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Architectures of Mammalian and Fungal Fatty Acid Synthases Presentation based on: T. Maier, S. Jenni, N. Ban, Science 311 , 1258 (2006). -- Mammalian fatty acid at 4.5 Å resolution S. Jenni, M. Leibundgut, T. Maier, N. Ban, Science 311 , 1263 (2006). - PowerPoint PPT Presentation
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Architectures of Mammalian and Fungal Fatty Acid Synthases
Presentation based on:
T. Maier, S. Jenni, N. Ban, Science 311, 1258 (2006). -- Mammalian fatty acid at 4.5 Å resolution
S. Jenni, M. Leibundgut, T. Maier, N. Ban, Science 311, 1263 (2006). -- Fungal fatty acid at 5 Å resolution
Agenda
1. fatty acid quick peak
2. catalytic cycle of fatty acid synthesis
3. mammalian fatty acid synthase structure
4. fungal fatty acid synthase structure
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1. Fatty acid quick peak
Common fatty acids are carboxylic acids with long hydrocarbon tails:
comes with a COOH head and a tail of many CH2.
2. Fatty acid catalytic cycle
2.1 Common elongation scheme
starter substrates Acetyl coenzyme A (Acetyl-CoA) and Malonyl-CoA transfer the active functionals to acyl carrierprotein (ACP).
ACP transports substrate to different reaction sites, catalyzed by different enzyme. A complete cycle gives the acyl group an additional two carbon units.
This step-wise elongation repeats until a substrate length of C16 to C18 is achieved.
Another enzyme then release the substrate from ACP, completing the synthesis process.
2.2 Step-by-step details of catalytic cycle
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: catalysts for different individual reactions
protein that releases the completed product from ACP.
A
B
C
DE
F
2.3 Different fatty acid synthase (FAS) systems
Type II FAS (bacteria) -- all reactions carried out by individual, monofunctional proteins.
Type I FAS (eukaryote) -- large, multifuctional polypeptides contains all necessary enzymes for the enlongation cycle.
Fungal FAS: 2.6-MD 66 dodecamer, catalytic domains distributed over two distinct subunits.
Vertebrates and Mammal FAS: 270-kD 2 homodimer, contains all catalytic activities.
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3. Mammalian fatty acid synthase
3.1 Functional proteins
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ACP: acyl protein carrier
MAT: malonyl-CoA-/acetyl- CoA-ACP-transacylase
KS: -ketoacyl synthase
KR: -ketoacyl reductase
DH: dehydratase
ER: -enoyl reductase
TE: thioesterase
3.2 overall structure and domain assignment
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210Å 180Å 90Å
Blue bubble:Electron density cloud viaX-ray crystallography
Colored proteins:Identified as specific domainvia mapping homologous protein structures
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3.2.1 KS domain
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Mammalian KS closely resembles the Escherichia coli KS I (FabB).So KS domain was fitted with E. coli FabB.
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3.2.2 MAT domain
Mammalian MAT is homologous to bacterial malonyl transferase (FabD).So we fit the MAT domain with Streptomyces coelicolor FabD.
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3.2.3 DH domain
Mammalian DH adopts a “double hot dog” fold that’s closely related to the fold of the dimeric bacterial dehydratases FabA and FabZ. So we fit DH with two monomers of dimeric E. coli FabA.
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3.2.4 ER domain
The best structural match for ER was obtained with a zinc-free bacterial quinone reductase. Here the particular model is quinone reductase of T. thermophilus.
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3.2.5 KR domain
KR belongs to the short-chain dehydrogenase family, and was modeled with E. coli FabG.
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3.2.6 ACP and TE domain
* ACP and TE domains could not be placed with confidence, likely due to their inherent flexibility.
* However…
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This blurred volume of electron density, which was observed only onone side, might be interpreted as arising from the C-terminal ACPand TE domains.
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3.2.7 Table of structural and functional analogs
3.3 intersubunit and interdomain connections
FAS is an intertwined dimer with a large dimerization interface.
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There are other substantialintersubunit contacts in the unassigned region of electrondensity map.
COLORED: identified domains
GREY: unassigned region.
3.4 active sites and the two reaction chambers
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Solid spheres: active sites
Hollow spheres: radii = length of the phosphopantheteine arm of ACP
4. Fungal fatty acid synthase
4.1 Functional proteins
ACP: acyl protein carrier
MPT: malonyl/palmitoyl transferase
KS: ketoacyl synthase
KR: ketoacyl reductase
DH: dehydratase
ER: enoyl reductase
AT: acetyl transferase
PT: phosphopantetheine transferase (for ACP activation)
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4.2 overall structure and domain assignment 230Å 230Å 260Å
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MPT: malonyl/palmitoyl transferase
KS: ketoacyl synthase
KR: ketoacyl reductase
DH: dehydratase
ER: enoyl reductase
AT: acetyl transferase
ACP and PT structures could not be identified.
White regions denote unidentified electron density.
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4.2.1 KS domain
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KS dimer domain was identified by finding the thiolase fold in the FAS electron density map (bacterial KS is known to adopt a thiolase fold and form homodimers).
Bacterial KS homolog fits almost perfectly into the density map.
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4.2.2 KR domain
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The 4-helix bundle is a characteristic trait of one of the dimerization interface in type-II tetrameric KR homolog of Brassica napus.It also contains a Rossmann fold.
The Brassica napus KR homolog fits FASelectron density remarkably well.
4.2.3 DH domain
The closest sequence homolog with known structure of fungal DH is the human Peroxisomal 2-enoyl-CoA hydratase 2 involved in oxidation of fatty acids. It is a pseudo-dimer, has two “hot dog”folds, and forms a large sheet.DH structures of bacterial FAS fits less well, since they are true homodimers.
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4.2.4 ER domain
The fungal ER, unlike other FAS systems,is a FMN-containing oxidoreductase, andno homology has been observed.Since a TIM-barrel fold was discovered nearthe FMN-binding pocket, the 21 known TIM-barrel superfamilies were examined, and a good fit was obtained with spinach glycolate oxidase.
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4.2.5 AT and MPT
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AT
MPT
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AT and MPT are homologous in sequence, catalyze similar reactions, and have same protein fold. Therefore they are both fittedwith malonyl transferase from Streptomycescoelicolor.
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In order to unambiguously assign AT andMPT domain, the locations of their N termini relative to the C terminus of DHwere observed.
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4.3 The reaction chambers
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** Two identical reaction chambers separated by central wheel.
** Each contains three copies of a full set of catalytic domains.
** All active sites are oriented towards interior. Red cones indicate entrances to active sites.
COLORED: fitted domainsGREY: unassigned region.
4.3 The reaction chambers
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A set of active sites in the reaction chamber with allenzymatic activities required for the synthesis cycle.
Green sphere: reaction chamber center.
4.3 The reaction chambers
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Schematic path of ACP, shuttlingsubstrate between the active sites.
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