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- EMDB-24868: CryoEM structure of modular PKS holo-Lsd14 stalled at the condens... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-24868 | ||||||||||||
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Title | CryoEM structure of modular PKS holo-Lsd14 stalled at the condensation step and bound to antibody fragment 1B2, composite structure | ||||||||||||
![]() | Composite map | ||||||||||||
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Function / homology | ![]() 6-deoxyerythronolide-B synthase / erythronolide synthase activity / macrolide biosynthetic process / DIM/DIP cell wall layer assembly / fatty acid synthase activity / phosphopantetheine binding / 3-oxoacyl-[acyl-carrier-protein] synthase activity / fatty acid biosynthetic process / oxidoreductase activity / plasma membrane / cytoplasm Similarity search - Function | ||||||||||||
Biological species | ![]() ![]() | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.1 Å | ||||||||||||
![]() | Bagde SR / Kim C-Y / Fromme JC | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Modular polyketide synthase contains two reaction chambers that operate asynchronously. Authors: Saket R Bagde / Irimpan I Mathews / J Christopher Fromme / Chu-Young Kim / ![]() Abstract: Type I modular polyketide synthases are homodimeric multidomain assembly line enzymes that synthesize a variety of polyketide natural products by performing polyketide chain extension and β-keto ...Type I modular polyketide synthases are homodimeric multidomain assembly line enzymes that synthesize a variety of polyketide natural products by performing polyketide chain extension and β-keto group modification reactions. We determined the 2.4-angstrom-resolution x-ray crystal structure and the 3.1-angstrom-resolution cryo–electron microscopy structure of the Lsd14 polyketide synthase, stalled at the transacylation and condensation steps, respectively. These structures revealed how the constituent domains are positioned relative to each other, how they rearrange depending on the step in the reaction cycle, and the specific interactions formed between the domains. Like the evolutionarily related mammalian fatty acid synthase, Lsd14 contains two reaction chambers, but only one chamber in Lsd14 has the full complement of catalytic domains, indicating that only one chamber produces the polyketide product at any given time. | ||||||||||||
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 | ![]() | 145.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 24.4 KB 24.4 KB | Display Display | ![]() |
Images | ![]() | 109 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7s6cMC ![]() 7s6bC ![]() 7s6dC C: citing same article ( M: atomic model generated by this map |
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Similar 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
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Composite map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.043 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : Dimeric holo-Lsd14 containing docking domains from DEBS module 3 ...
Entire | Name: Dimeric holo-Lsd14 containing docking domains from DEBS module 3 (holo-Lsd14-DD*) bound to two copies each of Fab 1B2 light and heavy chains and NADP, treated with 2-Acetaminoethyl-thio-3-oxobutanoate |
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Components |
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-Supramolecule #1: Dimeric holo-Lsd14 containing docking domains from DEBS module 3 ...
Supramolecule | Name: Dimeric holo-Lsd14 containing docking domains from DEBS module 3 (holo-Lsd14-DD*) bound to two copies each of Fab 1B2 light and heavy chains and NADP, treated with 2-Acetaminoethyl-thio-3-oxobutanoate type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Molecular weight | Theoretical: 450 KDa |
-Supramolecule #2: Polyketide synthase
Supramolecule | Name: Polyketide synthase / type: complex / ID: 2 / Parent: 1 |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Supramolecule #3: Fab 1B2 heavy chain, Fab 1B2 light chain
Supramolecule | Name: Fab 1B2 heavy chain, Fab 1B2 light chain / type: complex / ID: 3 / Parent: 1 |
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Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #1: 6-deoxyerythronolide-B synthase EryA2, modules 3 and 4, Lsd14 Pol...
