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- PDB-8uc6: Calpain-7:IST1 Complex -

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Basic information

Entry
Database: PDB / ID: 8uc6
TitleCalpain-7:IST1 Complex
Components
  • Calpain-7
  • IST1 homolog
KeywordsPEPTIDE BINDING PROTEIN / ESCRT-III / Abscission
Function / homology
Function and homology information


viral capsid secondary envelopment / MIT domain binding / abscission / ESCRT III complex disassembly / calcium-dependent cysteine-type endopeptidase activity / cytoskeleton-dependent cytokinesis / collateral sprouting / Sealing of the nuclear envelope (NE) by ESCRT-III / positive regulation of collateral sprouting / self proteolysis ...viral capsid secondary envelopment / MIT domain binding / abscission / ESCRT III complex disassembly / calcium-dependent cysteine-type endopeptidase activity / cytoskeleton-dependent cytokinesis / collateral sprouting / Sealing of the nuclear envelope (NE) by ESCRT-III / positive regulation of collateral sprouting / self proteolysis / multivesicular body assembly / Flemming body / positive regulation of epithelial cell migration / positive regulation of proteolysis / viral release from host cell / endoplasmic reticulum-Golgi intermediate compartment / Degradation of the extracellular matrix / establishment of protein localization / protein localization / azurophil granule lumen / protein transport / nuclear envelope / midbody / endopeptidase activity / Hydrolases; Acting on peptide bonds (peptidases); Cysteine endopeptidases / cadherin binding / protein domain specific binding / cell division / cysteine-type endopeptidase activity / intracellular membrane-bounded organelle / centrosome / chromatin / Neutrophil degranulation / protein-containing complex binding / proteolysis / extracellular exosome / extracellular region / identical protein binding / nucleus / cytosol
Similarity search - Function
Peptidase C2, calpain, domain III / calpain_III / Peptidase C2, calpain, large subunit, domain III / Peptidase C2, calpain family / Calpain large subunit, domain III superfamily / Calpain large subunit, domain III / Peptidase C2, calpain, catalytic domain / Calpain family cysteine protease / Cysteine proteinase, calpain-type, catalytic domain profile. / Calpain-like thiol protease family. ...Peptidase C2, calpain, domain III / calpain_III / Peptidase C2, calpain, large subunit, domain III / Peptidase C2, calpain family / Calpain large subunit, domain III superfamily / Calpain large subunit, domain III / Peptidase C2, calpain, catalytic domain / Calpain family cysteine protease / Cysteine proteinase, calpain-type, catalytic domain profile. / Calpain-like thiol protease family. / Vacuolar protein sorting-associated protein Ist1 / Regulator of Vps4 activity in the MVB pathway / Vacuolar protein sorting-associated protein IST1-like / MIT (microtubule interacting and transport) domain / MIT domain superfamily / MIT domain / Microtubule Interacting and Trafficking molecule domain / Papain-like cysteine peptidase superfamily
Similarity search - Domain/homology
IST1 homolog / Calpain-7
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.701 Å
AuthorsPaine, E. / Whitby, F.G. / Hill, C.P. / Sunquist, W.I.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1F31GM139318 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)5R01GM112080 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)S10OD030326 United States
CitationJournal: Elife / Year: 2023
Title: The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission.
Authors: Paine, E.L. / Skalicky, J.J. / Whitby, F.G. / Mackay, D.R. / Ullman, K.S. / Hill, C.P. / Sundquist, W.I.
History
DepositionSep 25, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 25, 2023Provider: repository / Type: Initial release
Revision 1.1May 15, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: Calpain-7
G: IST1 homolog
B: Calpain-7
E: IST1 homolog


Theoretical massNumber of molelcules
Total (without water)47,0104
Polymers47,0104
Non-polymers00
Water75742
1
C: Calpain-7
G: IST1 homolog


Theoretical massNumber of molelcules
Total (without water)23,5052
Polymers23,5052
Non-polymers00
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2350 Å2
ΔGint-17 kcal/mol
Surface area11230 Å2
MethodPISA
2
B: Calpain-7
E: IST1 homolog


Theoretical massNumber of molelcules
Total (without water)23,5052
Polymers23,5052
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2340 Å2
ΔGint-15 kcal/mol
Surface area11040 Å2
MethodPISA
Unit cell
Length a, b, c (Å)87.841, 87.841, 183.894
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number179
Space group name H-MP6522
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11(chain B and (resid 1 through 69 or resid 78 through 153))
21(chain C and (resid 1 through 69 or resid 78 through 153))
12chain E
22chain G

