing this, we can bypass hexokinase step when entering glycolitic pathway
Glycogen breakdown near branch point
PROBLEM: glycogen phosphorylase cannot hydrolyze glucose units 4 residues or less from branch point, limits break down

transferase debranches enzyme by breaking down 1-4 bond near branch and attaches to chain of 1-4 linkages
→1-6 linkage has less energy from 1-4 linkage
→breaking 1-6 linkage would not provide enough energy to create new 1-4 linkage

glucosidase debraches by removing 1-6 bond
Glucose-6-Phosphatase and Release of Glucose into Bloodstream
**Liver is doing #1 metabolic priority: maintaining blood levels of glucose**
→one easy way to do so is to mobilize glucose to glycogen
→→→generating glucose-6-phosphate

glucose-6-phosphate cannot exit cell, only unphosphorylated glucose can leave
→must dephosphorylate to allow glucose to enter blood stream

Glucose-6-phosphatase (in ER) hydrolyzes phosphate off, allows glucose to leave
Formation of UDP-Glucose
eventually builds glycogen

glucose-1-phosphate must be activated so that nucleophilic substitution is possible to form 1-4 or 1-6 glycositic linkage
→activated by being attached to UDP at 1-position, forms UDP-glucose

enzyme responsible for UDP-glucose is pyrophosphorylase
Glycogen Synthase
cannot start glycogen molecule de novo, needs primer

enzyme involved in glycogen synthesis

combines UDP-glucose with non-reducing end of glycogen chain with n residues (n > 4), UDP cofactor

forms new linkage between 1 carbon of UDP-glucose and 4-carbon of glycogen
Primes glycogen synthesis

first 8 glucose units attached to glycogenin added autocatalytically

Glycogenin primers gives rise to glycogen particle
Glycogen Branching Enzyme
simple mechanism

breaks 1-4 linkage and moves to make new 1-6 bond

not as complex as debranching enzyme mechanism because 1-4 bond has more energy that 1-6 so can easily form new bond, mildly exothermic
Regulation of Glycogen Phosphorylase
Active form (a) is phosphorylated, less active in dephosphorylated state (b)
→in process of physical exertion, becomes more active

Glucagon, calcium and epinephrine are activators of forward rxn, active form of glycogen phosphorylase
Phosphorylase Kinase is Active in Phosphorylated Form
Inactive phosphorylase kinase b is phosphorylated to active form, phosphorylase kinase a, by protein kinase A, a cAMP dependent kinase (hormonal regulation)

dephosphorylation of both phosphorylase and phosphorylase kinase A is catalyzed by phosphoprotein phosphatase-1
Glycogen Synthase Regulation
active in unphosphorylated state (a), inactive when phosphorylated (b)

phosphorylation by GSK-3, protein kinase A, and glycogen phosphorylase kinase

dephosphorylation by phosphoprotein phosphatase 1

P favors glycogen breakdown

want active if lots of glucose available that isn't being used

Glucagon/epinephrine tell us that we need glucose in blood, promote release of glucose from glycogen in muscle/liver, so we do not want activated glycogen synthase
Fuel Metabolism in Individual Organs
Interaction b/w muscle, adipose and liver very important

RBCs can only derive enrgy from glycolysis, must be anaerobic
→glucose is obligate metabolic fuel

Brain usually uses glucose for 100% of energy
→constitutes need for 1st metabolic priority to be maintaining blood glucose

Skeletal muscle will take up glucose, store a lot as glycogen when excess glucose available
→can catabolize aerobically or anaerobically

Liver - glucose is important for synthesis for fat, determining whether fat is stored
∙most of FAs in adipose tissues comes from fat in liver
∙Glycerol moeity of fat must come from dihydroxyacetone phosphate
→most abundant source comes from glucose
→if glucose not available, still need to generate dihydroxyacetate but will come from carbon skeletons and will be in much lowers concentrations
∙Major function is glucose homeostasis
→pyruvate -- citrate used to make FAs, fat in liver delivered by LDLs to other tissues
Relative Activities of Hexokinase and Glucokinase
Hexokinase I - found in skeletal muscle
→reaches muscle activity very very wuickly at low [ ]'s

Hexokinase IV (glucokinase) - found in liver
→low activity at low [blood glucose], but as [ ] ↑'s, activity ↑'s

pathways such as glycolysis have different purpose in liver than skeletal muscle
∙skeletal muscle - immediately used
∙in live - role is to synthesize glucose and release into blood, activity is lower so that molecules just synthesized are not immediately used when [glucose] already low
Cholesterol Biosynthesis
anabolic process, many different starting points can lead to cholesterol
→all have acetyl-CoA in common

cholesterol cannot be broken down in body, can only be removed by getting rid of steroids or bile acids (excretion) created from cholesterol
Cholesterol structure
27 carbons, all from acetyl-CoA
→important because cholesterol builds steroids, activity of steroids can be determined by # of carbons

4 carbon rings (1 5-carbon rings)

8-carbon side chain at C-17

composed os isoprene (5 carbon) units

3-hydroxy is only oxygen (very hydrophobic)
→so hydrophobic won't be dissolved in water, can only be transported by binding to hydrophilic compounds (proteins)

double bond b/w C-5 and C-6 (delta 5)
Many Compounds Contain Isoprene Units
Cholesterol is made from isoprenes, then makes steroid hormones, bile acids and vitamin D

other compounds:
∙vitamins A, E, K
∙plant hormones
∙quinone electron carriers
Overview of Cholesterol Biosynthesis
acetate → mevalonate → ispentenyl pyrophosphate → squalene → cholesterol
Acetyl-CoA Utilization
2 acetyl-CoAs fuse to form acetoacetyl-CoA (thiolase)

third acetyl-CoA is added to form HMG-CoA (HMG-CoA synthase)
Formation of Mevalonate is inhibited by Statins
HMG-CoA → melavonate (HMG-CoA reductase)
∙carboxy is reduced to hydroxy
∙electrons from 2 NADPH and release of CoA give mevalonate
∙inhibited by statins (can be used to control cholesterol levels, inhibit HMG-CoA reductase)
can get different statins by changing R1 and R2 groups
Activated Isoprenes are made by sequential phosphorylation and a decarboxylation
isoprenes are bicarbon units

2 sequential phosphorylations on C5 form pyrophosphase

phosphorylation at C3 followed by immediate decarboxylation (c1) and removal of phosphate on C3
Formation of Squalene
isoprenes fuse to form geranyl pyrophosphate

another isoprene is added to form farnesyl pyrophosphate

two farnesyl pyrophosphates fuse to form squalene (requires NADP)

Conversion of Squalene to Cholesterol
cyclase converts squalene 2,3-epoxide to lanosterol, precursor to cholesterol
→plants make stigmasterol instead, fungi ergosterol
Cholesterol stored and transported as esters
when we transport cholesterol, we can transport it as cholesterol or cholesteryl ester

need lipoproteins to transport

within cells, use ACAT (acyl-CoA cholesterol acyl transferease) to form ester

in HDL, use LCAT (lecithin-cholesterol acyl transferase) to form ester
De novo synthesis of cholesterol predominates
de novo synthesis in man is 1g/day

0.3 g per day from food

plasma levels of 150-200 mg/dL are maintained mainly by de novo synthesis
4 Main Mechanisms of Cholesterol Regulation
HMG-CoA reductase
→main biosynthetic control
→phosphorylation decreases activity
→level of enzyme also regulated

Acyl-CoA: cholesterol acyltransferase, ACAT

LDL/HDL ratios

Excretion as bile acids
Overview of Cholesterol Regulation
Insulin increases levels of HMG-CoA reductase → increase cholesterol biosynthesis (overeating)
→Want to get rid of glucose, main product is acetyl-CoA, used to make cholesterol

Glucagon has opposite action of insulin

Feedback mechanism from high levels of cholesterol: inhibit HMG-CoA reductase
→Protease that breakdowns HMG-CoA (NOT FEEDBACK INHIBITION)
→→→Feedback inhibition is transient, enzyme breakdown has longer effect because enzyme must be resynthesized

Cholesterol also stimulates ACAT to make cholesteryl esters
Transport of Cholesterol Occurs in Lipid-Protein Complexes
In intestine, chylomicrons (protein-lipid complexes) form, enter capillaries, acted upon by lipoprotein lipase
→Breaks chylomicrons up, form chylomicrons remnants, go to liver, make VLDL, go back to capillary
→VLDL broken down to make VLDL remnants
→→→Some back to liver
→→→Some make LDL, which can go back to liver or be delivered to extrahepatic tissues

leaves tissue as HDL
→when it goes back to liver, it ultimately makes LDL again
Human Plasma Lipoproteins
Cholesterol is hydrophobic and requires protein association for solubility

Number of different lipoprotein complexes are found in circulation and are characterized by unique sets of apoproteins and different ratios of lipid to protein resulting in particles of varying density

Each form of lipoprotein complex plays a particular role in cholesterol transport and control
Apolipoproteins of Human Plasma Lipoproteins
ApoB-100 - protein in LDL which binds to LDL receptor which is bound in cells (cells that only take up cholesterol if they have LDL receptor)

ApoD - specific to HDL
Bile Acid Synthesis
liver synthesizes 2 primary bile acids: chenodeoxycholic acid and cholic acid
→most bile acids (98%) are conjugated via an amide bond to either glycine or taurine
→both free and conjugated bile acids are secreted into gall bladder (for storage) and intestine

Conjugated bile acids are converted into free form in intestine

Bacteria convert primary bile acids to dozens of secondary bile acids including 7-deoxycholate and lithocholate

In gut, primary and secondary bile acids emulsify fat, 95% are reabsorbed and return to liver

In liver, primary and secondary bile acids are reconjugated to glycine or taurine and resecreted

5% of primary and secondary bile acids are excreted
→Main excretory pathway for cholesterol (90%)
→10% excreted as steroid hormones and derivatives

KNOW relative structure is similar to cholesterol, plus acid group and additional hydroxy group
Feedback Control of Bile Acid Synthesis (Simplified)
∙LXR turns on 7-hydroxylase
∙Cholesterol activated LXRα

∙bile acids feedback through many different pathways to inhibit LXRα or 7-hydroxylase
Fair tests of treatments
comparisons are key to all fair tests of treatments

testing treatments
Approaches to evaluating interventions
uncontrolled, noncomparative
∙clinical impression
∙case reports, case studies
∙cross-sectional study

controlled, comparative
∙case-control study
∙cohort study
∙clinical trial
→crossover or parallel
Studies can be observational or experimental
observational: exposures and events are observed

Experimental: intervention is applied, effect on outcome observed
∙clinical trial (prophylactic or treatment intervention in individuals)
∙community trial (prophylactic or treatment intervention in communities)
Epidemiologic Approaches to Diet and Disease
Cross-Sectional Studies (observational)
→correlation studies

