+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13583 | |||||||||
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Title | Structure of SidJ/CaM bound to SdeA in pre-glutamylation state | |||||||||
Map data | ||||||||||
Sample |
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Function / homology | Function and homology information Ligases / NAD+-protein-arginine ADP-ribosyltransferase / negative regulation of calcium ion transmembrane transporter activity / deNEDDylase activity / NAD+-protein-arginine ADP-ribosyltransferase activity / protein deneddylation / Transferases; Acyltransferases; Aminoacyltransferases / protein deubiquitination / K63-linked deubiquitinase activity / positive regulation of cyclic-nucleotide phosphodiesterase activity ...Ligases / NAD+-protein-arginine ADP-ribosyltransferase / negative regulation of calcium ion transmembrane transporter activity / deNEDDylase activity / NAD+-protein-arginine ADP-ribosyltransferase activity / protein deneddylation / Transferases; Acyltransferases; Aminoacyltransferases / protein deubiquitination / K63-linked deubiquitinase activity / positive regulation of cyclic-nucleotide phosphodiesterase activity / negative regulation of calcium ion export across plasma membrane / positive regulation of ryanodine-sensitive calcium-release channel activity / regulation of cell communication by electrical coupling involved in cardiac conduction / negative regulation of peptidyl-threonine phosphorylation / ligase activity / negative regulation of ryanodine-sensitive calcium-release channel activity / protein phosphatase activator activity / : / adenylate cyclase binding / catalytic complex / detection of calcium ion / regulation of cardiac muscle contraction / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / calcium channel inhibitor activity / regulation of release of sequestered calcium ion into cytosol by sarcoplasmic reticulum / cysteine-type peptidase activity / : / titin binding / regulation of calcium-mediated signaling / positive regulation of protein autophosphorylation / voltage-gated potassium channel complex / sperm midpiece / calcium channel complex / substantia nigra development / adenylate cyclase activator activity / regulation of heart rate / nucleotidyltransferase activity / sarcomere / protein serine/threonine kinase activator activity / regulation of cytokinesis / positive regulation of peptidyl-threonine phosphorylation / spindle microtubule / response to calcium ion / positive regulation of protein serine/threonine kinase activity / G2/M transition of mitotic cell cycle / spindle pole / calcium-dependent protein binding / host cell / myelin sheath / transferase activity / vesicle / transmembrane transporter binding / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / protein ubiquitination / G protein-coupled receptor signaling pathway / centrosome / nucleotide binding / calcium ion binding / protein kinase binding / protein-containing complex / proteolysis / extracellular region / membrane / nucleus / metal ion binding / plasma membrane / cytoplasm Similarity search - Function | |||||||||
Biological species | Legionella pneumophila (bacteria) / Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.91 Å | |||||||||
Authors | Adams M / Bhogaraju S | |||||||||
Funding support | European Union, 1 items
