Lecture 10 - Lipogenesis and Pentose Phosphate Pathway

Learning Outcomes:

  • Recall the overall scheme for the synthesis of fatty acids from glucose

  • Understand the reaction catalysed by and the regulation of acetyl CoA Carboxylase (ACC)

  • Describe the reaction sequence of fatty acyl synthase (FAS)

  • Apply knowledge of the regulators of acetyl CoA carboxylase to different physiological situations

  • Describe the process of esterification

  • Ingrate the pathways and processes that need to come together to support and supply lipogenesis

  • Summarise the effect of insulin on the activity of pyruvate dehydrogenase

  • Evaluate the probability of the different fates of acetyl CoA in lipogenic tissue

  • Explain the role of citrate in controlling the major anabolic and catabolic pathways

  • Outline the mechanisms for the return of oxaloacetate to the mitochondria after citrate cleavage

  • Describe the purpose of the pentose phosphate pathway and how it fits into lipogenesis

  • Illustrate the flow of substrates and key enzyme control points for lipogenesis

Lipogenesis:

  • Storing dietary glucose as fat

  • Build fatty acids (lipogenesis)

    • Mainly getting acetyl-coa from glucose

    • Everything in the pathway is driven by insulin

  • Join fatty acids to make triglycerides (3 fatty acids + glycerol) 3-phosphate (Esterification)

  • Need reductive power to synthesise fatty acids

    • NADPH – formed in the pentose phosphate pathway

  • Doing this in the tissues that respond to insulin – white adipose tissue or liver

  • Liver is a good place to synthesise fatty acids but not to store them – fatty liver

  • White adipose tissue (WAT) is a site of fatty acid synthesis

  • Lipogenesis occurs in the cytoplasm

  • Acetyl-coa go as far as citrate in the Krebs cycle to let it leave the mitochondria to the cytoplasm

  • Turn the citrate back into actyl-coa once it is in the cytoplasm to be used in lipogenesis

  • Need lots of reducing power to convert acetyl-coa to fatty acids

  • Reducing power comes from pentose phosphate pathway

  • If in the liver, need to make the decision to store as glycogen or fatty acids

  • WAT can only store as fatty acids

Acetyl-CoA Carboxylase (ACC)

  • Most regulated step in the process

  • Adds CO2 to acetyl-coa to make 3C chain, malonyl CoA

    • Requires biotin (vitamin), cofactors (Mg2+) and Used ATP

    • Takes the CO2 from biocarbonate (HCO3-) in the cell

  • Malonyl-CoA = activated Actyl-CoA

    • Ready for building new fatty acids

ACC Control:

  • Cycle of phosphorylation and dephosphoryaltion

  • Stimulated by insulin

  • Dephosphorylation by phosphates to make actice

  • Phosphorylation by kinase to make inactive

  • Insulin is actively telling the cells to turn acetyl-coa into malonyl-CoA ready for fatty acid synthesis

  • Activated allosterically by citrate

    • Lots of citrate going into the cytoplasm, this enzyme is activated

  • Inhibited allosterically, switched off, by its’ product – fatty acetyl-CoA

Lipogenesis = Reduction → dehydration → reduction (chain growing 2C at a time)

Beta oxidation = oxidation → hydration → oxidation (chain cutting 2C at a time)

Fatty Acyl Synthase Points:

  • Covalently binds to its substrates

  • Keep adding 2C via acetate

    • Do this by adding malonyl-CoA onto the FAS

  • Malonyl is decarboxylated (CO2 is removed) and leaves a negative charge on the malynol group

  • Negative charge attacks carbonyl carbon and attaches to the other chain

FAS Process

  • The chain produced is attached back to where the acetate was attached at the beginning, process repeats

  • FAS has left side for incoming malonyl CoA and right side for growing chain

Beta Oxidation vs. Fatty Acid Synthesis

Massive Multifunctional Problem:

  • Everything is happening on the one FAS enzyme

Input and Output

  • Each round of 2C addition requires:

    • 2 molecules of NADPH

    • BUT NO ATP (synthase)

      • Only ATP used was used when ACC turned acetyl-CoA into Malonyl-CoA

      • Carboxylation of acetyl-CoA does ot result in fixing CO2 as it was lost immediately when attatched with FAS (Fatty Acyl Synthase)

    • FAs released as FA-CoA when chain gets 14 to 18 carbons long

    • Desaturation of CH2-CH2 to -CH=CH= is done after release from FAS

  • We lack enzymes that introduce doubles bonds beyond carbon 9, long unsaturatesd omega-3/7 FA are essential in the diet

