Triglyceride and Fatty Acid Metabolism

Triglycerides (triacylglycerol) consist of three fatty acids each esterified to one of the carbons in glycerol

They have three physiological uses:

(1) Efficient Source of Fuel – fatty acids Þ acetyl CoA, glycerol Þ glycolysis

more highly reduced than carbohydrates so more energetic (9 kcal/g instead of 4 kcal/g)
hydrophobic, so easy to store – each gram of glucose requires 4 g of H₂O

(2) Structural Components – phospholipids, cholesterol

(3) Precursors of Signalling Molecules – prostaglandins, leukotrienes

Absorption and Transport of Lipids

Most Common Fatty Acids in Ingested Triglycerides:

palmitic acid (16 carbons, no double bonds) – 20% of diet

oleic acid (18 carbons, one double bond at C-9) – 50% of diet

linoleic acid (omega 6; 18 carbons, two double bonds at C-9 and C-12) – 10% of diet – essential fatty acid (from plants)

used to make arachadonic acid (20 carbons, four double bonds at C-5,8,11,14) – important precursor molecule

Triglycerides are insoluble in water and do not cross membranes – must be broken down and resynthesized at least twice

pancreatic lipase in small intestine breaks down into glycerol and fatty acids

olestra (olean) – fatty acids conjugated to sucrose – not a substrate for pancreatic lipase Þ not absorbed Þ laxative properties and does not allow for absorption of vitamins D, E, A, K

resynthesized into triglycerides in intestinal cells (except small fatty acids <10 carbons – can diffuse to blood)

released in chylomicron (a lipoprotein) – can cross membranes – to lymph then bloodstream

endothelial cells then break down fatty acids from lipoproteins (chylomicrons or VLDL) by means of lipoprotein lipase

lipoprotein lipase is induced by insulin

Lipid transport – Lipoprotein (triglyceride core, phospholipid/cholesterol shell, structural and recognition proteins) – 4 types:

(1) chylomicron – mostly triglyceride – used to transport triglycerides from intestine to tissues – made in intestinal ER

(2) VLDL (very low density lipoprot.) – made by liver for adipose tissue – denser than chylomicron (less triglyceride)

fatty acids are made from carbohydrates by the liver when it has enough glycogen

(3) LDL – formed when triglycerides cleaved from VLDL – high in cholesterol, transports cholesterol to tissues

(4) HDL – almost no triglyceride, mostly protein – functions as cholesterol "scavenger," absorbing from cells

Lipid Entry into Cells

During fasting, fatty acids from triglycerides are used as a source of fuel

hormone-sensitive lipase in adipose tissue produces glycerol and fatty acids and release them directly to the blood

fatty acids are detergents and will destroy membranes – hence bound to serum albumin

albumin is the most abundant protein in blood (40-50 g/L) – each molecule tightly binds 10 fatty acids (affinity = 10mM)

hormone sensitive lipase activated by protein kinase A phosphorylation ([cAMP] Ý )

Glycerol – to DHAP by glycerol kinase and glycerol DH (glycerol + ATP Þ glycerol-3P Þ DHAP + NADH)- gluconeogenesis

reverse reaction is used by adipose cells to make glycerol, but only liver has glycerol kinase (for forward reaction)
in late starvation (4-5 weeks), glycerol is the major source of blood glucose

Fatty Acids – absorbed by almost all tissues (except RBC and brain), converted into acetyl CoA by b -ox (in mito)

released from albumin and esterified to fatty acyl CoA (to prevent detergent activity) by fatty acyl CoA synthase/thiokinase (uses 2 ATP equivalents)
transported into mitochondria via carnitine shuttle (only use for carnitine) – three enzymes:

(1) CAT-1 – in cytosol – esterify fatty acid to acyl carnitine, releasing CoA

regulated step – malonyl CoA(-) – turn it off when synthesizing fatty acids in cytosol

(2) CAT-2 – in mitochondria – esterify fatty acid to acyl CoA, releasing carnitine

(3) carnitine translocase – antiport carnitine (out of mitochondria) and acyl-carnitine (into mitochondria)

Fatty Acid b -oxidation – occurs in mitochondria

b -oxidation is centered around b -carbon (third from end):

Four steps, first three analogous to succinate Þ OAA in TCA cycle

(1) Oxidation to trans double bond, producing FADH₂

analogous to succinate Þ fumerate

oxidation

(2) Addition of H₂O to produce –OH group

analogous to fumerate Þ malate

hydration

(3) Oxidation to ketone, producing NADH

analogous to malate Þ oxaloacetate

oxidation

(4) Nucleophilic cleavage by CoA producing Acetyl CoA

cleavage

Energetics:

Palmitate (16 carbons, no double bonds) goes through seven rounds of b -oxidation (last four carbons Þ 2 acetyl CoA) to produce 7 NADH (Þ 21 ATP), 7 FADH₂ (Þ 14 ATP), and 8 acetyl CoA (x 12 in TCA = 96 ATP)

total is 131 ATP – invested two to make acyl CoA when initially absorbed, so net total is 129 ATP from one fatty acid