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Citation | Journal: Nat Commun / Year: 2021 Title: Structural basis for protein glutamylation by the Legionella pseudokinase SidJ. Authors: Michael Adams / Rahul Sharma / Thomas Colby / Felix Weis / Ivan Matic / Sagar Bhogaraju / Abstract: Legionella pneumophila (LP) avoids phagocytosis by secreting nearly 300 effector proteins into the host cytosol. SidE family of effectors (SdeA, SdeB, SdeC and SidE) employ phosphoribosyl ...Legionella pneumophila (LP) avoids phagocytosis by secreting nearly 300 effector proteins into the host cytosol. SidE family of effectors (SdeA, SdeB, SdeC and SidE) employ phosphoribosyl ubiquitination to target multiple host Rab GTPases and innate immune factors. To suppress the deleterious toxicity of SidE enzymes in a timely manner, LP employs a metaeffector named SidJ. Upon activation by host Calmodulin (CaM), SidJ executes an ATP-dependent glutamylation to modify the catalytic residue Glu860 in the mono-ADP-ribosyl transferase (mART) domain of SdeA. SidJ is a unique glutamylase that adopts a kinase-like fold but contains two nucleotide-binding pockets. There is a lack of consensus about the substrate recognition and catalytic mechanism of SidJ. Here, we determined the cryo-EM structure of SidJ in complex with its substrate SdeA in two different states of catalysis. Our structures reveal that both phosphodiesterase (PDE) and mART domains of SdeA make extensive contacts with SidJ. In the pre-glutamylation state structure of the SidJ-SdeA complex, adenylylated E860 of SdeA is inserted into the non-canonical (migrated) nucleotide-binding pocket of SidJ. Structure-based mutational analysis indicates that SidJ employs its migrated pocket for the glutamylation of SdeA. Finally, using mass spectrometry, we identified several transient autoAMPylation sites close to both the catalytic pockets of SidJ. Our data provide unique insights into the substrate recognition and the mechanism of protein glutamylation by the pseudokinase SidJ. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_13583.map.gz | 51.9 MB | EMDB map data format | |
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Header (meta data) | emd-13583-v30.xml emd-13583.xml | 13.6 KB 13.6 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_13583_fsc.xml | 10 KB | Display | FSC data file |
Images | emd_13583.png | 60 KB | ||
Masks | emd_13583_msk_1.map | 83.7 MB | Mask map | |
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13583 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13583 | HTTPS FTP |
-Validation report
Summary document | emd_13583_validation.pdf.gz | 353.4 KB | Display | EMDB validaton report |
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Full document | emd_13583_full_validation.pdf.gz | 353 KB | Display | |
Data in XML | emd_13583_validation.xml.gz | 11.5 KB | Display | |
Data in CIF | emd_13583_validation.cif.gz | 15.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13583 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13583 | HTTPS FTP |
-Related structure data
Related structure data | 7ppoMC 7pqeC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_13583.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.008 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_13583_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : SidJ/CaM-SdeA
Entire | Name: SidJ/CaM-SdeA |
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Components |
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-Supramolecule #1: SidJ/CaM-SdeA
Supramolecule | Name: SidJ/CaM-SdeA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: Legionella pneumophila (bacteria) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
-Macromolecule #1: Ubiquitinating/deubiquitinating enzyme SdeA
Macromolecule | Name: Ubiquitinating/deubiquitinating enzyme SdeA / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO EC number: Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases |
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Source (natural) | Organism: Legionella pneumophila (bacteria) |