Macromolecule | Name: 6-deoxyerythronolide-B synthase EryA2, modules 3 and 4, Lsd14 Polyketide synthase fusion type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: 6-deoxyerythronolide-B synthase |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 174.085734 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MVTDSEKVAE YLRRATLDLR AARQRIRELE SEPIAIIGMA CRLPGGVDSP EGLWELVDSG TDAIAGFPLD RGWDVEGMYD PDAEAPGKT YVKEAGFLYD AGEFDAGFFG ISPREAVSMD PQQRLMLEAS WEAFERAGLD PARQRGTATG VFVGATATGY V SPAAEVPE ...String: MVTDSEKVAE YLRRATLDLR AARQRIRELE SEPIAIIGMA CRLPGGVDSP EGLWELVDSG TDAIAGFPLD RGWDVEGMYD PDAEAPGKT YVKEAGFLYD AGEFDAGFFG ISPREAVSMD PQQRLMLEAS WEAFERAGLD PARQRGTATG VFVGATATGY V SPAAEVPE GAEGFAITGN MTAVTSGRIS YTLGLQGPAV TIDTACSSSL VALHLACQSL RQGECTTALA GGVTVMPTPT AF TEFSRQR GLAPDGRCKS FAAAADGTNW AEGVAVLVVE RLSDARRNGH RVLAVVRGTA INQDGASNGL SAPNDLAQER VIR SALDNA GLTASDVDAV EAHGTGTTLG DPIEAQALLA AYGHERPAHR PLRVGSLKSN IGHAGPAAGV AGVIKMVMAM RHGV LPRSL HIDEPTPQVD WSSGAVTLLT EPVDWPDSDR PRRAGVSAFG ISGTNAHVIL EQAPTQDPQQ PAPPVPAAPW LLSAK TPAA LRAQARRLHT HLARHPHPDP TDIAHALATT RTPHEHRAAL VTDDHGTRGP ALAALAEGAP DACLISGTAL SKGRTV FVF PGQGSQWTGM GRELLHTSPE FAAYIAECET ALNDFVDWSL TDVLRGTEGA PGYDRVDVVQ PALFAVMVSL ARLWQHH GI HPDAVIGHSQ GEIAAAHIAG ALSLQDAARI VALRSQALLP LAGLGGMTSL ALPHDQALQL IQPWGQDLSI ASVNGPHS T VVSGTTHALD ELHTTCDTQG VRARRIPVDY ASHSAQVESI RDTVLQAATG INPQPTTIPL YSTVTGQPID GTQLDADYW YTNLRHTVRF EETTRALLGS GHRHFIETTA HPVLALALEE TIEATGSDAR VTGTLRRDHG DLTQLHTALA TAWTHGIDVD WTAVLGDRR TPFELPTYAF QRQRYWLEPG AGVGVPVGGT SAEARFWDAV EDEDLEALVA AIGADGDDAS WAGVLPALAG W RRRQREQS ALDDLRYKVT WKPTAVADGA SATGTWLVVV PESLAGGGWP VVVARAVDQA GGRPVVLSVD AADGADRSRL GL RIHEALG EGPVPDAVVS LLALDPSALP GLPDVPQALA STAALVQALL DLGLEARLWC VTSGAVSVSG ADGPSAPEQA AVW GFGRVA GLEHPHLWAG LVDLPPEADE RTAARLVGVL AGAGGEDQVA LRSSGVFVRR LVRAPASEVP AVRSWKPGGT VLVT GGTGG LGRQVARWLA RGGADHLLLV SRRGVDAPGA DELVDELTDL GARVTVAACD VADRDAVQRL LSEQVPSDAP LTAVI HTAA VLDDGVIDSL SPERMEQVLR VKVGGAVHLY ELTRESDLSA FVLFSSFGST FGLPGLGNYA PGNAALEALA EQWRAE GRP ATAVGWGTWA GGGMADGGVG ERGRTHGIHE LEPALATAAL EQALERDESS PVIIDIDWER FAVAFHAKRP TRGFELV PE AQAALEAADG GPGPDGGAGD PSELLDRLAA LPDAERDRAL LEVVRGNAAS VMSHGAMRTA TLEAVEPTRA FRDLGFDS L MAVELRNRIG AATGLRLAPT LVFDHPTPEA VVRHLRAELG LEGDGAPDPV FDELDGLERA LSSYTPDTDT RVKITKRLE SLLWEWTRSE ADAPDPVDAA DLAAVSDDEM FELIDRELGS ALEHHHHHH |
-Macromolecule #2: Fab 1B2 heavy chain
Macromolecule | Name: Fab 1B2 heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 26.447611 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MAEVQLVQSG GGLVQPGRSL RLSCTASGFT FGDYAMSWVR QAPGKGLEWV GFIRSKAYGG TTEYAASVKG RFTISRDDSK SIAYLQMNS LKTEDTAVYY CTRGGTLFDY WGQGTLVTVS SASTKGPSVF PLAPSSKSTS GGTAALGCLV KDYFPEPVTV S WNSGALTS ...String: MAEVQLVQSG GGLVQPGRSL RLSCTASGFT FGDYAMSWVR QAPGKGLEWV GFIRSKAYGG TTEYAASVKG RFTISRDDSK SIAYLQMNS LKTEDTAVYY CTRGGTLFDY WGQGTLVTVS SASTKGPSVF PLAPSSKSTS GGTAALGCLV KDYFPEPVTV S WNSGALTS GVHTFPAVLQ SSGLYSLSSV VTVPSSSLGT QTYICNVNHK PSNTKVDKKV EPKSCAALVP RGSAHHHHHH AA DYKDDDD KA |
