Lipid Metabolism: Fatty Acid Synthesis and Breakdown

Lipids (fats) exists in these forms

triacylglycerols (TAG), fatty acid, phospholipids, steroids, glycolipids

function of lipids

Energy storage, Constituents of  membranes, Anchors for membrane proteins, Cofactors for enzymes, Signaling molecules, Pigments, Detergents, Transporters, Antioxidants

Dietary lipids are broken down by lipases

Lingual lipase, Gastric lipase, Pancreatic lipase

Emulsification of lipids by bile salts occurs in the ______ → micelles

duodenum

Bile salts are synthesized in the ______ and stored in the _______

liver, gall bladder

Anabolism of fatty acids → requires ____ and _____

• takes place in cytosol in animals, chloroplast in plants

acetyl-CoA and malonyl-CoA, NADPH

Catabolism of fatty acids → produces ______ and ______

• takes place in the mitochondria

acetyl-CoA and NADH, FADH₂

Fatty Acid Synthase I (FAS I)

• FAS I in ______

vertebrates and fungi

Fatty Acid Synthase I (FAS I)

• FAS II in _______

plants and bacteria

Summary of FAS I action:

• Reactions occur in the ACP domain of FAS I

  • ACP= Acyl Carrier Protein

• Uses two enzyme-bound thiol (-SH) groups as activating groups

  • Prep step

• Catalyzes a repeating four-step cycle that elongates the fatty acyl chain

• Chain grows by two carbons after one turn of the cycle

• Process uses ______ as the electron donor

NADPH

Acyl Carrier Protein (ACP) Serves as a Shuttle in Fatty Acid Synthesis

• Contains a covalently attached prosthetic group _________

  • flexible arm to hold the acyl chain while carrying intermediates from one enzyme subunit to the next

• Delivers acetate (in the first step) or malonate (in all the next steps) to the FAS

• Shuttles the growing chain within the active site of FAS I during the four-step reaction

4’-phosphopantetheine

Prep Step: Chain Transfer and Charging ACP and FAS I with Acyl Groups

• Two thiols must be charged with the ________ before the condensation reaction can begin.

  • thiol from 4-phosphopantetheine in ACP

  • thiol from Cysteine in FAS I

correct acyl groups

Prep Step: Chain Transfer and Charging ACP and FAS I with Acyl Groups

• The acetate fragment of acetyl-CoA is transferred to ACP.

  • catalyzed by _______

• ACP passes this acetate to the Cys-SH of the KS domain of FAS I

  • alpha-ketoacyl-ACP synthase (KS) domain (region)

• ACP –SH group is recharged with malonyl from malonyl-CoA.

  • catalyzed by _________

Acetyl CoA-ACP acetyltransferase, Malonyl CoA-ACP transacylase

Step 1 of FA synthesis: Condensation

• Condensation reaction attaches acetyl group (or longer fatty acyl chain) to malonyl group

  • Occurs with simultaneous release of CO₂

  • This activates malonyl group to attack the acetyl carbonyl

    • Creates acetoacetyl-ACP, which is a b-keto intermediate

• Catalyzed by ______

• Decarboxylation of malonyl-CoA makes the reaction energetically favorable.

b-ketoacyl-ACP synthase

Step 2 of FA synthesis: Reduction

• Reduction #1

  NADPH reduces the -keto intermediate to an alcohol.

  Ketone carbon (-carbonyl) reduced to form an alcohol

  -hydroxybutyryl-ACP

  NADPH is e− donor

  Reaction is catalyzed by -ketoacyl-ACP reductase (KR)

B-ketoacyl-ACP reductase (KR)

Steps 3 and 4 of FA synthesis

• Step 3: Dehydration: -OH group from C-2 and -H from neighboring CH2 are eliminated, creating double bond (trans-alkene).

  • -OH and -H removed from C-2 and C-3 of B-hydroxybutyryl-ACP to form trans-D2-butenoyl-ACP

  • catalyzed by _______

• Step 4: Reduction #2: NADPH reduces double bond to yield saturated alkane

  • NADPH is the e- donor to reduce double bond of trans-D2-butenoyl-ACP to form butyryl-ACP.

  • catalyzed by _______

• Final Result: Increases chain length of the acyl group of the fatty acid by 2 carbons

B-hydroxyacyl-ACP dehydratase (DH), enoyl-ACP reductase (ER)

Fatty Acid Storage

• Fatty acids are converted to triacylglycerol (TAGs)

• TAGs are stored in ______

  • Ready for mobilization when energy is needed

adipose tissue

Fatty Acid Storage

• Very little of TAG are stored in the liver

  • Instead they are exported from the liver

  • Packaged with _________ and ___________

    • Different lipoprotein particles (Chylomicrons, VLDL, LDL, HDL)

cholesterol esters, phospholipids

Macromolecular aggregates made of

• Core of nonpolar lipids (TAG, C, CE)

• Monolayer of phospholipids

• Apolipoproteins

Solubilize and transport lipids in the blood

Lipoproteins

Fatty Acid Transport into Mitochondria

• Fats are degraded into fatty acids and glycerol in the cytoplasm of adipocytes.