Case-Control Studies (observational)

Cohort Studies (observational)

Clinical Trials (experimental)
→randomized, double-blind, placebo-controlled trials

Community Trials (experimental)

Metabolic Studies (experimental)
Cross-sectional studies
are also called frequency surveys or prevalence studies

assess exposure and outcome at same time

only assess prevalence, not incidence

the National Health and Nutrition Examination Survery (NHANES) is cross-sectional study
Case-control studies
retrospective studies that start with an outcome and then look back in time for exposure (factors believed to be related to disease or condition)

compares group of patients who have a condition or disease with group of patients who do not

cases have condition or disease, controls do not

case-control studies are best for testing rare diseases, new diseases, adverse drug rxns
Cohort studies
longitudinal study that follows group of people who share common characteristic or experience within defined period (e.g. are born, leave school, lose job, exposed to drug/vaccine)

follow group/groups over time

compare outcomes over time for exposed and non-exposed groups

can be used to calculate incidence rates

can examine multiple outcomes

cohort studies are better for common outcomes that rare outcomes
→can be useful in examining rare exposures that are common in certain groups
Randomized controlled trials
are best way to test therapies

treatment group is compared to control group

control may receive:
∙no treatment
∙placebo or sham
∙comparator (proven treatment)
helps to:
∙eliminate bias on treatment assignment
∙facilitate blinding (masking)
∙increases likelihood that unknown factors that make make a difference are evenly distributed
∙allocation must be concealed for proper randomization
∙an intention-to-treat analysis helps to preserve randomization
Metabolic study
first period of time, all participants have same diet

second period of time, diets different

difficult to do
Epidemiologic Approaches to Diet & Disease: Cross-Sectional & Correlation studies
Strength: contrasts in dietary intake are large, average diet is stable vs. individual diets, large population sizes
Limitations: correlation is not cause & effect, potential determinants of disease vary, limited by "food disappearance" data, data cannot be independently reproduced
Epidemiologic Approachesto Diet & Disease: Case-Control Studies
Strength: generating data is fairly easy & targeted, best when variable is clear-cut (i.e. smoking)

Limitations: # of subjects is typically small, selection of appropriate control group, many opportunities for methodologic bias (including biased recall of diet), many opportunities for confounding
Epidemiologic Approachesto Diet & Disease: OBSERVATIONAL STUDIES IN GENERAL
can never prove efficacy of therapy

are very important for:
∙monitoring health and disease in populations
∙finding associations that should be tested in controlled trials
∙assessing risks of therapies

should be used in formulation of hypotheses to be tested in clinical trials
Adherence Bias
good adherence is marker for other healthy behaviors
Formation of Mature Insulin
secreted by beta cells of pancreas in response to high levels of glucose

initially synthesized as single polypeptide (preproinsulin)

C domain helps protein fold properly (proinsulin) before it is cleaved off (mature insulin)
→C domain not degraded, stays in ER
→can measure level of C peptide in suspected diabetics
→injected insulin is in mature form already
Key Effects of Insulin

stimulates glycogen synthesis in muscles and liver

stimulates uptake of glucose by muscle cells, adipocytes and other cells

promotes uptake of branched amino acids, favoring build-up of muscle protein

stimulates protein synthesis, inhibits intracellular protein degradation
Effect of Glucagon
Primary target is liver
→for all practical purposes, no effect on muscle

stimulates glycogen breakdown

inhibits glycogen synthesis by diminishing by triggering phosphorylation of phosphorylase and glycogen synthase

inhibits FA synthesis by diminishing production of pyruvate and lowering activity of acetyl-CoA carboxylase
→shut down forward rxn in glycolysis, FA synthesis

stimulates glucose synthesis by liver by lowering F-2,6-BP

activates lipase in adipocytes
Effect of Catecholamines
can be considered neurotransmitters because secreted by sympathetic nervous system
→epinephrine and norepinephrine

strong effect on muscle, liver, adipose tissue

mobilizes glycogen in muscle and liver, TAGs in adipocytes

stimulates secretion of glucagon and inhibits secretion of insulin
→wants to amplify its own effect

Inhibits uptake of glucose by muscle → switch over to fatty acids

Increase amount of glucose released into blood by liver
Difference between glucagon and catecholamines
Glucagon is released in response to low levels of blood glucose

Catecholamines released in response to stress
→minor stress counts (i.e. running)
Insulin Secretion
Glucose enters cell through GLUT 2 transporters at rate equal to blood glucose concentration, creates ATP through TCA, glycolysis, OxPhos
→ High blood glucose, high rate of entering cell

ATP increase inhibits K⁺-channel, K⁺ no longer leaves cell at same rate, causes depolarization

Depolarization communicated to voltage-gated Ca²⁺ channels, opens channel, Ca²⁺ ions rush into cell, ER will also release its Ca²⁺ stores

Ca²⁺ important second messenger that causes insulin granule from Golgi to fuse with cell membrane, be secreted
Insulin Receptor
dimer of 2 alpha subunits, each associated with transmembrane beta protein
→dimerize upon insulin binding
→beta protein has domain in intracellular space (tyrosine kinase domains), autophosphorylation site

when insulin binds to alpha subunits, it induces conformational change in beta subunits, acting on tyrosine kinase domains and leading to its own autophosphorylation
→this phosphorylation begins signal transduction pathway
Insulin Signaling Pathway Involves Cascade of Protein Kinases
key thing to understand about insulin-signaling pathway is that it is complex, takes place in cytosol as well as nucleus

do not need to know whole thing, does not always do the same thing

Glucose Uptake and Glycogen Synthesis Promoted by Insulin
PKB, while crucial, is not specific to insulin
→phosphorylates things