-Macromolecule #3: Fab 1B2 light chain
Macromolecule | Name: Fab 1B2 light chain / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 25.715832 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: LFAIPLVVPF YSHSALDVVM TQSPLSLPVT PGEPASISCR SSQSLLHSNG YNYLDWYLQK PGQSPQLLIY LGSNRASGVP DRFSGSGSG TDFTLKISRV EAEDVGVYYC MQSLQTPRLT FGPGTKVDIK RTVAAPSVFI FPPSDEQLKS GTASVVCLLN N FYPRGAKV ...String: LFAIPLVVPF YSHSALDVVM TQSPLSLPVT PGEPASISCR SSQSLLHSNG YNYLDWYLQK PGQSPQLLIY LGSNRASGVP DRFSGSGSG TDFTLKISRV EAEDVGVYYC MQSLQTPRLT FGPGTKVDIK RTVAAPSVFI FPPSDEQLKS GTASVVCLLN N FYPRGAKV QWKVDNALQS GNSQESVTEQ DSKDSTYSLS STLTLSKADY EKHKVYACEV THQGLSSPVT KSFNRGEC |
-Macromolecule #4: 2'-MONOPHOSPHOADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: 2'-MONOPHOSPHOADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 1 / Formula: ATR |
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Molecular weight | Theoretical: 507.181 Da |
Chemical component information | ![]() ChemComp-ATR: |
-Macromolecule #5: 4'-PHOSPHOPANTETHEINE
Macromolecule | Name: 4'-PHOSPHOPANTETHEINE / type: ligand / ID: 5 / Number of copies: 1 / Formula: PNS |
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Molecular weight | Theoretical: 358.348 Da |
Chemical component information | ![]() ChemComp-PNS: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 4.2 mg/mL | ||||||||||||||||
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Buffer | pH: 7.2 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 120.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa | ||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV Details: Blot for 5 seconds before plunging in liquid ethane.. | ||||||||||||||||
Details | Size exclusion chromatography purified monodisperse holo-Lsd14-DD* + 1B2 complex was first incubated with KS substrate analog (2-Acetaminoethyl-thio-3-oxobutanoate) and TCEP at 30C for 20 minutes, followed by addition of NADP and methylmalonyl-CoA. The sample was further incubated at 30C for 15 minutes before applying to cryoEM grids. |
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Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 1278 / Average exposure time: 2.2 sec. / Average electron dose: 53.0 e/Å2 / Details: Images were collected as 50 frame movies. |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.7 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 79000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model |
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Details | Model was manually rebuilt in Coot and phenix.real_space_refine was used for refinement. | ||||||||
Refinement | Space: REAL / Protocol: OTHER | ||||||||
Output model | ![]() PDB-7s6c: |