Esterification

  • Join fatty acids to glycerol

  • Glycerol and 3 fatty acids forms ester bonds (condensation reaction)

  • Glycerol needs to be glycerol-3P

  • Fatty acids need to be fatty acyl CoAs

Esterification

  • Turn FA into fat

  • 3 fatty acid chains are added one at a time (mono-, di-, tri-)

  • Only the liver has the enzyme to convert glycerol into glycerol 3-phosphate (glycerol kinase)

  • If in fat tissue, go halfway down glycolysis and take glyceraldehyde 3-phosphate – reduce it to glycerol 3P

  • Glycolysis provides glycerol backbone and acetyl-coa building blocks for new FA

  • Esterification enzymes like to use FA-CoA not just fatty acids

  • Esterification enzymes and FAS are upregulated by insulin at a genetic level

  • FAS expression is down regulated when lots of fat is around – as in a Western diet

Glycerol 3P Generation:

  • Glucose → Glucose 6P → Dihydroxyacetone-P

  • Dihydroxyacetone-P (3C) can be exchanged with glyceraldehyde 3P and reduced to glycerol 3P

  • LIVER ONLY – Use glycerol kinase to add a phosphate to glycerol to make glycerol 3P

  • Can also start with lactate or amino acid skeleton as a substrate and work through gluconeogenisis until it gets to dihydroxyacetone phosphate – highly unlikely

    • Termed Glycerolneogenesis

Servicing Lipogenesis: Creating NADPH for Lipogenesis

  • Pentose Phosphate Pathway

  • PDH is activated by insulin and lots of acetyl CoA is formed

  • Some may enter Krebs cycle to make NADH and make ATP (necessary for malonyl CoA)

  • Some acetyl CoA is let out into the cytoplasm to go in the Krebs Cycle oxaloacetate to create citrate

    • Allows CoA to come back, allows PDH to continue

    • Transport or oxidise the citrate formed

      • If lots of cycles happening and it is not needed in the Krebs cycle, citrate will go back out of the mitochondrion into the cytoplasm and assist with fatty acid synthesis

    • Fate of citrate depends on the energy charge within the cell

ATP-Citrate Lyase:

  • Opposite of citrate formation

  • Break 6C citrate into 4C oxaloacetate and 2C acetyl-CoA

    • Requires ATP

  • This is how the acetyl-CoA gets in the cytoplasm to do this

  • ACL is inhibited by hydroxy-citrate (OHCit)

    • Found in brindle berry

    • Sold as a fat synthesis inhibitor – doesn’t actually work

  • Oxaloacetate needs to be transported back into the mitochondria or we will run out of Kreb cycles

  • Replaced by reducing oxaloacetate with NADH to give malate and to pyruvate, which produced NADPH (necessary for recucing FA into triglycerides). Pyruvate can go back in and be converted to oxaloacetate by pyruvate carboxylase

Malonyl CoA Inhibits Beta-Oxidation

  • Malonyl CoA inhibits CAT-I

    • CAT1 transports FA into the mitochondrian where they could undergo beta oxidation, which is now inhibited

  • Therefore, FA oxidation is inhibited by malonyl CoA

  • Insulin stimulates malonyl CoA production by Acetyl-CoA Carboxylase (ACC)

  • Therefore, Insulin inhibits (beta) fatty acid oxidation by malonyl production

The Reductant: NADPH

  • NADPH is a form of NADH involved in anabolic reactions

  • Same chemical properties, just bound to different enzymes

  • Pentose Phosphate Pathway produced NADPH proportional to demand for lipogenesis

Pentose Phosphate Pathway:

  • Take glucose 6-phosphate from glycolysis

  • Glucose-6-phosphate dehydrogenase acts on glucose 6-phosphate and produced an NADPH

  • Goes through other reactions and another enzyme produced an additional NADPH

  • Leaves a 5C sugar – ribulose 5-phosphate

  • Rest of the cycle is about rearranging the amount of carbon different sugars have to produce a 6C and 3C sugar to be put back into other pathways

  • Glucose 6-phosphate dehydrogenase is regulated by demand for NADPH

  • We use NADPH during lipogenesis, which produced NADP+ which is used in the pentose phosphate pathway to produce NADPH

    Pentose Phosphate Pathway

  • NADPH is a vital anti-oxidant in cells

  • In RBC a deficiency in G6PDH enzyme causes anemia – produced clumps of haemoglobin

Summary of Lipogenesis