Fatty acid biosynthesis

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Fatty acid biosynthesis

• The overall reaction for synthesis of palmitate
  is8 acetyl-CoA 7 ATP 14 NADPH 14 H
  ? CH3(CH2)14CO2- 7 ADP 7 Pi 14 NADP 7
  H2O
• Fatty acid synthesis is not the reverse of
  degradation, in terms of mechanism.
• Moreover, as you saw in carbohydrate synthesis,
  it takes more energy to make palmitate from 8
  acetyl-CoA than the energy derived from
  ?-oxidation of palmitate to 8 acetyl-CoA.

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Entry into the pathway

• This enzyme catalyzesacetyl-CoA HCO3- ATP ?
  malonyl-CoA ADP Pi
• Three enzymes carry this out
• biotin carrier protein (BCP) carries biotin
• biotin carboxylaseuses ATP to carboxylate biotin
  on BCP
• Transcarboxylase transfers carboxylate from
  biotin to acetyl-CoA
• This step is the one that is regulated, and the
  point at which energy is input into the system.
• The enzyme is analogous to pyruvate carboxylase,
  used to make PEP from pyruvate. The logic is
  similar as well. Acetyl-CoA is carboxylated
  transiently to drive its condensation onto the
  growing acyl chain.

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Entry into the pathway

• This enzyme catalyzesacetyl-CoA HCO3- ATP ?
  malonyl-CoA ADP Pi
• Three enzymes carry this out
• biotin carrier protein (BCP) carries biotin
• biotin carboxylaseuses ATP to carboxylate biotin
  on BCP
• Transcarboxylase transfers carboxylate from
  biotin to acetyl-CoA
• This step is the one that is regulated, and the
  point at which energy is input into the system.
• The enzyme is analogous to pyruvate carboxylase,
  used to make PEP from pyruvate. The logic is
  similar as well. Acetyl-CoA is carboxylated
  transiently to drive its condensation onto the
  growing acyl chain.

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Entry into the pathway

• This step is the one that is regulated, and the
  point at which energy is input into the system.
• The enzyme is analogous to pyruvate carboxylase,
  used to make PEP from pyruvate.
• The logic is similar as well. Acetyl-CoA is
  carboxylated transiently to drive its
  condensation onto the growing acyl chain.

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Fatty acid synthase7 reactions for 7 enzymes

• This is a large multidomain enzyme, which
  catalyzes 7 reactions (one for each
  domain/subunit)
• Acetyl-CoA-ACP transacetylase The acetyl group
  of acetyl-CoA is transferred to a cysteine thiol
  on the ?-ketoacyl-ACP synthase domain.
• Malonyl-CoA-ACP transferase The malonyl group of
  malonyl-CoA is transferred to ACP (acyl carrier
  protein), to which it is attached via
  phosphopantetheine (i.e. same linkage as in CoA).
  All reactions are carried out on acyl chains
  attached to ACP, which is at the core of the FA
  synthase.

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Fatty acid synthase7 reactions for 7 enzymes

• ?-ketoacyl-ACP synthase acetyl-ACP and
  malonyl-ACP condense, releasing CO2, to form
  ?-ketoacyl-ACP. The acyl chain is now on ACP,
  where it will stay for the remainder of the
  reactions in this cycle.
• Note that the 2 carbons from acetyl-CoA will be
  the last 2 carbons of the fatty acid.
• Also note that the carboxylate added to
  acetyl-CoA to make malonyl-CoA is the one that is
  released as CO2.
• This decarboxylation helps drive the reaction
  forward, which would otherwise be entropically
  quite unfavorable.

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Fatty acid synthase7 reactions for 7 enzymes

• ?-ketoacyl-ACP reductase Using NADPH, the ketone
  is reduced to a hydroxyl. (Note that the carbon
  bearing the hydroxyl is chiral this compound is
  in the D-configuration.)
• ?-hydroxyacyl-ACP dehydrase Water is eliminated,
  making a trans-?2-enoyl-ACP.
• enoyl-ACP reductase Using NADPH, the double bond
  is saturated.

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Fatty acid synthase7 reactions for 7 enzymes

• Now you have an acyl-ACP.
• This is transferred to the thiol of
  ?-ketoacyl-ACP synthase by the transacetylase
  (reaction 1), where it will then react with
  another molecule of malonyl-ACP, starting a new
  cycle.
• Then the same 3 reduction/dehydration/reduction
  reactions will occur.
• Seven iterations of this cycle will give a
  palmitoyl-ACP.
• thioesterase Palmitate is released from the
  enzyme by hydrolysis of palmitoyl-ACP. The enzyme
  is rather specific for C16.