PKB promotes glycogen synthesis by phosphorylating glycogen synthase kinase 3 (GSK3), inactivating it
→P-GSK3 = inactive, GSK3 = active
→GSK3's job is to phosphorylate glycogen synthase (GS) and inactivate it, but P-GSK3 means it can't make P-GS, leaving GS on and activated, glycogen synthesized

Increases number of glucose transporters in plasma membrane by adding GLUT 4 transporters
→GLUT 4 usually sequestered in intracellular vesicles, but increase in PKB → increase in GLUT 4

Exercise considered stress → increases catecholamine releases, mobilizes glycogen and fat stores, down regulates insulin release
→exercise can also be thought of as mechanism for glucose uptake independent of insulin
→conditioning allows body to take up more glucose in presence of catecholamines
→→→major reason why exercise is great for all, but especially Type II diabetics who are severely insulin resistant
Catecholamine receptors and signaling
when catacholamines bind to β-adrenergic receptor, ATP is transferred to cAMP through actions of adenyl cyclase
→cAMP them activates PKA

PKA is kinase that phosphorylates many different substrates
→most important are phosphorylase kinase (PK) and glycogen phosphorylase (GP)
→P-PK = active, P-GP = active
∙P-PK → P-PG → breakdown of glycogen to G1P, eventually to glucose

Signal in cascade amplified because enzymes like adenyl cyclase turn over and activate multiple substrates, resulting in massive glycogen mobilization and glucose into blood stream
Formation of cAMP
when catacholamines bind to β-adrenergic receptor, ATP is transferred to cAMP through actions of adenyl cyclase
Signal transduction & α-adrenergic receptor
catecholamines can also signal through α-adrenergic receptor

upon hormone binding, g-protein coupled rxn occurs from GDP to GTP, leading to activation of phospholipase C (PLC)
→ acts to remove head group from PIP2, cleaving it to diacylglycerol (DG) and IP3

→unique in that it needs two things to be active:
1. needs to bind to DG in plasma membrane (from cleave of PIP2)
2. needs calcium, which is released from ER upon IP3 binding to calcium channel
Formation of DP & IP3 second messengers
Glycogen metabolism in muscle
Exercise ups levels of catecholamines

using β-adrenergic receptors, cAMP will be produced, turns on PKA
∙PKA → P-PK (on) → P-GP (on)
∙helps w/ massive mobilzation of glycogen stores
∙PKA does not directly phosphorylate GP, must go through PK

also want to inhibit glycogen synthesis, so PKA will also P-GS to inactivate it
→GSK3 will also acto to P-GS

α-adrenergic effects of the catecholamines in the muscle, which can influence the balance by acting on GP, which will turn off (aka phosphorylate) P-GS
Glucose → insulin → more GLUT4 → more glucose in cell

in process, dephosphorylates hormone sensitive lipase (P-HSL on, HSL off), inactivating it and favoring TAG synthesis over mobilizatoin of fatty acids

FA delivered to liver cells and mobilized into energy sources with increased levels of catecholamines and glucagon (starvation)
Fatty Acid Metabolism in fed and fasting states
Acetyl-CoA carboxylase is first committed step in FA synthesis

malonyl CoA is added 2 at a time from acetyl-CoA

controlled by phosphorylation

P-AcCoA carboxylase = off, AcCoA carboxylase = on

insulin promotes dephosphorylation of AcCoA carboxylase, activating it, which makes sense → when we have a lot of glucose we want to synthesize FA to store in adipocytes
Diabetic Ketoacidosis
Hyperglycemia (over 300mg/dL)

Ketonemia and ketonuria
→TCA intermediates have been depleted by gluconeogesis, so liver starts producing ketone bodies, lots of them in blood (ketonemia) and urine (ketonuria)

Acidosis (pH <7.35)
→ketone bodies are acidic, drops blood pH

Low bicarbonate
→blood pH has dropped so bicarbonate buffering system steps in to try and raise pH but becomes depleted in process

Water deficit
→glucose binds to water because of its hygroscopic property, b/c blood sugar is so high, some of glucose ends up making it past kidney and getting peed out, taking water with it

Potassium deficit (5 mEq/kg body weight)
→to adjust for low pH, K⁺ leaves cells in exchange for H⁺ entering, some of K⁺ is excreted, screws with CNS

High blood urea nitrogen (BUN)
→ gluconeogenesis is going crazy, lots of amino groups have been liberated, liver made them to urea to be disposed of, so urea in blood is also high


Treatment: hydration, respiration, electrolytes, insulin (moderate)
Pretty much only in liver (90%) and kidney
→only other organ that does anything even like this is adipose tissue, up to DHAP step

begins with carbon skeleton supplied by three sources: glucogenic amino acids (alanine), lactic acid and glycerol
∙when using carbon skeletons from glucogenic amino acids or lactic acid, oxaloacetate must be formed within mitosol
∙glycerol from TAG breakdown in adipose tissue travels to liver and follows different path for gluconeogenesis
→converted to glycerol-3-phosphate, then DHAP, eventually to glucose (skipping many steps taken by lactic acid and glucogenic amino acids)
∙glycerol only primary source of carbon skeletons from gluconeogenesis under starvation conditions

set of enzymatic rxns that convert carbon skeleton starting materials to glucose is basically reverse of glycolysis with 3 exceptions: kexokinase, PFK-1, pyruvate kinase
∙these rxns are too exergonic to perform in reverse, must be bypassed
b/c reverse of glycolysis is not energetically favorable, need large amount of ATP to drive gluconeogenesis, energy supplied by FA β-oxidation
First Bypass of Gluconeogenesis
bypasses pyruvate kinase rxn of glycolysis

lactic acids from anaerobic metabolism in muscle or glucogenic amino acids from protein breakdown enter hepatocyte mitochondria matrix and are converted to pyruvate
→alanine's alpho-keto acid is pyruvate

Pyruvate then converted to oxaloacetate in rxn catalyzed by pyruvate carboxylase
→still in mitosol

oxaloacetate must be transported to cytstol → converted to malate
→difficult process b/c oxaloacetate cannot cross inner mitochondrial membrane
→converted to malate, shuttled out of mitochondria, reoxidized back to oxaloacetate and NADH w/in cytosol
→rxn also serves to produce reducing equivalents (NADH) that will be used to drive GAPDH rxn of glycolysis in reverse

last step is to convert oxaloacetate to phosphoenolpyruvate (PEP)
→catalyzed by PEP carboxylase
→can take place in cytosol (glucogenic amino acids) on w/n mitochondrial matrix (lactate)
→for lactate, reducing equivalents for reverse GAPDH rxn have already been formed through conversion of lactate to pyruvate, rxn catalyzed by lactate DH
→→→w/o need to generate NADH, oxaloacetate converted to PEP w/in mitosol
Important points about first bypass in gluconeogenesis
Pyruvate carboxylase uses biotin cofactor

anaplerotic rxn, meaning rxn increases number of TCA intermediates, therefore accelerates TCA and rate of acetyl-CoA oxidation

Acetyl-CoA activates pyruvate carboxylase
→means that when we build up acetyl-CoA, rate of rxn increases, leading to production of more oxaloacetate, and increase if rate of acetyl-CoA oxidation
→rxn proceeds by hydrolysis of ATP, energy expensive

glucogenic amino acids that aren't alanine (glutamate, glutamine) can also be used for gluconeogenesis even though they do not form pyruvate
→accomplished by converting TCA intermediates, i.e. α-ketoglutarate, which leads to increase in oxaloacetate
Alternate paths from pyruvate to PEP in first bypass of gluconeogenesis
takes place in cytosol when starting point is glucogenic amino acids
→crosses inner mitochondrial membrane as malate

takes place in mitochondrial matrix when starting point is lactate
Second Bypass of Gluconeogenesis
→glycosis rxn: F6P → F1,6bP
→gluconeogenesis: F1,6bP → F6P

in gluconeogenesis, rxn catalyzed by FBPase1

both forward and reverse rxn are regulated by F2,6bP
→acts as indicator of how much F6P is available for glycolysis
→high levels of F2,6bP stimulate glycolysis, inhibit gluconeogenesis
Role of F2,6BP in glycolysis and gluconeogenesis
dboth forward and reverse rxn are regulated by F2,6bP
→acts as indicator of how much F6P is available for glycolysis
→high levels of F2,6bP stimulate glycolysis, inhibit gluconeogenesis

levels determined by activity of enzyme with competing subunits, PFK-2/FBPase-2
→PFK2 converts F6P to F2,6bP, stimulating glycolytic pathway, inhibiting gluconeogenesis
→FBPase-2 catalyzes opposite rxn, stimulating gluconeogenesis

Activity of enzyme determined by phosphorylation, under hormonal control
→well fed state (high insulin): no need for gluconeogenesis, favors unphosphorylated state, activating PFK-2, leadings to larges amounts of F2,6bP, stimulating glycolysis
fasting state (high glucagon/catecholamines): elevated level of cAMP activates PKA, which phosphorylates PFK-2/FBPase-2, which activates FBPase-2, decreasing F2,6bP, stimulating gluconeogenesis
Third Bypass of Gluconeogenesis
bypasses hexokinase step

dephosphorylates glucose-6-phosphate

catalyzed by glucose-6-phosphatase

G6P to glucose is essential to release glucose into blood stream
→enzyme performs same function in mobilization of glycogen
Gluconeogenesis & Fatty Acid Oxidation
Important facts from earlier:
∙β-oxidatoin of FA yields acetyl-CoA, which cannot contribute to overall increase in TCA intermediates and therefore cannot be used as starting material in gluconeogenesis
→still important b/c it supplies energy to drive gluconeogenesis
∙an increase in any TCA intermediate will lead to increase in oxaloacetate, thus accelerating oxidation of acetyl-CoA, providing materials for gluconeogenesis
∙use of AAs, pyruvate and lactic acid for gluconeogenesis must proceed through oxaloacetate, TCA intermediate, generated by conversion of pyruvate (via pyruvate carboxylase) or by conversion of AA to other TCA intermediates
→in starvation conditions, gluconeogenesis will deplete oxaloacetate, causing TCA to slow down, leading to build up of acetyl-CoA

during exercise, lactic acid is generated in muscle performing anaerobic metabolism → lactic acid goes to liver → converted to pyruvate → tends to be converted to oxaloacetate → increases TCA intermediates, starting materials for gluconeogenesis
∙catecholamines are high → activate PKA → activate hormone sensitive lipase → FA oxidation → increase in acetyl-CoA
∙this is good in exercise b/c oxaloacetate from lactate will undergo gluconeogenesis, driven by energy of acetyl-CoA oxidation, will help restore muscle's glycogen stores
→→→assuming athlete not starving, plenty of TCA intermeidates, meaning acetyl-CoA is being oxidized rapidly, rapid oxidation produces high-energy electrons primed for OxPhos

