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- EMDB-22488: Cryo-EM structure of Grafix-stabilized PaFS prenyltransferase oct... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-22488 | |||||||||
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Title | Cryo-EM structure of Grafix-stabilized PaFS prenyltransferase octamer with a peripheral cyclase domain - symmetry expanded class C | |||||||||
![]() | Final reconstruction Symmetry Expanded Class C | |||||||||
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Function / homology | ![]() fusicocca-2,10(14)-diene synthase / alcohol biosynthetic process / mycotoxin biosynthetic process / ketone biosynthetic process / geranylgeranyl diphosphate synthase / farnesyltranstransferase activity / isoprenoid biosynthetic process / lyase activity / metal ion binding Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 9.4 Å | |||||||||
![]() | Faylo JL / van Eeuwen T / Murakami K / Christianson DW | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural insight on assembly-line catalysis in terpene biosynthesis. Authors: Jacque L Faylo / Trevor van Eeuwen / Hee Jong Kim / Jose J Gorbea Colón / Benjamin A Garcia / Kenji Murakami / David W Christianson / ![]() Abstract: Fusicoccadiene synthase from Phomopsis amygdali (PaFS) is a unique bifunctional terpenoid synthase that catalyzes the first two steps in the biosynthesis of the diterpene glycoside Fusicoccin A, a ...Fusicoccadiene synthase from Phomopsis amygdali (PaFS) is a unique bifunctional terpenoid synthase that catalyzes the first two steps in the biosynthesis of the diterpene glycoside Fusicoccin A, a mediator of 14-3-3 protein interactions. The prenyltransferase domain of PaFS generates geranylgeranyl diphosphate, which the cyclase domain then utilizes to generate fusicoccadiene, the tricyclic hydrocarbon skeleton of Fusicoccin A. Here, we use cryo-electron microscopy to show that the structure of full-length PaFS consists of a central octameric core of prenyltransferase domains, with the eight cyclase domains radiating outward via flexible linker segments in variable splayed-out positions. Cryo-electron microscopy and chemical crosslinking experiments additionally show that compact conformations can be achieved in which cyclase domains are more closely associated with the prenyltransferase core. This structural analysis provides a framework for understanding substrate channeling, since most of the geranylgeranyl diphosphate generated by the prenyltransferase domains remains on the enzyme for cyclization to form fusicoccadiene. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 1.8 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 17.6 KB 17.6 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 7 KB | Display | ![]() |
Images | ![]() | 50.4 KB | ||
Masks | ![]() ![]() | 27 MB 27 MB | ![]() | |
Others | ![]() ![]() | 20.7 MB 20.7 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 444.1 KB | Display | ![]() |
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Full document | ![]() | 443.6 KB | Display | |
Data in XML | ![]() | 13.6 KB | Display | |
Data in CIF | ![]() | 17.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Final reconstruction Symmetry Expanded Class C | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.16 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
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Density Histograms |
-Mask #2
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-Half map: #2
File | emd_22488_half_map_1.map | ||||||||||||
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Density Histograms |
-Half map: #1
File | emd_22488_half_map_2.map | ||||||||||||
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Sample components
-Entire : Phomopsis amygdali fusicoccadiene synthase (PaFS)
Entire | Name: Phomopsis amygdali fusicoccadiene synthase (PaFS) |
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Components |
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-Supramolecule #1: Phomopsis amygdali fusicoccadiene synthase (PaFS)
Supramolecule | Name: Phomopsis amygdali fusicoccadiene synthase (PaFS) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all Details: PaFS octameric prenyltransferase domain is well-resolved in structure, with some density for a variably positioned cyclase domain alongside the octamer. Molecular weight reflects the ...Details: PaFS octameric prenyltransferase domain is well-resolved in structure, with some density for a variably positioned cyclase domain alongside the octamer. Molecular weight reflects the calculated molecular weight of a prenyltransferase octamer plus one cyclase domain. |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() ![]() |
Molecular weight | Theoretical: 430 KDa |
-Macromolecule #1: Fusicoccadiene synthase
Macromolecule | Name: Fusicoccadiene synthase / type: protein_or_peptide / ID: 1 Details: The final reconstruction accounts for an octamer of the C-terminal transferase domain, residues 414-719. Density on the periphery of the octamer accounts for the N-terminal cyclase domain Enantiomer: LEVO / EC number: fusicocca-2,10(14)-diene synthase |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MGSSHHHHHH SSGLVPRGSH MEFKYSEVVE PSTYYTEGLC EGIDVRKSKF TTLEDRGAIR AHEDWNKHIG PCREYRGTLG PRFSFISVAV PECIPERLEV ISYANEFAFL HDDVTDHVGH DTGEVENDEM MTVFLEAAHT GAIDTSNKVD IRRAGKKRIQ SQLFLEMLAI ...String: MGSSHHHHHH SSGLVPRGSH MEFKYSEVVE PSTYYTEGLC EGIDVRKSKF TTLEDRGAIR AHEDWNKHIG PCREYRGTLG PRFSFISVAV PECIPERLEV ISYANEFAFL HDDVTDHVGH DTGEVENDEM MTVFLEAAHT GAIDTSNKVD IRRAGKKRIQ SQLFLEMLAI DPECAKTTMK SWARFVEVGS SRQHETRFVE LAKYIPYRIM DVGEMFWFGL VTFGLGLHIP DHELELCREL MANAWIAVGL QNDIWSWPKE RDAATLHGKD HVVNAIWVLM QEHQTDVDGA MQICRKLIVE YVAKYLEVIE ATKNDESISL DLRKYLDAML YSISGNVVWS LECPRYNPDV SFNKTQLEWM RQGLPSLESC PVLARSPEID SDESAVSPTA DESDSTEDSL GSGSRQDSSL STGLSLSPVH SNEGKDLQRV DTDHIFFEKA VLEAPYDYIA SMPSKGVRDQ FIDALNDWLR VPDVKVGKIK DAVRVLHNSS LLLDDFQDNS PLRRGKPSTH NIFGSAQTVN TATYSIIKAI GQIMEFSAGE SVQEVMNSIM ILFQGQAMDL FWTYNGHVPS EEEYYRMIDQ KTGQLFSIAT SLLLNAADNE IPRTKIQSCL HRLTRLLGRC FQICDDYQNL VSADYTKQKG FCEDLDEGKW SLALIHMIHK QRSHMALLNV LSTGRKHGGM TLEQKQFVLD IIEEEKSLDY TRSVMMDLHV QLRAEIGRIE ILLDSPNPAM RLLLELLRV |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.5 |
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Grid | Details: unspecified |
Vitrification | Cryogen name: ETHANE / Instrument: LEICA EM CPC / Details: Blot for 2.5 seconds before plunging. |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 6544 / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Refinement | Protocol: AB INITIO MODEL |
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