Regulation of oxidation:

the most significant regulation is CAT-1 (i.e., availability of acyl CoA in mitochondria)

Special Cases:

(1) Odd Number of Carbons – end up with three-carbon acyl CoA (propionyl CoA)

propionyl CoA carboxylase (requires biotin) adds a COO^- to the a -carbon, producing methylmalonyl CoA

methylmalonyl CoA mutase (requires B₁₂) moves the COO^- to the b -carbon, producing succinyl CoA Þ TCA

deficiency of this enzyme leads to methylmalonyl acidurea Þ mental retardation

(2) Unsaturated Fatty Acids (Containing Double Bonds) – other enzymes move double bond forward and make trans

enter b -oxidation at step 2. Above

(3) Branched Fatty Acids – common in plants – b -oxidation occurs down to last carbon or undergoes a -oxidation

odd carbon leaves as CO₂ (oxygen added to a carbon by monooxygenase)

Refsum’s disease – if unable to a -oxidize Þ phytanic acid builds up

(4) Long Fatty Acids – longer than 12 carbons are oxidized in peroxisomes

Ketone Body Synthesis – another use for fatty acids, occurs in liver mitochondria

Acetoacetyl CoA is made from two acetyl CoA molecules (via thiolase), or from reversal of last step of b -oxidation

CoA group is removed in 2-step process:

(1) condensation with acetyl CoA to produce HMG CoA via HMG CoA synthase

(2) removal of acetyl CoA via HMG CoA lyase

This produces acetoacetate, which can be further reduced to 3-Hydroxybuterate

ratio of acetoacetate to 3-HO butyrate is dependent on ratio of NADH/NAD

Utilization of ketone bodies:

3-hydroxybuterate is first oxidized to acetoacetate, producing one NADH

acetoacetate is converted to acetoacetyl CoA (along TCA succinyl CoA Þ succinate)

two acetyl CoA’s are liberated, producing 24 ATP

one GTP lost when succinyl CoA Þ succinate, so net 23 ATP

the additional NADH means 3-hydroxybutyrate carries 26 ATP of energy

similar energy to glucose

Fatty Acid Biosynthesis – occurs in cytosol of adipose tissue and liver

Fatty acid synthesis is very much like oxidation in reverse

Transport – First, acetyl CoA must be transported from the mitochondria to the cytosol – uses 2 ATP

made into citrate (first part of TCA), but citrate transported out of mitochondria

acetyl CoA released from citrate producing OAA (uses ATP)

OAA Þ malate Þ pyruvate (malic enzyme: produces 50% of NADPH) Þ mitochondria Þ OAA (by pyruvate carboxylase, uses another ATP)

First carbon in fatty acid is from acetyl CoA, but the rest are from malonyl CoA

malonyl CoA is produced from acetyl CoA by acetyl CoA carboxylase(requires ATP and biotin) – adds COO^- to acetyl part of acetyl CoA
carboxylase activity is upregulated by two mechanisms:

(1) polymerization – induced by citrate(+), make fatty acids when much TCA citrate (too much carbohydrate)

(2) dephosphorylation – by cAMP dependent phosphetase (kinse is NOT cAMP dependent)

protein phosphate is cAMP dependent which indirectly controls phosphorylation

insulin is (+) regulator of FA synthase and acetyl CoA carboxylase

Synthesis takes place by means of a single protein, fatty acid synthase

enzyme has two arms – cysteine residue and acyl carrier protein (ACP); both can from thioester bond like CoA

first acetyl CoA binds ACP, then is transthiolated to the cysteine residue

new malonyl CoA binds ACP, then terminal carbon from first acetyl attaches to a carbon of malonyl, producing CO₂

b -keto acyl intermediate reduced to OH by NADPH, dehydrated to double bond, reduced again to single bond

growing chain transthiolated to cysteine residue again, process starts over

Each round requires 2 NADPH (7 rounds x 2 NADPH = 14 NADPH)

for each round, one NADPH is produced by malic enzyme (produces 50% of NADPH in body; pentose pathway is responsible for the rest)

Regulation is primarily by acetyl CoA carboxylase activity

also insulin-controlled transcription levels

Fatty Acid Modification – elongation or desaturation

(1) Elongation – can add carbons to fatty acid, up to 24 total (although rarely beyond 18)

in mitochondria – through acetyl CoA and NADH (essentially reversal of oxidation)
in cytosol – through malonyl CoA and NADPH (similar to fatty acid synthesis)

(2) Desaturation – add cis double bond – catalyzed by a monooxygenase (called fatty acid desaturase) in two steps:

(1) uses molecular O₂ to produce an alcohol
(2) uses NADPH to oxidize to a double bond

cannot add double bonds more than 9 carbons from end, so linoleic acid (Þ arachadonic acid) must come from diet

Triglyceride Synthesis

Two acyl CoA groups are added to glycerol-3P (made from DHAP) – this forms phosphatidic acid

phosphate is removed, forming diacylglycerol

a third acyl CoA is added, forming triacylglycerol

Note: The small intestine resynthesizes triglycerides from 2-monoacylglycerol after absorption