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDSelection detailsAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
111METMETSERSER(chain B and (resid 1 through 69 or resid 78 through 153))BC1 - 691 - 69
121PROPROPROPRO(chain B and (resid 1 through 69 or resid 78 through 153))BC78 - 15378 - 153
211METMETSERSER(chain C and (resid 1 through 69 or resid 78 through 153))CA1 - 691 - 69
221PROPROPROPRO(chain C and (resid 1 through 69 or resid 78 through 153))CA78 - 15378 - 153
112VALVALLYSLYSchain EED324 - 3643 - 43
212VALVALLYSLYSchain GGB324 - 3643 - 43

NCS ensembles :
ID
1
2

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Components

#1: Protein Calpain-7 /


Mass: 18603.771 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CAPN7 / Cell line (production host): RIPL / Production host: Escherichia coli (E. coli) / Strain (production host): BL21-Codon Plus / Variant (production host): DE3 / References: UniProt: Q9Y6W3
#2: Protein/peptide IST1 homolog


Mass: 4901.372 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: IST1 / Cell line (production host): RIPL / Production host: Escherichia coli (E. coli) / Strain (production host): BL21-Codon Plus / Variant (production host): DE3 / References: UniProt: P53990
#3: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 42 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.18 Å3/Da / Density % sol: 43.53 %
Crystal growTemperature: 294 K / Method: vapor diffusion, sitting drop / pH: 4.2
Details: Both proteins were purified and concentrated to 20 mg/ml in buffer containing 25 mM Tris, pH 7.2, 150 mM NaCl, 1 mM TCEP, 0.5 mM EDTA. The proteins were mixed in a 1:2 molar ratio (CAPN7: ...Details: Both proteins were purified and concentrated to 20 mg/ml in buffer containing 25 mM Tris, pH 7.2, 150 mM NaCl, 1 mM TCEP, 0.5 mM EDTA. The proteins were mixed in a 1:2 molar ratio (CAPN7:IST1). Crystals grew at 21 C (294 K) in Rigaku Wizard Cryo condition D5 (25% (v/v) 1,2-Porpanediol, 100 mM Sodium phosphate dibasic/Citric acid pH 4.2, 5% (w/v) PEG 3000, 10% (v/v) Glycerol. Crystals were transferred briefly to crystallization buffer supplemented with 25% added glycerol prior to plunging in liquid nitrogen

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRL / Beamline: BL14-1 / Wavelength: 1.19499 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jul 27, 2019
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.19499 Å / Relative weight: 1
ReflectionResolution: 2.7→39.631 Å / Num. obs: 12228 / % possible obs: 100 % / Redundancy: 114.3 % / CC1/2: 0.998 / Rmerge(I) obs: 0.855 / Rpim(I) all: 0.08 / Rrim(I) all: 0.859 / Net I/σ(I): 11
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. measured allNum. unique obsCC1/2Rpim(I) allRrim(I) allNet I/σ(I) obs% possible all
2.7-2.83104.16.8116452615810.650.6666.8431.599.9
8.96-39.63930.144387744170.9990.0150.14539.699.1

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Processing

Software
NameVersionClassification
XDSdata reduction
Aimless0.7.4data scaling
PHENIX1.16refinement
PDB_EXTRACT3.27data extraction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 2KW3

2kw3


Resolution: 2.701→39.631 Å / SU ML: 0.37 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 27.38 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2847 1221 10.03 %
Rwork0.2112 10949 -
obs0.2184 12170 99.95 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 114.56 Å2 / Biso mean: 48.851 Å2 / Biso min: 17.98 Å2
Refinement stepCycle: final / Resolution: 2.701→39.631 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2779 0 0 42 2821
Biso mean---46.43 -
Num. residues----345
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDNumberRefine-IDRmsType
11B874X-RAY DIFFRACTION3.534TORSIONAL
12C874X-RAY DIFFRACTION3.534TORSIONAL
21E142X-RAY DIFFRACTION3.534TORSIONAL
22G142X-RAY DIFFRACTION3.534TORSIONAL
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / % reflection obs: 100 %

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection Rwork
2.701-2.80880.37091310.28361178
2.8088-2.93660.29531310.24891188
2.9366-3.09130.34111310.21651173
3.0913-3.28490.26551330.20651191
3.2849-3.53840.27541340.19481200
3.5384-3.89420.23091340.17451198
3.8942-4.45710.24941360.17661229
4.4571-5.61290.29661390.21151239
5.6129-39.6310.3071520.24181353

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