Sources pof energy in fasting state In adipose tissue: In fasting state, the stored TAG will be the major source of energy. -Stored TAG in adipose tissue is hydrolysed by hormone sensitive lipase (HSL) into free fatty acids and glycerol. -Glycerol will move to liver -Adipose tissue oxidize some free fatty acids to get energy. The remaining fatty acids will move to blood, carried by albumin and enter peripheral tissues (except brain) to be oxidized and produce energy. -N.B. Blood brain barrier is impermeable to fatty acids, so brain can’t use fatty acids as source of energy.

Liver in fasting state: liver can use the following sources of energy: 1- Free fatty acids (from adipose tissue) is oxidized to produce energy 2- Glycerol (from adipose tissue), amino acids (from degradation of muscle protein), and lactate (from muscles), all are used as substrates of gluconeogenesis in liver to produce glucose. This glucose could be released into blood and up-taken by brain to be used as source of energy. 3- Liver can degrade the stored glycogen (by glycogenolysis) to produce glucose that is used as source of energy. **** In fasting state, liver synthesize ketone bodies, which is then released and up-taken by all peripheral tissues including brain to be oxidized to produce energy. Liver can’t use ketone bodies as source of energy.

Muscles in fasting state: Muscles can use either fatty acids (from adipose tissues) or ketone bodies (from liver) Metabolism of lipids in fasting state I-Fatty acid oxidation II- Ketone bodies synthesis and degradation

I-Fatty acid oxidation 1- hydrolysis of TAG to release free fatty acids (lipolysis) and mobilization of fatty acoids to peripheral tissues : Fatty acids are the part to be oxidized so the first step is release of fatty acids from TAG. This process is initiated by hormone- sensitive lipase (HSL) which removes fatty acids from TAG. HSL is activated by anti-insulin hormones (adrenaline, glucagon, GH, cortison) and inhibited by insulin. HSL has two forms, phosphorylated form (active form) and dephosphirylated form (inactive).

In fasting state, anti-insulin hormones (glucagon, adrenaline, thyroxine) are released, bind to their cell membrane receptors, activate adenylate cyclase which convert ATP into cAMP. cAMP activate protein kinase A which phosphorylate HSL and activate it. However in fed state, insulin is released and activate protein phosphatase enzyme which dephosphorylate HSL and inactivate it.

Activation of adipose tissue HSL

2- Activation of free fatty acids: Free fatty acids is mobilized from adipose tissue to all tissues (except brain) to be oxidized. Before oxidation, fatty acids should be activated into acyl CoA. Site of activation: Long chain fatty acids are activated in cytoplasm of cells. In the presence of ATP and CoA, the enzyme thiokinase (or called Acyl CoA synthetase) catalyses the activation of long chain fatty acids into acyl CoA. RCOOH+2ATP + CoASH → RCO ~ SCoA acyl CoA

3- Beta oxidation of fatty acids: It is the major pathway of oxidation (catabolism or breakdown) of saturated fatty acids in which two carbons are removed from activated fatty acid, producing acetyl CoA, NADH and FADH 2 Site: in the mitochondria of all tissues particularly in the liver. So there is no fatty acid oxidation in RBCs which have no mitochondria. Note that: fatty acids with less than 12 carbons (short and medium chain fatty acids) are activated in mitochondria then oxidized. Since activation of long chain fatty acids (more than 12 C) occurs in cytoplasm (their thiokinase are cytoplasmic enzymes) and the oxidation occur in mitochondria, so long chain acyl CoA should be transported.

Transport of long chain fatty acyl CoA into mitochondria: Carnitine shuttle Long chain fatty acyl CoA cannot penetrate mitochondrial membrane (as mitochondrial membrane is impermeable to CoA). They need a carrier to transport them into mitochondria. This carrier is Carnitine which transports active fatty acid by the help of 3 enzymes: Carnitine acyltransferase I (CAT-1) Carnitine - acylcarnitine translocase Carnitine acyltransferase II (CAT-2) Steps of transport: 1) acyl group is transferred from acylCoA into carnitine by CAT-1 to give acyl carnitine and free CoA which remains in cytoplasm. 2) Acyl carnitine is transported into mitochondria by the help of Carnitine - acylcarnitine translocase. 3) CAT-2 catalyses the transfer of acyl group from acyl carnitine to CoA to give acyl CoA and free carnitine which go back to cytoplasm by translocase enzyme. Inhibitor of carnitine shuttle: malonyl CoA which inhibits carnitine acyltransferase I Sources of carnitine: diet (meat products), synthesized from amino acids lysine and methionine in liver and kidney.

Mitochondria

Steps of β- oxidation: β-oxidation consists four sequential steps. These steps are repeated until all the carbons of an even-chain fatty acyl-CoA are converted to acetyl-CoA. Reactions: 1. Oxidation: FAD accepts 2 hydrogens from a fatty acyl-CoA in the first step. A double bond is produced between the α- and β- carbons, and an enoyl-CoA is formed. -Enzyme: acyl-CoA dehydrogenase 2. Hydration: H 2 O is added across the double bond, and a β- hydroxyacyl-CoA is formed. -Enzyme: enoyl-CoA hydratase 3. Oxidation: β-Hydroxyacyl-CoA is oxidized by NAD+ to a β- ketoacyl CoA. ‒ Enzyme: L-3-hydroxyacyl-CoA dehydrogenase

4. Thiolysis: The bond between the (α- and β-carbons of the β- ketoacyl CoA is cleaved by a thiolase that requires coenzyme A. Acetyl CoA is produced from the two carbons at the carboxyl end of the original fatty acyl CoA, and the remaining carbons form a fatty acyl CoA that is two carbons shorter than the original. -Enzyme: β-ketothiolase 5. The shortened fatty acyl CoA repeats these four steps. Repetitions continue until all the carbons of the original fatty acyl CoA are converted to acetyl CoA. a. The 16-carbon palmitoyl CoA undergoes seven repetitions. b. In the last cycle, a four-carbon fatty acyl CoA (butyryl CoA) is cleaved to two acetyl CoAs.

Energy yield from β- oxidation: Each β- oxidation cycle yields : -acyl CoA with 2 carbon atoms less -one NADH+H + (give 3 ATP via respiratory chain) -one FADH 2 (which give 2 ATP via respiratory chain) - one acetyl CoA (oxidized through Kreb's cycle to yield 12 ATP). Remember that the last cycle produce two acetyl CoA. Question: Oxidation of one molecule of palmitic acid yields 129 ATP ( How?). Palmitic acid need 7 cycles to be completely converted into acetyl CoA. 7x 3ATP (from NADH+H+) = 21 7x 2ATP (from FADH2 ) = 14 8 x12 (from a cetyl CoA) = 96 Total 131, since 2ATP are used for activation, so the net ATP yield is = 129 ATP

Energy yield from one molecule of palmitic acid (16C): palmitic undergo 7 cycles of oxidation With Production of 8 acetyl CoA 8 acetyl CoA x12 ATP = 96 ATP 7 NAH x3 = 21 7FADH2 x 2= 14 Total =131 2ATP utilized for FA activation, so net is 129 ATP C16 → C14 + Acetyl CoA

Regulation of β- oxidation: Oxidation of fatty acids is controlled by the rate of release of free fatty acids from adipose tissues (lipolysis), which is inhibited by food intake and insulin, and stimulated by starvation, adrenaline, glucagon, thyroxin, glucocorticoids and growth hormone (the factors that regulate the action of HSL).

fatty acid β-oxidationfatty acid synthesis Greatest flux of pathway (diet regulation) Increase in fasting state Decrease in fed state Increase in fed state Decrease in fasting state Hormonal state that favor pathwayAnti-insulin hormonesInsulin Major tissue site (organ)All tissues especially liverLiver (mainly), adipose tissue Site inside the cellsMitochondriacytoplasm PrecursorAcyl CoAAcetyl CoA Regulatory enzymeCAT-1Acetyl CoA carboxylase (ACC) Two-carbon donor/productAcetyl CoA (two carbons product) Malonyl CoA (two carbons donor) Allosteric ActivatorCitrate, ATP Allosteric InhibitorPalmitate Carrier of acyl/acetyl groups between mitochondria and cytosol Carnitine (caryy long chain acyl CoA from cytoplasm to mitochondria Citrate (caryy acetyl CoA from mitochondria into cytoplasm) H-carrier (Coenzymes)FAD, NADNADPH End productAcetyl CoAPalmitate Steps of the pathwayOxidation, Hydration Oxidation Thiolysis (cleavage) Condensation Reduction Dehydration Reduction Comparison between Fatty acid oxidation, Fatty acid synthesis