Molecular weight | Theoretical: 111.200375 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: HHHHHHSAGL EVLFQGPMVG FSLYTDDTVK AAAQYAYDNY LGKPYTGSVE SAPANFGGRM VYRQHHGLSH TLRTMAYAEL IVEEARKAK LRGETLGKFK DGRTIADVTP QELKKIMIAQ AFFVAGRDDE ASDAKNYQKY HEQSRDAFLK YVKDNESTLI P DVFKDQED ...String: HHHHHHSAGL EVLFQGPMVG FSLYTDDTVK AAAQYAYDNY LGKPYTGSVE SAPANFGGRM VYRQHHGLSH TLRTMAYAEL IVEEARKAK LRGETLGKFK DGRTIADVTP QELKKIMIAQ AFFVAGRDDE ASDAKNYQKY HEQSRDAFLK YVKDNESTLI P DVFKDQED VNFYARVIED KSHDWESTPA HVLINQGHMV DLVRVKQPPE SFLQRYFSSM QRWIGSQATE AVFGIQRQFF HA TYEVVAG FDSDNKEPHL VVSGLGRYVI GEDGQPIREA PKKGQKEGDL KVFPQTYKLK ENERLMRVDE FLKLPEIQNT FPG SGKHLQ GGMPGMNEMD YWNRLNSLNR ARCENDVDFC LKQLQTAHDK AKIEPIKQAF QSSKGKERRQ PNVDEIAAAR IIQQ ILANP DCIHDDHVLI NGQKLEQQFF RDLLAKCEMA VVGSLLNDTD IGNIDTLMRH EKDTEFHSTN PEAVPVKIGE YWIND QRIN NSSGNITQKK HDLIFLMQND AWYFSRVNAI AQNRDKGSTF KEVLITTLMT PLTSKALVDT SQAKPPTRLF RGLNLS EEF TKGLIDQANA MIANTTERLF TDHSPEAFKQ IKLNDLSKMS GRTNASTTTE IKLVKETWDS NVIFEMLDPD GLLHSKQ VG RHGEGT(86N)SEF SVYLPEDVAL VPVKVTLDGK TQKGENRYVF TFVAVKSPDF TPRHESGYAV EPFLRMQAAK LAEV KSSIE KAQRAPDLET IFNLQNEVEA VQYSHLSTGY KNFLKNTVGP VLENSLSGLM ESDTDTLSKA LAAFPSDTQW SAFNF EEAR QAKRQMDAIK QMVGNKVVLD ALTQCQDALE KQNIAGALDA LKKIPSEKEM GTIRRELREQ IQSARQELES LQRAVV TPV VTDEKKVRER YDALIENTSK KITELETGKL PNLDAVKKGI SNLSNLKQEV TVLRNEKIRM HVGTDKVDFS DVEKLEQ QI QVIDTKLADA YLLEVTKQIS A |
-Macromolecule #2: Calmodulin-dependent glutamylase SidJ
Macromolecule | Name: Calmodulin-dependent glutamylase SidJ / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: Ligases |
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Source (natural) | Organism: Legionella pneumophila (bacteria) |
Molecular weight | Theoretical: 91.826992 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: HHHHHHSAGL EVLFQGPMVK QYYFARRGET STHDTSLPPP VKVLSGRSIP LKEIPFETTR NELVQIYLTS VDQLIKSNKL NSIPSQQIA SHYLFLRSLA NSETDGIKKN QILSLAKPLG IYLASKEPHV WKTINELIEK SEYPIIHYLK NNRAHSNFML A LIHEYHKE ...String: HHHHHHSAGL EVLFQGPMVK QYYFARRGET STHDTSLPPP VKVLSGRSIP LKEIPFETTR NELVQIYLTS VDQLIKSNKL NSIPSQQIA SHYLFLRSLA NSETDGIKKN QILSLAKPLG IYLASKEPHV WKTINELIEK SEYPIIHYLK NNRAHSNFML A LIHEYHKE PLTKNQSAFV QKFRDSSVFL FPNPIYTAWL AHSYDEDSSF NPMFRERLST NFYHSTLTDN LLLRTEPKEV TL SSEHHYK KEKGPIDSSF RYQMSSDRLL RIQGRTLLFS TPQNDVVAVK VQKRGEPKST LEEEFQMADY LLKHQSRLDV YSK LPQPLG QYSVKKSEIL EISRGSLDFE RFKTLIGDSK DLEVYVYKAP LTYFTYLHDK NQDLEDLTAS VKTNVHDLFV LLRE GIMFP QLADIFHTHF GEDEREDKGR YQALVQLLNV LQFQLGRIDK WQKAVEYVNL RSSGLADLGD SLPITSLFTS SDFTK HYFS ALLTGGYHPT FFDKSSGTAN SLFTGKRRLF GNYLYLNTIA EYLLVIQLTL GSYGDKVTRD MMDKPKKEAV WRELAN VMF TSCAEAIHIM TGIPQSRALT LLKQRANIEK HFRQTQFWMT PDYSKLDEDT LQMEQYSIYS GEPEYEFTDK LVSGVGL SV DGTHQDLGGY NRESPLRELE KLLYATVTLI EGTMQLDKEF FKQLQQVEKI LSGEIKTDAN SCFEAVAQLL DLARPRCH F QKRLVLSYYE EAKLKYPSAP TDAYDSRFQV VAKTNAAITI QRFWRETRKN LSENSDIESE KPESERTTDK RLK |
-Macromolecule #3: Calmodulin
Macromolecule | Name: Calmodulin / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 19.066006 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: HHHHHHSSGL EVLFQGPHMM ADQLTEEQIA EFKEAFSLFD KDGDGTITTK ELGTVMRSLG QNPTEAELQD MINEVDADGN GTIDFPEFL TMMARKMKDT DSEEEIREAF RVFDKDGNGY ISAAELRHVM TNLGEKLTDE EVDEMIREAD IDGDGQVNYE E FVQMMTAK |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #5: CALCIUM ION
Macromolecule | Name: CALCIUM ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: CA |
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Molecular weight | Theoretical: 40.078 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 4 K / Instrument: FEI VITROBOT MARK I |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 47.45 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |