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- PDB-6pen: Structure of Spastin Hexamer (whole model) in complex with substr... -

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Basic information

Entry
Database: PDB / ID: 6pen
TitleStructure of Spastin Hexamer (whole model) in complex with substrate peptide
Components
  • EYEYEYEYEY
  • Spastin
KeywordsMOTOR PROTEIN / AAA+ ATPase / Microtubule Severing
Function / homology
Function and homology information


cytokinetic process / microtubule-severing ATPase / microtubule severing ATPase activity / microtubule severing / endoplasmic reticulum tubular network / positive regulation of microtubule depolymerization / cytoskeleton-dependent cytokinesis / mitotic nuclear membrane reassembly / nuclear membrane reassembly / Sealing of the nuclear envelope (NE) by ESCRT-III ...cytokinetic process / microtubule-severing ATPase / microtubule severing ATPase activity / microtubule severing / endoplasmic reticulum tubular network / positive regulation of microtubule depolymerization / cytoskeleton-dependent cytokinesis / mitotic nuclear membrane reassembly / nuclear membrane reassembly / Sealing of the nuclear envelope (NE) by ESCRT-III / membrane fission / anterograde axonal transport / microtubule bundle formation / mitotic spindle disassembly / protein hexamerization / exit from mitosis / axonal transport of mitochondrion / beta-tubulin binding / positive regulation of cytokinesis / mitotic cytokinesis / alpha-tubulin binding / endoplasmic reticulum to Golgi vesicle-mediated transport / lipid droplet / axon cytoplasm / axonogenesis / isomerase activity / protein homooligomerization / spindle pole / midbody / microtubule binding / cytoplasmic vesicle / nuclear membrane / microtubule / endosome / axon / centrosome / endoplasmic reticulum membrane / protein-containing complex binding / perinuclear region of cytoplasm / ATP hydrolysis activity / extracellular exosome / nucleoplasm / ATP binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Spastin, chordate / Spastin / Vps4 oligomerisation, C-terminal / MIT domain / Microtubule Interacting and Trafficking molecule domain / : / Vps4 C terminal oligomerisation domain / AAA ATPase, AAA+ lid domain / AAA+ lid domain / ATPase, AAA-type, conserved site ...Spastin, chordate / Spastin / Vps4 oligomerisation, C-terminal / MIT domain / Microtubule Interacting and Trafficking molecule domain / : / Vps4 C terminal oligomerisation domain / AAA ATPase, AAA+ lid domain / AAA+ lid domain / ATPase, AAA-type, conserved site / AAA-protein family signature. / ATPase family associated with various cellular activities (AAA) / ATPase, AAA-type, core / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / BERYLLIUM TRIFLUORIDE ION / Spastin
Similarity search - Component
Biological speciesHomo sapiens (human)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsHan, H. / Schubert, H.L. / McCullough, J. / Monroe, N. / Sundquist, W.I. / Hill, C.P.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)RO1 GM112080 United States
CitationJournal: J Biol Chem / Year: 2020
Title: Structure of spastin bound to a glutamate-rich peptide implies a hand-over-hand mechanism of substrate translocation.
Authors: Han Han / Heidi L Schubert / John McCullough / Nicole Monroe / Michael D Purdy / Mark Yeager / Wesley I Sundquist / Christopher P Hill /
Abstract: Many members of the AAA+ ATPase family function as hexamers that unfold their protein substrates. These AAA unfoldases include spastin, which plays a critical role in the architecture of eukaryotic ...Many members of the AAA+ ATPase family function as hexamers that unfold their protein substrates. These AAA unfoldases include spastin, which plays a critical role in the architecture of eukaryotic cells by driving the remodeling and severing of microtubules, which are cytoskeletal polymers of tubulin subunits. Here, we demonstrate that a human spastin binds weakly to unmodified peptides from the C-terminal segment of human tubulin α1A/B. A peptide comprising alternating glutamate and tyrosine residues binds more tightly, which is consistent with the known importance of glutamylation for spastin microtubule severing activity. A cryo-EM structure of the spastin-peptide complex at 4.2 Å resolution revealed an asymmetric hexamer in which five spastin subunits adopt a helical, spiral staircase configuration that binds the peptide within the central pore, whereas the sixth subunit of the hexamer is displaced from the peptide/substrate, as if transitioning from one end of the helix to the other. This configuration differs from a recently published structure of spastin from , which forms a six-subunit spiral without a transitioning subunit. Our structure resembles other recently reported AAA unfoldases, including the meiotic clade relative Vps4, and supports a model in which spastin utilizes a hand-over-hand mechanism of tubulin translocation and microtubule remodeling.
History
DepositionJun 20, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 4, 2019Provider: repository / Type: Initial release
Revision 1.1Dec 11, 2019Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Jan 1, 2020Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.3Jan 22, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year / _citation_author.identifier_ORCID
Revision 1.4Mar 13, 2024Group: Data collection / Database references / Derived calculations
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_struct_conn_angle / struct_conn
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_auth_comp_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr2_label_atom_id / _pdbx_struct_conn_angle.ptnr2_label_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id

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Assembly

Deposited unit
A: Spastin
B: Spastin
C: Spastin
D: Spastin
E: Spastin
G: EYEYEYEYEY
F: Spastin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)330,96720
Polymers328,4697
Non-polymers2,49713
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Spastin / Spastic paraplegia 4 protein


Mass: 54498.328 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SPAST, ADPSP, FSP2, KIAA1083, SPG4 / Production host: Escherichia coli (E. coli) / References: UniProt: Q9UBP0, microtubule-severing ATPase
#2: Protein/peptide EYEYEYEYEY


Mass: 1479.453 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#3: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM
#4: Chemical
ChemComp-BEF / BERYLLIUM TRIFLUORIDE ION


Mass: 66.007 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: BeF3
#5: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Mg
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Spastin Hexamer with a substrate peptide bound in the central pore
Type: COMPLEX / Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 57 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.16_3549: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 119984 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00122510
ELECTRON MICROSCOPYf_angle_d0.4640835
ELECTRON MICROSCOPYf_dihedral_angle_d3.0298868
ELECTRON MICROSCOPYf_chiral_restr0.0331753
ELECTRON MICROSCOPYf_plane_restr0.0013299

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