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- PDB-6pxj: Crystal structure of human thrombin mutant I16T -

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Basic information

Entry
Database: PDB / ID: 6pxj
TitleCrystal structure of human thrombin mutant I16T
Components
  • Thrombin heavy chain
  • Thrombin light chain
KeywordsHYDROLASE / Trypsin -like proteases / Ionic interaction
Function / homology
Function and homology information


positive regulation of lipid kinase activity / cytolysis by host of symbiont cells / positive regulation of phospholipase C-activating G protein-coupled receptor signaling pathway / thrombospondin receptor activity / Defective factor XII causes hereditary angioedema / thrombin / regulation of blood coagulation / neutrophil-mediated killing of gram-negative bacterium / ligand-gated ion channel signaling pathway / Defective F8 cleavage by thrombin ...positive regulation of lipid kinase activity / cytolysis by host of symbiont cells / positive regulation of phospholipase C-activating G protein-coupled receptor signaling pathway / thrombospondin receptor activity / Defective factor XII causes hereditary angioedema / thrombin / regulation of blood coagulation / neutrophil-mediated killing of gram-negative bacterium / ligand-gated ion channel signaling pathway / Defective F8 cleavage by thrombin / Platelet Aggregation (Plug Formation) / negative regulation of astrocyte differentiation / negative regulation of platelet activation / positive regulation of collagen biosynthetic process / negative regulation of cytokine production involved in inflammatory response / positive regulation of blood coagulation / negative regulation of fibrinolysis / Gamma-carboxylation of protein precursors / Transport of gamma-carboxylated protein precursors from the endoplasmic reticulum to the Golgi apparatus / Common Pathway of Fibrin Clot Formation / Removal of aminoterminal propeptides from gamma-carboxylated proteins / fibrinolysis / regulation of cytosolic calcium ion concentration / Intrinsic Pathway of Fibrin Clot Formation / Peptide ligand-binding receptors / positive regulation of release of sequestered calcium ion into cytosol / Regulation of Complement cascade / acute-phase response / negative regulation of proteolysis / Cell surface interactions at the vascular wall / lipopolysaccharide binding / positive regulation of receptor signaling pathway via JAK-STAT / growth factor activity / positive regulation of insulin secretion / platelet activation / response to wounding / positive regulation of protein localization to nucleus / Golgi lumen / Regulation of Insulin-like Growth Factor (IGF) transport and uptake by Insulin-like Growth Factor Binding Proteins (IGFBPs) / positive regulation of reactive oxygen species metabolic process / blood coagulation / antimicrobial humoral immune response mediated by antimicrobial peptide / Thrombin signalling through proteinase activated receptors (PARs) / heparin binding / regulation of cell shape / positive regulation of cell growth / G alpha (q) signalling events / collagen-containing extracellular matrix / blood microparticle / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / cell surface receptor signaling pathway / positive regulation of protein phosphorylation / G protein-coupled receptor signaling pathway / endoplasmic reticulum lumen / serine-type endopeptidase activity / signaling receptor binding / calcium ion binding / positive regulation of cell population proliferation / proteolysis / extracellular space / extracellular exosome / extracellular region / plasma membrane
Similarity search - Function
Prothrombin/thrombin / Thrombin light chain / Thrombin light chain domain superfamily / : / Thrombin light chain / Kringle domain / Kringle / Kringle, conserved site / Kringle superfamily / Kringle domain signature. ...Prothrombin/thrombin / Thrombin light chain / Thrombin light chain domain superfamily / : / Thrombin light chain / Kringle domain / Kringle / Kringle, conserved site / Kringle superfamily / Kringle domain signature. / Kringle domain profile. / Kringle domain / Vitamin K-dependent carboxylation/gamma-carboxyglutamic (GLA) domain / Gamma-carboxyglutamic acid-rich (GLA) domain / Gamma-carboxyglutamic acid-rich (GLA) domain superfamily / Vitamin K-dependent carboxylation domain. / Gla domain profile. / Domain containing Gla (gamma-carboxyglutamate) residues. / Kringle-like fold / Serine proteases, trypsin family, histidine active site / Serine proteases, trypsin family, serine active site / Peptidase S1A, chymotrypsin family / Serine proteases, trypsin family, histidine active site. / Serine proteases, trypsin family, serine active site. / Serine proteases, trypsin domain profile. / Trypsin-like serine protease / Serine proteases, trypsin domain / Trypsin / Trypsin-like serine proteases / Thrombin, subunit H / Peptidase S1, PA clan, chymotrypsin-like fold / Peptidase S1, PA clan / Beta Barrel / Mainly Beta
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / MOLECULAR REPLACEMENT / Resolution: 1.7 Å
AuthorsStojanovski, B. / Chen, Z. / Koester, S.K. / Pelc, L.A. / Di Cera, E.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI) United States
Citation
Journal: Sci Rep / Year: 2019
Title: Role of the I16-D194 ionic interaction in the trypsin fold.
Authors: Stojanovski, B.M. / Chen, Z. / Koester, S.K. / Pelc, L.A. / Di Cera, E.
#1: Journal: EMBO J. / Year: 1989
Title: THE REFINED 1.9 A CRYSTAL STRUCTURE OF HUMAN ALPHA-THROMBIN: INTERACTION WITH D-PHE-PRO-ARG CHLOROMETHYLKETONE AND SIGNIFICANCE OF THE TYR-PRO-PRO-TRP INSERTION SEGMENT.
Authors: Bode, W. / Mayr, I. / Baumann, U. / Huber, R. / Stone, S.R. / Hofsteenge, J.
History
DepositionJul 26, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 18, 2019Provider: repository / Type: Initial release
Revision 1.1Oct 11, 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 / 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_seq_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.ptnr2_auth_seq_id
Revision 1.2Nov 20, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
L: Thrombin light chain
H: Thrombin heavy chain
B: Thrombin heavy chain
A: Thrombin light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)67,9387
Polymers67,7294
Non-polymers2083
Water8,341463
1
L: Thrombin light chain
H: Thrombin heavy chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)34,0494
Polymers33,8652
Non-polymers1842
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area3060 Å2
ΔGint-14 kcal/mol
Surface area12790 Å2
MethodPISA
2
B: Thrombin heavy chain
A: Thrombin light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)33,8893
Polymers33,8652
Non-polymers241
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2210 Å2
ΔGint-18 kcal/mol
Surface area12680 Å2
MethodPISA
Unit cell
Length a, b, c (Å)81.595, 151.399, 50.561
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number18
Space group name H-MP21212
Components on special symmetry positions
IDModelComponents
11L-128-