• Fatty acids are transported to other tissues for fuel through the blood.

• B-oxidation of fatty acids occurs in mitochondria.

• Small fatty acids (< 12 carbons) diffuse freely across mitochondrial membranes.

• Larger fatty acids (most free fatty acids) are transported via ______

acyl-carnitine/carnitine transporter

Catabolism of lipids

• _______ of TAG is catalyzed first by hormone-sensitive lipase, and then by other lipases

  • Hydrolysis releases free fatty acids (and glycerol)

Hydrolysis

Hormone-sensitive lipase (HSL)

• When insulin is low, epinephrine is high

• Epinephrine binds to the receptor

  • activates adenylyl cyclase

  • Causes rise in cellular cAMP levels

• cAMP activates a protein kinase

• Active Protein Kinase phosphorylatesHSL

  • Phosphorylated HSL = ____

  • Dephosphorylated HSL = _____

• Activated HSL causes hydrolysis of TAGs

• Hydrolysis of TAGs produces free fatty acids

active, inactive

To enter matrix, fatty acids need to be in ______ form

Fatty acid entry into mitochondria via the acyl-carnitine/carnitine transporter. After fatty acyl–carnitine is formed at the outer membrane or in the intermembrane space, it moves into the matrix by facilitated diffusion through the transporter in the inner membrane. In the matrix, the acyl group is transferred to mitochondrial coenzyme A, freeing carnitine to return to the intermembrane space through the same transporter. Acyltransferase I is inhibited by malonyl-CoA, the first intermediate in fatty acid synthesis (see Fig. 21–2). This inhibition prevents the simultaneous synthesis and degradation of fatty acids.

acyl CoA

Stages of Fatty Acid Oxidation

• Stage 1 consists of oxidative conversion of two-carbon units into acetyl-CoA via ______ with concomitant generation of NADH and FADH2.

  • involves oxidation of B-carbon to thioester of fatty acyl-CoA

• Stage 2 involves oxidation of acetyl-CoA into CO2 via ________

  • generation of GTP, NADH and FADH2

• Stage 3 generates ATP from NADH and FADH2 via the _________

B-oxidation, TCA cycle, e- transport chain

The B-Oxidation Pathway

• Each pass (4-step sequence) removes one acetyl group (2C) in the form of acetyl-CoA.

• Repeated passes produces more acetyl CoA

• Acetyl CoA enters the TCA cycle to produce energy

In each pass through this four-step sequence, one acetyl residue (shaded in pink) is removed in the form of acetyl-CoA from the carboxyl end of the fatty acyl chain—in this example palmitate (C16), which enters as palmitoyl-CoA. (b) Six more passes through the pathway yield seven more molecules of acetyl-CoA, the seventh arising from the last two carbon atoms of the 16-carbon chain. ______ molecules of acetyl-CoA are formed in all.

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Fatty Acid Breakdown: Step 1: Dehydrogenation of Alkane

• Oxidation

• Catalyzed by ________ on the inner-mitochondrial membrane

• Results in trans double bond, different from naturally occurring unsaturated fatty acids

• Electrons from bound FAD transferred directly to the e- transport chain via electron-transferring flavoprotein (ETF)

• Step 1 is analogous to the succinate dehydrogenase reaction in the citric acid cycle

acyl-CoA dehydrogenase (AD)

Fatty Acid Breakdown: Step 2: Dehydrogenation of Alkane

• Re-hydration

• Catalyzed by _____

• Hydrolysis of alkene

• Addition of water produces an alcohol

• Step 2 is analogous to the fumarase reaction in the citric acid cycle

  • Stereospecific addition

enoyl-CoA hydratase

Fatty Acid Breakdown: Step 3: B-Oxidation

• Oxidation

• Catalyzed by __________

• The enzyme uses NAD redox cofactor as the hydride acceptor.

  • Electrons are transferred to the e- transport chain

• Reaction of Step 3 is analogous to malate dehydrogenase reaction of the citric acid cycle

B-hydroxyacyl CoA dehydrogenase

Fatty Acid Breakdown: Step 4: B-Oxidation

• Catalyzed by _________

• The ketone carbon in -ketoacyl-CoA is electrophilic.

• SH in CoA-SH acts as a nucleophile and attacks the ketone carbon

  • This releases one molecule of acetyl-CoA.