under fasting/starving conditions, glucagon/catecholamines are released into blood stream → stimulate β-oxidation of FA to acetyl-CoA
∙problem is if we are not taking in glucose, hepatocytes cannot convert glucose → pyruvate → oxaloacetate to replenish TCA intermiedates, provide starting materials for gluconeogenesis
→→→body becomes dependent on glucogenic AAs to supple TCA intermediates, provide starting materials for gluconeogenesis
→→→process is slow, leading to build up of acetyl-CoA, leads to production of ketone bodies, which can supply brain energy and lessen amount of AAs needed for gluconeogenesis, helping fulfill 2nd metabolic priority: preserve protein

meal high in FAs and protein, low in carbs will NOT stimulate insulin secretion b/c no carbs
→FA oxidation with occur, glucogenic AAs ingested will be used to make pyruvate or TCA intermediates
Using amino acids in gluconeogenesis and maintaining nitrogen balance
Under starvation conditions when glycogen has been depleted, liver uses large store of AAs in muscle to make glucose and replenish depleted TCA intermediates, first step in this process is deamination of AAs to yield respective α-keto acid, which is then converted to oxaloacetate

When protein is being degraded for metabolic fuel, excess nitrogen MUST BE ELIMINATED
→in muscle, amino groups from catabolized AAs are transferred to α-keto acids (pyruvate or α-ketoglutarate) forming alanine and glutamate/glutamine, AAs then transported to liver for deamination
→in liver: AAs deaminatoed to form NH4+ for excretion, α-keto acids used for gluconeogenesis
**→Deamination rxns accomplish two important goals: allow free nitrogen to be detoxified and supply α-keto acid carbon skeletons for gluconeogenesis**
Alanine and Gluconeogenesis
serves as major non-toxic carrier of nitrogen in blood stream

when it reaches liver, it undergoes transamination rxn
→transfers its amino group to α-ketoglutarate, forming pyruvate (from the α-keto acid of pyruvate) and glutamate (from α-ketoglutarate)
→rxn catalyzed by aminotransferase, uses pyridoxal phosphate (PLP) as cofactor
→pyruvate used to produce oxaloacetate for gluconeogenesis
→glutamate undergoes oxidative deamination, removing ammonium ion so it can be detoxified in urea cycle, and restoring TCA intermediate α-ketoglutarate
→→→rxn catalyzed by glutamate DH
→→→importance of this enzyme is that it is one exception that can use either NAD+ or NADP+ as cofactor
Glutamine and Gluconeogenesis
other major non-toxic carrier of nitrogen in blood

you can attach 2 nitrogens for price of one TCA intermediate
→glutamine is basically α-ketoglutarate with 2 nitrogens attached

in hepatocyte mitochondria, one nitrogen is liberated by glutaminase, forming glutamate, glutamate then undergoes oxidative deamination to form α-ketoglutarate
The Urea Cycle
serves to take extra amino groups that are cleaved off and combines two of them to form urea, more easily trashed compound that isn't as toxic while floating around in blood

occurs exclusively in liver

5 steps:
1. CPS 1
2. ornithine transcarbamoylase
3. argininosuccinatae synthetase
4. argininosuccinase
5. arginase

very circuitous route for making urea → could have been formed more simply by attaching ammonium ion to extra phosphate bond left over on carbamoyl phosphate in step 1
→drawn out mechanism forms several significant connections to TCA, helps body maintain second metabolic priority: PRESERVE PROTEIN
Step 1 of Urea Cycle: CPS 1
Carbamoyl phosphate synthetase 1 (CPS 1)

takes place in mitochondria of liver cells and utilizes 2 ATP to attach an NH₄⁺ and phosphate to bicarbonate molecule in three-step rxn
1. one ATP used to add high energy phosphate bond to one end of bicarbonate, forming carbonic phosphoric anhydride
2. nucleophilic substition, ammonium ion replaces phosphate, forming carbamate
3. one additional ATP burned to phosphorylate other end of bicarbonate, forming final product carbamoyl phosphate
Step 2 of Urea Cycle: Ornithine Transcarbamoylase
Ornithine - nonstandard alpha-amino acid that is regenerated at end of every urea cycle

using extra phosphate group from last step for energy, carbomyl phosphate is added to ornithine, forming citrulline

last step that occurs in mitochondria, reset occurs in cytosol
Step 3 in Urea Cycle: Argininosuccinate synthetase
2 ATP used (effectively), 1 NH₄⁺ added (as aspartate)

citrulline transported out of mitochondria, into cytosol

First, ATP is hydrolyzed to AMP and pyrophosphate (PPi), AMP is attached to citrulline
→energy expenditure amounts to 2 ATP b/c pyrophosphate is immediately cleaved into 2 inorganic phosphates, meaning we have used 2 high energy bonds that will need to be reformed

