Lect26 F2021 FA synthesis part2 PDF

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11/14/21

Fatty Acid Synthesis (continued) BCH 442: Mon 11/15/2021 Garrett & Grisham, Chapter 24

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Comparison of b-Oxidation to FFatty atty Acid SSynthesis ynthesis

1) Cellular location, 2) acyl group carrier, 3) electron acceptor/donor, 4) stereochemistry of hydration/dehydration reaction, 5) form of the C2 units.

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The citrate-malatepyruvate shuttle.

Cells Provide Cytosolic Acetyl-CoA and NADPH for Fatty Acid Synthesis

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Acetate Units Are Committed to Fatty Acid Synthesis by Formation of Malonyl-CoA Bicarbonate (form of CO2)

Acetyl-coA (2C)

Malonyl-coA (3C)

The acetyl-CoA carboxylase (ACC) reaction produces malonylCoA for fatty acid synthesis.

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Acetyl-CoA Carboxylase in Animals is a Multifunctional Protein Equilibrium between 2 monomeric/oligomeric forms of ACC Inactive protomers (250kDa)

Active polymer (megaDa) Citrate shifts equilibrium toward active polymer of ACC

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ACC Phosphorylation Modulates Activation by Citrate and Inhibition by Palmitoyl-CoA

The activity of acetyl-CoA carboxylase is modulated by phosphorylation. The dephospho-form (unmodified form) of the enzyme is activated by low [citrate] and inhibited only by high levels of fatty acyl-CoA. In contrast, the phosphorylated enzyme is activated by high levels of citrate.

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Allosteric R Regulation egulation and

Hormone Regulation of Acetyl-CoA Carboxylase

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Malonyl-CoA Regulation of Fatty Acid Uptake into Mitochondria @ CPT-1 (CAT-I)

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Fatty Acid Synthase (FAS) Cytosolic, multi-enzyme complex, Very large Acetate units are attached to growing FA acid chain, 1 unit at a time Requires 2 SH sulfhydryl groups

CoA-like moiety of ACP 9

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Acyl Carrier Proteins Carry the Intermediates in Fatty Acid Synthesis

Fatty acids are conjugated both to coenzyme A and to acyl carrier protein through the sulfhydryl group of phosphopantetheine prosthetic groups. 10

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Palmitate biosynthesis from acetyl-CoA and malonyl-CoA (i) Acetyl & malonyl bldg blocks introduced as ACP-conjugates

(ii) Decarboxylation drives condensation catalyzed by b-ketoacyl-ACP synthase (KSase)

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b -ketoacyl-ACP synthase: also called “KS” b -ketoacyl-ACP reductase: also called “KR” b -hydroxyacyl-ACP dehydratase: also called “DH”

2,3-trans enoyl-ACP reductase: also called “ER” Enzyme activities (5-7) part of the Malonyl-coA/acetyl-coA-ACPtransacylase: also called “MAT”

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FAS in Animals: Head to tail dimer

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Structure of the Fungal Fatty Acid Synthase: closed barrel structure

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Structure of Mammalian Fatty Acid Synthase: Asymmetric X-shaped structure

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The pathway of palmitate synthesis from acetylCoA and malonyl-CoA

malonyl-CoAacetyl-CoA-ACP transacylase (MAT)

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A Mechanism for the Mammalian Ketoacyl-ACP Synthase (KS)

An acetyl group is transferred from CoA to malonyl-CoA-acetyl-CoA-ACP transacylase (MAT) then to the acyl carrier protein (ACP), and then to the ketoacyl-ACP-synthase. Next a malonyl group is tranferred to MAT and then to the acyl carrier protein. C-C bond formation follows.

