Lipid and Palmitate Biosynthesis

Lipid Biosynthesis: An Overview

• Lipid synthesis and breakdown follow distinct metabolic routes.
• Synthesis predominantly occurs in the cytosol, while breakdown mainly takes place in the mitochondria.
• Fatty acid synthesis involves acyl carrier proteins (-SH groups), whereas breakdown involves CoA (-SH groups).
• Enzymes for synthesis comprise a single polypeptide (fatty acid synthase), while breakdown relies on multiple enzymes.
• Biosynthesis utilizes NADPH/NADP+, and breakdown NADH/NAD+.

Palmitate Biosynthesis

• The net reaction for palmitate biosynthesis requires 8 acetyl-CoA, 7 ATP, and 14 NADPH to produce palmitate, 7 ADP, 7 Pi, 14 NADP+, 8 CoASH, and 6 H2O.
• Palmitate rapidly becomes Palmitoyl-CoA for use in triacylglycerol (TAG)& phospholipid synthesis.
• Acetyl-CoA, made in the mitochondrial matrix, must be transported to the cytosol for fatty acid synthesis.
• The Tricarboxylate transport system (citrate-malate-pyruvate shuttle) facilitates the transfer of Acetyl-CoA into the cytoplasm.
• Acetyl-CoA combines with oxaloacetate to form citrate, which then crosses the mitochondrial membrane into the cytosol.
• ATP-citrate lyase in the cytosol converts citrate back into acetyl-CoA and oxaloacetate; this process requires ATP.
• Malic enzyme produces NADPH for fatty acid biosynthesis.
• Eight acetyl-CoA molecules yield eight NADPH molecules through the malic enzyme pathway which supports palmitate synthesis.
• The pentose phosphate pathway also provides NADPH required for fatty acid synthesis through oxidation reactions.

Acetyl-CoA Carboxylase (ACC)

• Acetyl-CoA carboxylase (ACC) is the key enzyme, catalyzing the first committed step in fatty acid synthesis.
• ACC in animals is a multifunctional protein that polymerizes.
• The inactive protomer form turns into the active polymer form.
• Covalent modification regulates ACC activity.
• Unphosphorylated ACC is active at low citrate concentrations, while phosphorylated ACC requires high citrate levels for activation.
• Glucagon and epinephrine hormones trigger AMP-dependent phosphorylation inactivating ACC.
• Insulin triggers dephosphorylation activating ACC to store glucose and energy as FA.
• Biotin is on a flexible tether which delivers carboxyl groups from the carboxylase to the carboxyltransferase.

Fatty Acid Synthase (FAS)

• Mammals use homodimeric fatty acyl synthase I (FAS I) for fatty acid synthesis.
• FAS-I consists of 270-kD polypeptides containing all reaction centers for fatty acid production.
• Yeast and fungi use two multifunctional polypeptide chains for FAS activities.
• Plants and bacteria employ separate, independent enzymes, referred to as fatty acid synthase II (FAS II).
• The steps in fatty acid synthesis are similar across organisms.
• Fatty Acid Synthase (FAS) is a dimer of multifunctional polypeptides.
• A megasynthase is an example.
• It has autonomous domains, each with a specific but different catalytic function.
• Autonomous domains includes a catalytic function of subunits associated as a head-to-tail dimer
• Reactions mirror previous pages.
• Reactions take place in the multienzyme complex.
• Acyl Carrier Protein (ACP) carries intermediates of FA biosynthesis
• Somewhat larger versions of Coenzyme A specialized for FA biosynthesis.

Enzymes Activities

• Malonyl/acetyl transferase (MAT)
• β-Ketoacyl-ACP Synthase (KSase or KS)
• β-Ketoacyl-ACP Reductase (KR)
• 2,3-trans-Enoyl-ACP Reductase (ER)
• Acetyl & Malonyl groups transferred from CoA to ACP to growing fatty acid chain
• Forms thioesters with acyl groups using a product of catabolism for anabolic processes
• Long flexible chain facilitates transport of substrate between various enzymatic domains of multienzyme complex Fatty Acid Synthase

Palmitate Biosynthesis steps (bacteria and plants)

• Acetyl and malonyl groups are moved from CoA to ACP via malonyl/acetyl transferase (MAT).
• An acetyl group is transferred to β-ketoacyl-ACP synthase (KSase or KS).
• Formation of a C-C single bond takes place, between acetyl (or acyl group) & malonyl group involving β-ketoacyl-ACP synthase (KS or KSase).
• Formation is driven by decarboxylation with carbon from CO2 added to acetyl-CoA.
• Carbon is added to acetyl-CoA to make malonyl-CoA from acetyl-CoA carboxylase (ACC).
• Carbon is not incorporated into the growing fatty acid chain.
• Reduction of β-Carbonyl to β-Hydroxyl Group uses NADPH and β-ketoacyl-ACP reductase (KR)
• Dehydration to Form C=C Double bonds through B-ketoacyl-ACP reductase (KR and loss of H2O occurs to form an α, β C=C double bond.
• Reduction to form Saturate FA performed by 2,3-trans-enoyl-ACP reductase (ER) and uses NADPH.