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Fatty acid synthase7 reactions for 7 enzymes

• Fatty acid synthesis is not exactly the reverse
  of degradation. There are several key differences
  among them are

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Elongation of fatty acids

• Fatty acid elongation systems utilize similar
  mechanisms as fatty acid synthase to make fatty
  acids with gt16 carbons.
• The main difference is the location (smooth ER in
  animals) and the fact that it takes place on
  acyl-CoA, rather than acyl-ACP.
• Malonyl-CoA is still used as the acetyl donor,
  followed by reduction, dehydration, and reduction
  by NADPH.

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Desaturation of fatty acids

• Anaerobic pathway (certain prokaryotes)
• At the 10-C stage, a new enzyme
  (?-hydroxydecanoyl-ACP-dehydrase) dehydrates and
  then isomerizes the trans-?2-enoyl-ACP to
  cis-?3-enoyl-ACP.
• After completion of fatty acid synthesis, the
  end product will be palmitoleic acid (161?9).

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Desaturation of fatty acids

• Aerobic pathway (eukaryotes)
• C16 and C18 acyl-CoAs are desaturated by acyl-CoA
  desaturase, using O2 and electrons from 2 cyt b5.
  The electrons originate ultimately from NADPH
  cytochrome b5 reductase uses flavin as an adapter
  to catalyzeNADPH 2 cyt b5(Fe3) gt NADP H
  2 cyt b5(Fe2)
• Cyt b5 is used to reduce and thus activate
  molecular oxygen, which then oxidizes the acyl
  chain.
• This is a 4 e- reduction/oxidation
• 2 come from NADPH via cyt b5 ("activation
  investment")
• 2 come from the acyl chain
• 2 cyt b5(Fe2) gt 2 cyt b5(Fe3) 2 e-
• stearoyl-CoA gt oleoyl-CoA 2 H 2 e-
• O2 4 H 4 e- gt 2 H2O

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Desaturation of fatty acids

• Thus, the anaerobic pathway introduces the double
  bond during FA synthesis, while the aerobic
  pathway does it after the FA is made.
• This difference has to do partly with the
  different organization of FA synthase
• The multi-subunit enzyme in E.coli, in which each
  activity resides on a separate polypeptide,
  allows substitution of alternate subunits, such
  as 3-ketoacyl-ACP synthase II.
• This is not possible with the mammalian enzyme,
  where these activities are domains of one large
  polypeptide.
• It also has to do with the fact that many
  bacteria can live in anaerobic environments, and
  thus need an O2-independent way to do this.
  (Mammals in an anaerobic environment have bigger
  things to worry about!)

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Triacylglycerol synthesis

• Creation of backbone (glycerol-phosphate) in one
  of 2 ways
• Glycerol kinaseglycerol ATP ? glycerol-3-Pi
  ADP
• Glycerol-3-Pi dehydrogenaseDHAP NADH H ?
  glycerol-3-Pi NAD(These are the same enzymes
  used to degrade glycerol.)

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Triacylglycerol synthesis

• Then acyl transferase use acyl-CoA as a substrate
  to make the ester linkages to glycerol
• glycerol-3-Pi acyl-CoA ? acylglycerol-3-Pi
  CoA
• acylglycerol-3-Pi acyl-CoA ? diacylglycerol-3
  -Pi CoA
• Diacylglycerol-3-Pi (phosphatidic acid) is the
  precursor to triacylglycerols and
  glycerolphospholipids.

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Triacylglycerol synthesis

• To make triacylglycerols, it must first undergo
  phosphate removal by phosphatidic acid
  phosphatase
• diacylglycerol-3-Pi H2O ? diacylglycerol Pi
• Then acyl transferase can act at the C3 position
• diacylglycerol acyl-CoA ? triacylglycerol CoA

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Glycerophospholipid synthesis

• The first 2 stages (backbone synthesis and fatty
  acid attachment) have already been shown.
• The next stage is headgroup attachment.
• This involves the formation of a phosphodiester
  linkage 2 alcohols are linked through a
  phosphate.
• There are 3 main ways to do this.

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Glycerophospholipid synthesis

• Activate the backbone. This is accomplished by
  attachment to a cytidine nucleotide
• diacylglycerol-3-Pi CTP ? CDP-DAG PPI
• CDP-DAG headgroup-OH ? headgroup-O-Pi-DAG
  CMP

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Glycerophospholipid synthesis

• Activate the headgroup. This is also
  accomplished by attachment to a cytidine
  nucleotide
• headgroup-O-Pi CTP ? CDP-headgroup PPI
• CDP-headgroup diacylglycerol-3-Pi ?
  headgroup-O-Pi-DAG CMP

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Glycerophospholipid synthesis

• Either way, the overall reaction ofDAG-3-Pi
  headgroup-OH ? headgroup-O-Pi-DAGis coupled to
  CTP ? CMP PPi.
• Using CDP-DAG is most common, especially in
  bacteria.

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Glycerophospholipid synthesis

• Exchange headgroups. The energy invested in the
  phosphoester bond is conserved by a
  transesterification reaction
• headgroup1-O-Pi-DAG headgroup2-OH ?
  headgroup2-O-Pi-DAG headgroup1-OH

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