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Perhaps a clearer view of steps in fatty acid synthesis Condensing enzyme

O O || || + H3C-C-S- KSase OOC-CH2-C-S-ACP malonyl-ACP acetyl-S-KSase repeat

O || H3C-CH2-CH2-C-S-ACP butyryl-ACP

NADPH+ H+

H O | || H3C-C=C-C-S-ACP

O O || || H3C-C-CH2-C-S-ACP acetoacetyl-ACP Reduction

NADPH + H+ NADP+

H2O

O H || | H3C-C-CH2-C-S-ACP Dehydration

HO |

NADP+

Reduction (saturation of C=C double bond)

KSase-SH + CO2

beta-hydroxybutyryl-ACP

H |

crotonyl-ACP

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Recall: FA b -oxidation

Olefin formation

b-keto acid

b-hydroxyl group

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Last step of the 16C fatty acid synthesis: catalyzed by palmitoylthioesterase

Thioesterases Medium-chain FAS: 10, 12, 14 carbon fatty acids

Other enzymes are needed further elongate fatty acids to 18:0, stearic acid and 18:1w9 oleic acid. We can’t make 18:2w6 or 18:3w3, but we can elongate and desaturate—see more later.

Lon chain FAS 16 Fatty acid

We can make 20:4w6, 20:5w3, and 22:6w3 from the 18 carbon precursors and these in turn are made into powerful hormones with many functions.

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C16 Fatty Acids May Undergo Elongation and Unsaturation • Additional elongation - in mitochondria and ER • Introduction of cis double bonds: • Prokaryotes use an O2-independent process • Eukaryotes use an O2-dependent process • Eukaryotes add double bond to middle of the chain - and need the power of O2 to do it

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Elongation of fatty acids in mitochondria is initiated by the thiolase reaction.

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Introduction of single cis C=C double bond • Done by both prokaryotes and eukaryotes • O2-independent pathway for bacteria • vs. O2 dependent in eukaryotes • Fundamental difference between the two: • Can introduce C=C in several places in chain with O2dependent scheme • With O2-independent scheme restricted on where dehydrogenation = desaturation takes place • (needs to be near the b -carbonyl or b-hydroxy group & thioester group at end of chain)

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Double bonds are introduced into the growing fatty acid chain in E. coli by specific dehydrases

10 Carbon chain Introduction of cis C=C double bond in bacteria: Specific dehydrase enzymes

10 Carbon chain with 1 cis C=C bond

16 Carbon chain 18 Carbon chain

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Unsaturation Reactions Occur in Eukaryotes in the Middle of an Aliphatic Chain

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Desaturation reaction may be followed by additional chain elongation • Additional chain elongation following single desaturation step • Oleoyl-CoA produced (18C with C=C at D9) can be elongated by two carbons to form 20:1 cis D11 fatty acyl-coA • If starting fatty acid is C-16 palmitate, similar reaction to produce palmitoyloleoyl-CoA (16:1 cis D9) can be elongated to an 18:1 cis D11 fatty acid (cis-vaccenic acid) • C-16/C-18 fatty acids can be elongated to FA chains with C-22 or C-24 (found in sphingolipids)—sphingolipids are important in structure and signaling 28

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Combinations of desaturase and elongase reactions Essential FA Omega-6

Omega-6 side Essential FA

Omega-3. Animals cannot add C=C beyond D9

Animal cannot change omeg 6 or omeg 3 numbers 29

!3 and !6 – Essential Fatty Acids with Many Functions

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Arachidonic Acid Synthesis in Eukaryotes

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Cytokines and other triggers of inflammation are mediated by eicosanoids, made from arachidonic acid (20:4w6) and are needed to fight infections.

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Regulation of Fatty Acid Synthesis

Regulation of fatty acid synthesis and fatty acid oxidation are coupled as shown. 34

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Hormonal Signals Regulate ACC and Fatty Acid Biosynthesis

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Recall: regulation of ACC carboxylase

Hormone Regulation of Acetyl-CoA Carboxylase

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ACC Phosphorylation Modulates Activation by Citrate and Inhibition by Palmitoyl-CoA

The activity of acetyl-CoA carboxylase is modulated by phosphorylation. The dephospho-form (unmodified form) of the enzyme is activated by low [citrate] and inhibited only by high levels of fatty acyl-CoA. In contrast, the phosphorylated enzyme is activated by high levels of citrate.

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