Palmitate Biosynthesis End

• The process repeats by transferring butyryl-ACP to KSase and adding another malonyl group on ACP.
• At the end- each additional round adds two more carbons to growing fatty acyl group
• β-ketoacyl-ACP synthase (KSase) cannot accommodate substrates larger than 16 carbons
• FA biosynthesis concludes with hydrolysis of palmitoyl-ACP into palmitate & free ACP using palmitoyl thioesterase, TE
• One acetyl-CoA combines with 7 Malanoyl-CoA to produce one Palmitate.
• End product: 1 acetyl-CoA + 7 Malanoyl-CoA + 14 NADPH + 13 H+ + H2O -> Palmitate + 7 HCO3 + 8 CoASH + 14 NADP+.
• The whole process consumes 49 ATP equivalents for the conersion of palmitate into palmitate CoA in animal cells.

Elongation and Desaturation of Fatty Acids

• Elongases, found in two cellular locations, facilitate fatty acid elongation.
• In the ER, they introduce two-carbon units using malonyl-CoA (decarboxylation).
• In mitochondria, they add two-carbon units using acetyl-CoA (thiolase).
• Desaturases in E. coli:
• Bacteria can introduce double bonds at the end of the chain .
  • The double bond is near the β-carbonyl group and the thioester group via an oxygen-independent pathway.)
• Animals can add double bonds anywhere in the chain (oxygen-dependent pathway) but only before the fatty acid reaches full length.
• Unsaturation reactions occur in eukaryotes within the middle of an aliphatic chain, catalyzed by enzymes like stearoyl-CoA desaturase.

Essential Fatty Acids: Omega-3 and Omega-6

• Linoleic and α-linolenic acids are termed essential fatty acids and cannot be synthesized by animals.
• Linoleic acid is a precursor to arachidonic acid, both classified as omega-6 fatty acids.
• alpha-Linolenic acid gives rise to eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), termed omega-3 fatty acids.

Regulation of Fatty Acid Metabolism

• Fatty acid biosynthesis and beta-oxidation are reciprocally regulated.
• Malonyl-CoA, at high concentrations, serves as a central regulator, inhibiting fatty acyl-CoA entry into mitochondria (blocking the carnitine shuttle) and stimulating fatty acid synthesis; beta-oxidation is inhibited and fatty acid synthesis is turned on.
• Citrate activates the conversion of acetyl-CoA to Malonyl-CoA, boosting fatty acid synthesis.
• Fatty acyl-CoA inhibits the conversion of acetyl-CoA to malonyl-CoA, preventing fatty acid biosynthesis through feedback inhibition.
• Hormonal regulation: -Glucagon inhibits fatty acids synthesis in low glucose conditions and stimulates glycogen breakdown (and fatty acid beta-oxidation). -Insulin promotes fatty acid synthesis and inhibits fatty acid breakdown.

Complex Lipid Synthesis - Glycerophospholipids

• Complex lipid synthesis involving Glycerol or DHAP can be used to synthesize TAGs
• Organisms depend upon different lipid biosynthetic pathways
• Eukaryotes synthesize: TAGs, Sphingolipids, and cholesterol
• Bacteria synthesize simple phospholipids

Building Blocks for Complex Lipids

• Glycerol, DHAP, or dietary monoacylglycerols can be used to synthesize TAGs (triacylglycerols).
• CDP-Diacylglycerol is a precursor of phosphatidylinositol, phosphatidylglycerol, and cardiolipin in eukaryotes.

Key Points

• Sphingolipids and triacylglycerols
  • Only eukaryotes synthesize Sphingolipids and triacylglycerols
• PE (phosphatidylethanolamine)
  • Constitute approximately 75% of phospholipid in E. coli
• E. coli lack
  • PC (phosphatidylcholine), PI (phosphatidylinositol), sphingolipids and cholesterol
• Bacteria
  • Some bacteria produce PC while most do not

Sphingolipid Biosynthesis

• Sphingolipid biosynthesis begins with the condensation of serine and palmitoyl-CoA.
• This process occurs at high levels in neural tissue.
• The enzyme 3-ketosphinganine synthase, which depends on pyridoxal phosphate (PLP) as a coenzyme is needed to condense serine and palmitoyl-CoA.
• Ketone reduction requires NADPH.
• This is followed by acylation and double bond formation.
• The product is ceramide used for the formation of sphingolipids.

Bile Acids synthesis and function

• Bile acids are polar carboxylic acid derivatives of cholesterol
• Bile acids aid in the solubilizing of dietary lipids, acting as detergents.
• Bile acids are made in the liver and stored in the gallbladder to be secreted when needed into the intestines
• Oxidation and Hydroxylation processes are important in this pathway
• In the intestines they become oxidized to cholic acid
• Taurine is added to make taurocholic acid
• Glycine is added to make glycocholic acid

Steroid Hormone

• Steroid hormones are crucial signalling molecules
• They do not bind the cell surface of the plasma membrane
• Steroid hormones diffuse directly through the plasma membrane.
• Hormones then bind to receptors in the nucleus, thus they can control gene expression such as transcriptions