Aspartate is added in nucleophilic substitution rxn, replacing AMP and brining with it second ammonia group to for argininosuccinate
Step 4 in Urea Cycle: Argininosuccinase
carbon skeleton of aspartate cleaved off, ejecting fumarate molecule, forming arginine
→fumarate is TCA intermediate
Step 5 in Urea Cycle: Arginase
arginase hydrolyzes a bond to form urea, regenerates ornithine, which can enter mitochondria to re-enter cycle at step 2

urea is released into bloodstream, collected by kidney for excretion
Urea Cycle Regulation
can be regulated by:
∙N-acetyl glutamate
Urea Cycle Regulation by Ornithine
can be formed from glutamate, whose carbon skeleton is alpha-ketoglutarate

essential member of urea cycle
→cycle's speed is dependent on concentration, production of ornithine is dependent on [glutamate] and [alpha-ketogluratate]
→in starvation, gluconeogenesis is actively drawing upon TCA intermediates to form glucose, glucogenic AAs are being broken down into amino groups and TCA intermediates to feed process
→since urea cycle must keep up with gluconeogenesis disposal of ammonium groups, level of ornithine must also stay high
→→→limits rate of AA catabolism, preserving protein
Linkage of TCA and Urea Cycle
Urea Cycle Regulation by N-acetyl glutamate
allosteric activator of first step of urea cycle (step catalyzed by carbamoyl phosphate synthetase), spurring forward rxn

formed by action of N-acetyl glutamate synthase, which takes glutamate and attachs acetyl group from acetyl-CoA

another significant connecter b/w urea cycle and TCA, serves to help preserve protein
→high levels of glutamate are required to form N-acetyl glutamate, which causes urea cycle to move forward more quickly during step 1
→if gluconeogenesis and AA catabolism are occurring too quickly, glutamate/alpha-keto glutarate levels will be low, less N-acetyl glutamate will be formed, urea cycle will slow down to prevent protein from being catabolyzed too quickly

also significant indicator of how much excess ammonia needs to be disposed of, since alpha-ketoglutarate is primary acceptor of ammonia groups during transmaination rxns that occur as amino acids are being broken down
→when glutamate is high, 1st rxn of urea cycle moves forward more quickly, trashing nitrogen more quickly as well, occurs during nonstarvation conditions when TCA intermediates are plentiful
Urea Cycle Regulation by Aspartate
2nd amino donor in urea cycle (Step 3) but carbon skeleton is TCA intermediate oxaloacetate

Glutamate DH removes amino group from glutamate, forming alpha-ketoglutarate and 1 NADH, extra amino group is used by aspartate aminotransferase to aminate oxaloacetate to aspartate
→when TCA intermediates like oxaloacetate are low, levels of asparate will be low, urea cycle will slow down, preserving proteins
Urea Cycle Regulation by Fumarate
multiple fates:
∙can be converted into malate by fumarase, re-enter mitochondria, continue along with its role in TCA, becoming oxaloacetate by action of malate DH and generating 1 nADH
∙if cell is actively undergoing gluconeogenesis, fumarate can stay in cytosol, be converted to malate, then be converted to oxaloacetate by cytosolic malate DH, also generating 1 NADH
→keeps oxaloacetate out of mitochondria, allows cytosolic PEP caroxykinase to begin gluconeogenesis, converting it to PEP
Urea Cycle Energetics
slightly energetically favorable

1 NADH is generated in rxn where glutamate is deaminated, 1 NADH generated when fumarate is ejected from cycle, moves through steps to be oxaloacetate

leaves us with 2 NADH which will form 5 ATP
→since 4 used in urea cycle, net gain of 1 ATP from cycle
Metabolic Priorities during Starvation
Priority 1: provide glucose to brain and RBCS

Priority 2: preserve protein
→protein breakdown for energy is self-limiting in this respect, but it is also just a slow process in general

high levels of catecholamines and low levels of insulin that would expect during time of starvation help body accomplish these goals:
∙high catecholamines stimulate FA mobilization
∙FA that have been mobilized can move to liver to be broken down into acetyl-CoA and used to produce ketone bodies
∙because muscle and fat cells are insulin-dependent for uptake of glucose, when insulin is low, these tissues cease
The Kidney
minor player in body's regulation of glucose homeostasis

like liver, capable of performing gluconeogenesis, but only scales up to fulfill about 10% of body's demand

unlike liver, incapable of performing urea cycle
→significant b/c when it breaks down glutamine, it dumps 2 ammonium ions into blood

ammonium dump can be useful under starvation conditions
→with liver pumping out ketone bodies, blood pH drops
→since ketone bodies have negative charge, anion gap occurs in blood
→kidney exchanges NH4+ ions for H+ floating around in blood, raises blood pH, helps avert acidosis for as long as possible
Diabetes Mellitus
group of disease that are characterized by elevations in blood glucose or hyperglycemia due to some sort of insulin deficiency

Type I - auto-immune directed destruction of pancreatic beta cells that produce insulin

Type II - due to defective insulin secretion or insulin resistance
→obesity and other metabolic disorders are strongly associated with type II

potentially life threatening symptoms: diabetic ketoacidosis (mainly type I) and non-ketotic hyperosmolar coma
Theories about Type II diabetes
Lipostat theory

Hormones that may be involved:
Lipostat Theory
hypothesis that suggests signals from adipose tissue play significant role in eating behavior and metabolic activity, these may be deregulated in type II
hormone secreted by adipose tissue when fat cells reach limit for storing TAGs, hormone travels to hypothalamus where it binds, causing hypothalamus to suppress appetite, stimulates hypothalamus to transduce signal via SNS to adipose tissue

when signal reaches fat cells as norepinephrine (NE), NE sets forth series of signal transduction rxns:
∙levels of enzyme adenyl cyclase rise, levels of cAMP rise, levels of PKA rise

PKA does 3 main things w/in fat cell that promote mobilization of FAs, oxidation of FAs and thermogenesis
→activates hormone-sensitive lipase that can hydrolyze fats
→phosphorylates lipid-droplet-sealing protein, perilipin, to provide hydrolytic enzymes access to TAGs
→moves to nucleus, upregulates expression of uncoupling protein (UCP), proton-pore protein that disconnects OxPhos from ATP formation, allowing proton gradient to generate heat instead

Leptin believed to make cells more sensitive to insulin

Leptin activates AMP-dependent protein kinase (AMPK), kinase that likely phosphorylates some of same downstream insulin substrates as well
→AMPK's primary function is to minimzie energy expenditure and maximize usage of energy reserves during times when AMP is abundant, indicating ATP is low
hormone secreted almost exclusively by adipose cells

activates AMPK

mice with defects in adiponectin as less sensitive to insulin
Therapies for Diabetes
exercise can increase levels of AMP w/in cells, activate AMPK, helps sensitize cells to insulin

drugs like Avandia elevate adiponectin secretoin from adipose tissue, increase activity of AMPK in other tissue
Primary Sources of Energy
→Protein → Glucose

Free FAs

Glucose vs. Free Fatty Acids
Brain Glucose Utilization
Requires 120 grams/day, ~25% of total body energy requirements

Brain function dependent on glucose level about 70mg/dL
Fuels Used by Various Body Tissues
Fasting State
bring about changes in way glucose is handled in body

w/in first few hours, body uses all exogenous glucose, followed by mobilization of glycogen stores, which lasts about a day

gluconeogenesis ramps up around same time glycogen stores begin to run out, bas this does a number on body b/c it is taking protein from muscle and other sources and converting it to glucose
→not good, but ketogenesis kicks in around day 8
Regulation Hormones
→involved in glucose uptake in cells, except for special organs (brain)
→during fasting conditions, glucose not plentiful → insulin levels decrease to save glucose fur use by brain, encourage FFA mobilization

Glucagon, epinephrine and cortisol:
→increase during fasting state in order to raise and maintain blood sugar
→rise causes glycogenolysis, gluconeogenesis, FFA mobilization
Fed State
insulin is secreted by beta cells in pancreas and allows glucose uptake into liver and muscles
Diabetes diagnosis
if after fasting, blood glucose is >126 mg/dL
Bolus insulin
take 0-120 minutes after meal to prevent high amount of glucose in blood
Basal insulin
needed in between meals and at night in order to mimic basal levels normally found in body
Gestational diabetes
glucose/other energy sources cross through placenta and supply fetus

if glucose levels aren't well controlled in mother, effects can be amplified in fetus

insulin from baby's pancreas also responds
→if mother doesn't have well-controlled glucose levels, insulin levels in fetus will be out of balance and can cause increased growth and fat deposition

Exogenous insulin cannot cross placenta
→can control glucose in mother w/o affecting fetus
Type I diabetes
clinical manifestations can be seen when only 10% of beta cells exist

treatment is to administer proper amount of exogenous insulin
Type II diabetes
insulin resistance

more and more insulin needed to uptake set amount of glucose into tissues, will eventually lead to beta cell deficiency and dysfunction
→operating at such high capacity, they become exhausted quickly
Periods of most rapid growth
Infancy and adolescence, not childhood
Nutrients essential for brain development




Nutrients essential for growth of adipose tissue

Fat soluble vitamins A, D, E and K
Nutrients essential for growth of musculature

vitamin C
Nutrients essential for bone mineralization


vitamin D
Nutrients essential for sexual maturation
Nutrients essential for blood formation


Failure to thrive
describes inadequate growth, may result from insufficient nutrition, numerous medical conditions, environmental circumstances

Iron Deficiency
most common nutrient deficiency

can result in delayed speech, impaired growth, delayed cognitive development, anemia
Water Absorption
Na⁺-K⁺ ATPase
→ATPase keeps intracellular sodium concentration low

→powered by sodium concentration gradient, two sodium ions pull one glucose and more than two hundred water molecules through transporter

results from decreased absorption in small intestine and net secretion in large intestine, which produces large stool volume with significant water and electrolyte loss
Oral Rehydration Therapy
goal: replenish and maintain adequate hydration and nutritional status during or following diarrheal episode

involves oral administration of solutions specifically designed to utilize SGLT-1 transporter to replace fluid and electrolytes rapidly
wasting disease, results from lack of protein and energy in diet

emaciated appearnce, apathy, lower body temp

damage can be irreversible
arises when energy intake is almost adequate but protein is lacking

bloated abdomen, edematous limbs
Glucogenic & Ketogenic Amino Acids
Glucogenic - generate pyruvate or Krebs cycle metabolites

Ketogenic - generate acetyl-CoA and ketone bodies
Vitamin A
needed for vision, regulation of gene expression, and control of cell proliferation
Muscle Fuel Sources
fat is basic muscle fuel

glycogen provides booster power
Activity and Fuel Use
High carbohydrate intake can boost muscle glycogen stores

low performance can point to inadequate glycogen repletion
Effects of Cytokines
induce NO production, fever, muscle proteolysis, decreased albumin production by liver, increased uptake of zinc into liver, sequestration of iron by reticuloendothelial system, proliferation of fibroblasts
Physiological Response to Stress
increased cardiac output, body temp, oxygen consumption

increased secretion of interleukins (cytokine), tumor necrosis factor (cytokine), catecholamines, glucotricoids, glucagon, insulin
Stages of Folate Deficiency
serum folate drops

RBC folate drops, liver stores decrease

hypersegmented neutrophils appear

MCV increases

hemoglobin drops (late event)