Fatty acids

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Fatty acids

Oleate C181 Linoleate C182 Linolenate
C183 Arachidonate C204
Myristate C140 PalmitateC160 Stearate C180
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Fatty acids have major biological functions

• Used to synthesize glycerophospholipids and
  sphingolipids
• Covalently modify various proteins
• Fuel molecules that can be broken down to
  generate energy
• Derivatives serve as hormones (prostaglandins)
  and intracellular messengers (DAG, IP3)

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Fatty acids synthesis (FAS)
FAS I Mammals and fungi 1 gene encoding for a
polypeptide (mammals) or 2 polypeptides (lower
eukaryotes-yeasts) containing all the multiple
catalytic activities Cytosolic FAS
II Bacteria, plants, parasites and eukaryotic
organelles of prokaryotic origin Each component
is encoded by a separate gene that catalyze a
single step in the pathway Cytosolic
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Fatty Acid Synthase (FAS)
A multienzyme complexe which contains 7 distinct
catalytic centers.
ACP or acyl carrier protein contains a
pantetheine chain (similar to CoA) long enough
to allow the substrate to reach all seven
catalytic sites.
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Condensation of acetyl ACP and malonyl ACP,
catalyzed by acyl-malonyl ACP condensing enzyme.
This results in the formation of acetoacetyl ACP
Reduction of acetoacetyl ACP
Dehydration
Reduction of crotonyl ACP to butyryl-ACP
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ACP Acyl carrier protein CE condensing enzyme
This cycle is repeated 7 times to give a C16
product palmitoyl-S-ACP (length of the chain is
monitored by the thioesterase TE) Longer chains
are synthesized by elongases
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Trypanosoma brucei
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Bloodstream form Myristoylation (C14) of FA on
GPI anchor Procyclics C16 or C18
VSG 10 millions per cell ? 10 total
proteins Crucial for parasite survival
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Background of fatty acids research in T. brucei
Bloodstream forms were thought not to be able to
synthesize FA de novo Good evidence of FA
uptake Stimulation of host myristate
mobilization Strategies for efficient salvage of
myristate from the bloodstream Finding that T.
brucei can synthesize FA after a study on the
incorporation of radiolabeled laurate (C12) was
shown to be incorporated in palmitate (C16).
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Which system makes fatty acids in Trypanosoma
brucei ?
FA synthesis is based on a type II or an hybrid
form between Type I and II based on the
polypeptidic nature of the predicted proteins
encoded by the genes found in the genome
BUT 1-FA synthesis in T. brucei is membrane
associated- not cytosolic 2-Type II in T. brucei
is localized to the mitochondrion and myristate
is needed in the ER where the GPI remodeling
occurs 3- Does not respond to typical type II
inhibitor (triclosan) 4- RNAi silencing of ACP
has no effect on FA synthesis Bulk FA synthesis
is unconventional involving neither type I or
type II pathway.
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Elongation system
Elongase
The growing acyl chain is esterified to CoA
instead of ACP
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T. brucei 4 putative elongase (ELO)
genes ELO1-3 Chromosome 7 (tandem) ELO4-
Chromosome 5
Techniques used Silencing by RNAi Knock outs of
the 4 elongases Experiments are conducted on
membrane preparations or on whole parasites
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Experiment done on PCF
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Fig.3
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Sequential fatty acid synthesis by ELO1-3
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Notes on WT -Robust synthesis with primers
between C4 and C12 -C4 and C12 primers themselves
were about ½ as efficient as intermediate
primers -C4 elongation is mostly to C14 with
little intermediates while C6 and C8 shows
accumulation of intermediates
Sequential fatty acid synthesis by ELO1-3
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WT parasites membranes C6-CoA, C16-CoA
Fig.5
0 saturated FA 1 mono-unsaturated 2
di-unsaturated 3 tri-unsaturated 4
tetra-unsaturated o origin
Mostly saturated FA are being produced only
traces of unsaturated
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Fatty acid synthesis in cultured parasites
Fig.6
Elo1-3 share the same EnCR (RNAi inhibition
incomplete)
ELO pathway is responsible for the majority of FA
synth. ELO pathway appears to be regulated (FA
synthesis increases in low-lipid medium)
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Other kinetoplastids
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• Efficient synthesis from butyryl-CoA
• but not from Acetyl-CoA
• Synthesis of FA more robust in T. cruzi
• than Leishmania
• Products ranging from C8 to C18

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Summary
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T. brucei
ER associated Pathway essential in PCF Not
essential in BSF
FAS II pathway is mitochondrial and produces at
low level
Toxoplasma gondii FAS II appears to be essential
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What are the advantages of using the elongase
pathway?

• Synthesis is at the site of product utilization
• The ELO pathway is very versatile and can be
  regulated
• to adapt to the parasites needs when migrating
  from vector
• to host or depending on the stage of the life
  cycle