• The net reaction is thiolysis of the carbon-carbon bond (hence thiolase)

• The carbon chain is shortened by 2 carbon atom

  • Palmitic acid (C-16) is converted to Myristic (C-14) acid

acyl-CoA acetyltransferase (aka thiolase)

Summary of β-oxidation

• Catabolic reactions of fatty acids: Energy release!

• Location: Mitochondrial matrix

• Fatty acid-acyl CoA substrate enters the matrix

  • Vehicle: _______

• Definition of β-oxidation: A sequence of four reactions acting on the fatty acid-acyl CoA

• Outcome: Shortening of the fatty acid chain, 2C units a time

• The two-carbon units are released as acetyl CoA

  • The final thiolytic cleavage releases two units of acetyl CoA

carnitine shuttle

Fatty Acid Catabolism for Energy

• For palmitic acid (C16)

  • Repeating the previous four-step process six more times (seven total) results in _______ molecules of acetyl CoA

    • FADH2 is formed in each cycle (seven total).

    • NADH is formed in each cycle (seven total).

    • Last step produces two molecules of Acetyl CoA

      • From acetoacetyl CoA

• Acetyl-CoA enters TCA cycle

  • This makes more GTP, NADH, and FADH2.

• Electrons from all FADH2 and NADH enter electron transport chain to produce ATP

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• Synthesis of ketone bodies is termed ________

• During a fasting state, liver has elevated levels of fatty acid

• Liver produces acetyl CoA by fatty acid breakdown

  • Process: β-oxidation

• Elevated Acetyl CoA levels cause upregulation of pyruvate carboxylase

  • In which pathway, have you seen this enzyme?

  • Pyruvate to OAA

  • Gluconeogenesis!

• Acetyl CoA enters the ketogenic pathway, instead of coupling with OAA

  • This protects OAA from entering gluconeogenesis (or Krebs)

    • Management of cellular resources

ketogenesis

Ketone Bodies

• Entry of acetyl-CoA into citric acid cycle requires _______

• When oxaloacetate is depleted, acetyl-CoA is converted into ketone bodies

  • This occurs when there is low carb intake, or diabetes

• Three forms of ketone bodies can leave the liver, and are delivered by the blood to various tissues

  • Acetone

  • Acetoacetate

  • B-hydroxybutyrate

• Cardiac muscle, skeletal muscle, renal cortex, and brain can use ketone bodies as a fuel source

  • Acetone is not really used; it is eliminated in breathing

oxaloacetate

Formation of Ketone Bodies: Generating Free CoA (Step 1)

• Reverse of the last step of B-oxidation

  • _______(4C) is produced from two molecules of acetyl CoA (2C)

  • Thiolase

Acetoacetyl CoA

Formation of Ketone Bodies: Generating Free CoA (Step 2)

• A third molecule of acetyl-CoA is incorporated in this step

• _______ (6C) is produced

• HMG CoA synthase

HMG CoA

Formation of Ketone Bodies: Degradation of HMG-CoA (Step 3)

• HMG CoA is broken down by HMG CoA lyase (present in the mitochondrial matrix) to produce the ketone bodies

• ________ now enter blood

• Acetone is exhaled

  • Pulmonary exchange of gases

• Acetoacetate andB-hydroxybutyrate can move to the brain (and other tissues) for use in energy production

• Ketone bodies are essentially “masked versions” of acetyl CoA

• They are “unmasked” once they enter the tissue (e.g. brain, heart)

Acetone, acetoacetate, and B-hydroxybutyrate

Ketone Bodies as Fuel

• D-β-Hydroxybutyrate reaches other tissues where it finally produces acetyl CoA in 3 steps

• Step 1: Oxidized to acetoacetate by _______

• Step 2: Thioester formation with CoA using _________

• Step 3: Breakdown of acetoacetyl CoA by ______

• The acetyl-CoA thus formed is used for energy production

Overall you can see that ketone bodies #1 and #2 are “unmasked” in the tissue to “reveal” acetyl CoA

B-hydroxybutyrate dehydrogenase, B-ketoacyl CoA transferase, thiolase

TAGs are hydrolyzed to yield free fatty acids through lipase action influenced by _____

epinephrine

______ enables the movement of fatty acid acyl CoA to the matrix for B-oxidation

Carnitine shuttle

Two-carbon units in fatty acids are oxidized in a four-step B-oxidation process into acetyl-CoA

• NADH and FADH2 produced yield ATP via _____

electron-transport chain

Under fasting or untreated diabetic conditions, acetyl-CoA formed in the liver is converted to ______ that serve as fuels for other tissues

ketone bodies

Once the ketone bodies reach the _____, they get re-converted to acetyl CoA

• This acetyl CoA can produce energy within the tissue

tissues