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- PDB-3itn: Crystal structure of Pseudo-activated Procaspase-3 -

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Basic information

Entry
Database: PDB / ID: 3itn
TitleCrystal structure of Pseudo-activated Procaspase-3
Components
  • ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor
  • Caspase-3Caspase 3
KeywordsHYDROLASE/HYDROLASE INHIBITOR / caspase-3 / Apoptosis / Hydrolase / Phosphoprotein / Protease / S-nitrosylation / Thiol protease / Zymogen / HYDROLASE-HYDROLASE INHIBITOR COMPLEX
Function / homology
Function and homology information


caspase-3 / Stimulation of the cell death response by PAK-2p34 / phospholipase A2 activator activity / anterior neural tube closure / intrinsic apoptotic signaling pathway in response to osmotic stress / leukocyte apoptotic process / positive regulation of pyroptotic inflammatory response / glial cell apoptotic process / NADE modulates death signalling / luteolysis ...caspase-3 / Stimulation of the cell death response by PAK-2p34 / phospholipase A2 activator activity / anterior neural tube closure / intrinsic apoptotic signaling pathway in response to osmotic stress / leukocyte apoptotic process / positive regulation of pyroptotic inflammatory response / glial cell apoptotic process / NADE modulates death signalling / luteolysis / response to cobalt ion / cysteine-type endopeptidase activity involved in apoptotic signaling pathway / death-inducing signaling complex / cyclin-dependent protein serine/threonine kinase inhibitor activity / cellular response to staurosporine / Apoptosis induced DNA fragmentation / Apoptotic cleavage of cell adhesion proteins / cysteine-type endopeptidase activity involved in execution phase of apoptosis / Caspase activation via Dependence Receptors in the absence of ligand / death receptor binding / SMAC, XIAP-regulated apoptotic response / axonal fasciculation / Activation of caspases through apoptosome-mediated cleavage / Signaling by Hippo / SMAC (DIABLO) binds to IAPs / SMAC(DIABLO)-mediated dissociation of IAP:caspase complexes / cysteine-type endopeptidase activity involved in apoptotic process / fibroblast apoptotic process / execution phase of apoptosis / negative regulation of cytokine production / epithelial cell apoptotic process / platelet formation / Other interleukin signaling / positive regulation of amyloid-beta formation / pyroptotic inflammatory response / Apoptotic cleavage of cellular proteins / negative regulation of B cell proliferation / T cell homeostasis / negative regulation of activated T cell proliferation / neurotrophin TRK receptor signaling pathway / B cell homeostasis / protein maturation / negative regulation of cell cycle / response to X-ray / Caspase-mediated cleavage of cytoskeletal proteins / regulation of macroautophagy / response to amino acid / cell fate commitment / Pyroptosis / response to tumor necrosis factor / response to glucose / response to UV / response to glucocorticoid / keratinocyte differentiation / striated muscle cell differentiation / Degradation of the extracellular matrix / intrinsic apoptotic signaling pathway / erythrocyte differentiation / response to nicotine / apoptotic signaling pathway / hippocampus development / sensory perception of sound / protein catabolic process / regulation of protein stability / response to hydrogen peroxide / protein processing / neuron differentiation / response to wounding / positive regulation of neuron apoptotic process / response to estradiol / heart development / peptidase activity / neuron apoptotic process / protease binding / response to lipopolysaccharide / aspartic-type endopeptidase activity / response to hypoxia / learning or memory / response to xenobiotic stimulus / positive regulation of apoptotic process / cysteine-type endopeptidase activity / neuronal cell body / apoptotic process / DNA damage response / protein-containing complex binding / proteolysis / nucleoplasm / nucleus / cytosol / cytoplasm
Similarity search - Function
Rossmann fold - #1460 / Peptidase family C14A, His active site / Caspase family histidine active site. / Peptidase C14, caspase non-catalytic subunit p10 / Peptidase family C14A, cysteine active site / Caspase family cysteine active site. / Caspase family p10 domain profile. / Peptidase C14A, caspase catalytic domain / Caspase, interleukin-1 beta converting enzyme (ICE) homologues / Peptidase C14, p20 domain ...Rossmann fold - #1460 / Peptidase family C14A, His active site / Caspase family histidine active site. / Peptidase C14, caspase non-catalytic subunit p10 / Peptidase family C14A, cysteine active site / Caspase family cysteine active site. / Caspase family p10 domain profile. / Peptidase C14A, caspase catalytic domain / Caspase, interleukin-1 beta converting enzyme (ICE) homologues / Peptidase C14, p20 domain / Caspase family p20 domain profile. / : / Caspase domain / Caspase-like domain superfamily / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
Ac-Asp-Glu-Val-Asp-CMK / Caspase-3
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.63 Å
AuthorsWalters, J. / Pop, C. / Scott, F.L. / Drag, M. / Swartz, P.D. / Mattos, C. / Salvesen, G.S. / Clark, A.C.
CitationJournal: Biochem.J. / Year: 2009
Title: A constitutively active and uninhibitable caspase-3 zymogen efficiently induces apoptosis.
Authors: Walters, J. / Pop, C. / Scott, F.L. / Drag, M. / Swartz, P. / Mattos, C. / Salvesen, G.S. / Clark, A.C.
History
DepositionAug 28, 2009Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 2, 2010Provider: repository / Type: Initial release
Revision 1.1Jul 13, 2011Group: Version format compliance
Revision 1.2Dec 12, 2012Group: Other
Revision 1.3Sep 6, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_conn / struct_ref_seq / struct_ref_seq_dif
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.pdbx_leaving_atom_flag / _struct_ref_seq.db_align_beg / _struct_ref_seq.db_align_end / _struct_ref_seq_dif.details