HOH

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Components

#1: Protein/peptide Thrombin light chain / Coagulation factor II


Mass: 4096.534 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: GR were disordered in the structure / Source: (gene. exp.) Homo sapiens (human) / Gene: F2 / Production host: Cricetulus griseus (Chinese hamster) / References: UniProt: P00734, thrombin
#2: Protein Thrombin heavy chain / Coagulation factor II


Mass: 29768.168 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: I-T: mutant I to T. (original sequence is I). LKETWTANVGKG were disordered in the structure. GE were disordered in the structure.
Source: (gene. exp.) Homo sapiens (human) / Gene: F2 / Production host: Cricetulus griseus (Chinese hamster) / References: UniProt: P00734, thrombin
#3: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL


Mass: 92.094 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C3H8O3
#4: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 463 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY
Has protein modificationY

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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: 2.45 Å3/Da / Density % sol: 49.78 %
Crystal growTemperature: 295 K / Method: vapor diffusion, hanging drop / Details: 200 mM Mg formate, 20% PEG 3350

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: ROTATING ANODE / Type: RIGAKU MICROMAX-007 HF / Wavelength: 1.5418 Å
DetectorType: RIGAKU RAXIS IV++ / Detector: IMAGE PLATE / Date: Mar 14, 2019
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.5418 Å / Relative weight: 1
ReflectionResolution: 1.7→40 Å / Num. obs: 68725 / % possible obs: 98.5 % / Observed criterion σ(F): -1.5 / Observed criterion σ(I): -1.5 / Redundancy: 6.2 % / Rsym value: 0.053 / Net I/σ(I): 23.9
Reflection shellResolution: 1.7→1.73 Å / Redundancy: 5.8 % / Num. unique obs: 3328 / Rsym value: 0.703 / % possible all: 97.3

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Processing

Software
NameVersionClassification
REFMAC5.8.0232refinement
PDB_EXTRACT3.25data extraction
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 1PPB

1ppb


Resolution: 1.7→31.77 Å / Cor.coef. Fo:Fc: 0.969 / Cor.coef. Fo:Fc free: 0.958 / SU B: 2.223 / SU ML: 0.071 / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R: 0.099 / ESU R Free: 0.097
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS U VALUES : REFINED INDIVIDUALLY
RfactorNum. reflection% reflectionSelection details
Rfree0.2031 3335 4.9 %RANDOM
Rwork0.1727 ---
obs0.1742 65368 98.3 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å
Displacement parametersBiso max: 111.01 Å2 / Biso mean: 32.968 Å2 / Biso min: 17.79 Å2
Baniso -1Baniso -2Baniso -3
1-0 Å20 Å20 Å2
2--0.01 Å20 Å2
3----0.01 Å2
Refinement stepCycle: final / Resolution: 1.7→31.77 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4474 0 13 463 4950
Biso mean--43.17 45.11 -
Num. residues----552
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0130.0134623
X-RAY DIFFRACTIONr_bond_other_d0.0020.0174283
X-RAY DIFFRACTIONr_angle_refined_deg1.8531.6476246
X-RAY DIFFRACTIONr_angle_other_deg1.4241.5869945
X-RAY DIFFRACTIONr_dihedral_angle_1_deg8.0575554
X-RAY DIFFRACTIONr_dihedral_angle_2_deg29.68521.192260
X-RAY DIFFRACTIONr_dihedral_angle_3_deg16.30415822
X-RAY DIFFRACTIONr_dihedral_angle_4_deg16.4171540
X-RAY DIFFRACTIONr_chiral_restr0.1020.2566
X-RAY DIFFRACTIONr_gen_planes_refined0.0110.025128
X-RAY DIFFRACTIONr_gen_planes_other0.0020.021032
LS refinement shellResolution: 1.7→1.744 Å / Rfactor Rfree error: 0
RfactorNum. reflection% reflection
Rfree0.307 252 -
Rwork0.313 4565 -
obs--94.32 %

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