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor


Theoretical massNumber of molelcules
Total (without water)29,1692
Polymers29,1692
Non-polymers00
Water4,017223
1
A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor

A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor

A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor

A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor


Theoretical massNumber of molelcules
Total (without water)116,6748
Polymers116,6748
Non-polymers00
Water1448
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_665-x+1,-y+1,z1
crystal symmetry operation3_656-x+1,y,-z+11
crystal symmetry operation4_566x,-y+1,-z+11
Buried area16780 Å2
ΔGint-54 kcal/mol
Surface area35220 Å2
MethodPISA
2
A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor

A: Caspase-3
B: ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor


Theoretical massNumber of molelcules
Total (without water)58,3374
Polymers58,3374
Non-polymers00
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation3_656-x+1,y,-z+11
Buried area6800 Å2
ΔGint-18 kcal/mol
Surface area19210 Å2
MethodPISA
3


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area1300 Å2
ΔGint1 kcal/mol
Surface area11700 Å2
MethodPISA
Unit cell
Length a, b, c (Å)69.224, 84.607, 96.171
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number23
Space group name H-MI222

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Components

#1: Protein Caspase-3 / Caspase 3 / CASP-3 / Apopain / Cysteine protease CPP32 / CPP-32 / Yama protein / SREBP cleavage activity 1 / ...CASP-3 / Apopain / Cysteine protease CPP32 / CPP-32 / Yama protein / SREBP cleavage activity 1 / SCA-1 / Caspase-3 subunit p17 / Caspase-3 subunit p12


Mass: 28633.586 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CASP3, CPP32 / Plasmid: pcDNA3 / Production host: Escherichia coli (E. coli) / References: UniProt: P42574, caspase-3
#2: Protein/peptide ACETYL-ASP-GLU-VAL-ASP-CHLOROMETHYL KETONE inhibitor


Type: Peptide-like / Class: Inhibitor / Mass: 534.946 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: Calbiochem / References: Ac-Asp-Glu-Val-Asp-CMK
#3: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 223 / 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: 2.42 Å3/Da / Density % sol: 49.22 %
Crystal growTemperature: 291.15 K / Method: vapor diffusion, hanging drop / pH: 8.5
Details: Proteins were dialyzed in a buffer of 10 mM Tris-HCl (pH 8.5) and 1 mM DTT. The protein was concentrated to 10 mg/mL using Amicon ultrafree centrifugal filter devices, and inhibitor, Ac-DEVD- ...Details: Proteins were dialyzed in a buffer of 10 mM Tris-HCl (pH 8.5) and 1 mM DTT. The protein was concentrated to 10 mg/mL using Amicon ultrafree centrifugal filter devices, and inhibitor, Ac-DEVD-CMK reconstituted in DMSO, was then added at a 5:1 inhibitor:peptide ratio (w/w). The protein was diluted to a concentration of 8 mg/mL by adding 10 mM Tris-HCl (pH 8.5), concentrated DTT, and concentrated NaN3 so that the final buffer consisted of 10 mM Tris-HCl (pH 8.5), 10 mM DTT, and 3 mM NaN3. Crystals were obtained by the hanging drop vapor diffusion method. Concentrated protein (2 L) was mixed 1:1 with a reservoir solution that contained 100 mM sodium citrate (pH 5), 3 mM NaN3, 10 mM DTT, and 17% PEG 6000 (w/v). The crystallization plates were incubated at 18 C., VAPOR DIFFUSION, HANGING DROP, temperature 291.15K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 22-ID / Wavelength: 1 Å
DetectorType: MARMOSAIC 300 mm CCD / Detector: CCD / Date: Oct 13, 2006
RadiationMonochromator: Rosenbaum-Rock monochromator high-resolution double-crystal Si(220) sagittal focusing, Rosenbaum-Rock vertical focusing mirror
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 1.63→33.79 Å / Num. all: 35601 / Num. obs: 32677 / % possible obs: 91.79 % / Observed criterion σ(F): 3 / Observed criterion σ(I): 3 / Redundancy: 5.45 % / Rmerge(I) obs: 0.52 / Rsym value: 0.444
Reflection shellHighest resolution: 1.63 Å / % possible all: 99

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Processing

Software
NameClassification
HKL-2000data collection
CNSrefinement
DENZOdata reduction
SCALEPACKdata scaling
CNSphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: Caspase-3 PDB code 2J30

2j30


Resolution: 1.63→33.79 Å / σ(F): 1 / Stereochemistry target values: Engh & Huber
RfactorNum. reflectionSelection details
Rfree0.2242 3063 Randomly selected 10% of reflections
Rwork0.1987 --
all0.22 35601 -
obs0.1987 30659 -
Refinement stepCycle: LAST / Resolution: 1.63→33.79 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1963 0 0 223 2186
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONc_bond_d0.004748
X-RAY DIFFRACTIONc_angle_d1.26877

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