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- PDB-7w70: Crystal structure of the PDZ-C domain fragment of Kangiella koree... -

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Basic information

Entry
Database: PDB / ID: 7w70
TitleCrystal structure of the PDZ-C domain fragment of Kangiella koreensis RseP orthologue
ComponentsZinc metalloproteaseMetalloproteinase
KeywordsHYDROLASE / Intramembrane protease
Function / homology
Function and homology information


Hydrolases; Acting on peptide bonds (peptidases); Metalloendopeptidases / metalloendopeptidase activity / proteolysis / membrane / metal ion binding
Similarity search - Function
Peptidase M50, putative membrane-associated zinc metallopeptidase / Peptidase M50 / Peptidase family M50 / PDZ domain 6 / PDZ domain / PDZ domain profile. / Domain present in PSD-95, Dlg, and ZO-1/2. / PDZ domain / PDZ superfamily
Similarity search - Domain/homology
Zinc metalloprotease
Similarity search - Component
Biological speciesKangiella koreensis DSM 16069 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.15 Å
AuthorsMiyoshi, K. / Nogi, T.
Funding support Japan, 1items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)19H03170 Japan
CitationJournal: Sci Adv / Year: 2022
Title: Mechanistic insights into intramembrane proteolysis by site-2 protease homolog RseP.
Authors: Yuki Imaizumi / Kazunori Takanuki / Takuya Miyake / Mizuki Takemoto / Kunio Hirata / Mika Hirose / Rika Oi / Tatsuya Kobayashi / Kenichi Miyoshi / Rie Aruga / Tatsuhiko Yokoyama / Shizuka ...Authors: Yuki Imaizumi / Kazunori Takanuki / Takuya Miyake / Mizuki Takemoto / Kunio Hirata / Mika Hirose / Rika Oi / Tatsuya Kobayashi / Kenichi Miyoshi / Rie Aruga / Tatsuhiko Yokoyama / Shizuka Katagiri / Hiroaki Matsuura / Kenji Iwasaki / Takayuki Kato / Mika K Kaneko / Yukinari Kato / Michiko Tajiri / Satoko Akashi / Osamu Nureki / Yohei Hizukuri / Yoshinori Akiyama / Terukazu Nogi /
Abstract: Site-2 proteases are a conserved family of intramembrane proteases that cleave transmembrane substrates to regulate signal transduction and maintain proteostasis. Here, we elucidated crystal ...Site-2 proteases are a conserved family of intramembrane proteases that cleave transmembrane substrates to regulate signal transduction and maintain proteostasis. Here, we elucidated crystal structures of inhibitor-bound forms of bacterial site-2 proteases including RseP. Structure-based chemical modification and cross-linking experiments indicated that the RseP domains surrounding the active center undergo conformational changes to expose the substrate-binding site, suggesting that RseP has a gating mechanism to regulate substrate entry. Furthermore, mutational analysis suggests that a conserved electrostatic linkage between the transmembrane and peripheral membrane-associated domains mediates the conformational changes. In vivo cleavage assays also support that the substrate transmembrane helix is unwound by strand addition to the intramembrane β sheet of RseP and is clamped by a conserved asparagine residue at the active center for efficient cleavage. This mechanism underlying the substrate binding, i.e., unwinding and clamping, appears common across distinct families of intramembrane proteases that cleave transmembrane segments.
History
DepositionDec 2, 2021Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Sep 7, 2022Provider: repository / Type: Initial release
Revision 1.1Nov 29, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Zinc metalloprotease
B: Zinc metalloprotease


Theoretical massNumber of molelcules
Total (without water)18,5272
Polymers18,5272
Non-polymers00
Water2,990166
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area1230 Å2
ΔGint-6 kcal/mol
Surface area9370 Å2
MethodPISA
Unit cell
Length a, b, c (Å)40.673, 42.408, 97.949
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121
Space group name HallP2ac2ab
Symmetry operation#1: x,y,z
#2: x+1/2,-y+1/2,-z
#3: -x,y+1/2,-z+1/2
#4: -x+1/2,-y,z+1/2

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Components

#1: Protein Zinc metalloprotease / Metalloproteinase


Mass: 9263.323 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Kangiella koreensis DSM 16069 (bacteria)
Strain: DSM 16069 / KCTC 12182 / SW-125 / Gene: Kkor_1905 / Production host: Escherichia coli (E. coli)
References: UniProt: C7R5Z1, Hydrolases; Acting on peptide bonds (peptidases); Metalloendopeptidases
#2: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 166 / 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: 2.28 Å3/Da / Density % sol: 46.05 %
Description: The entry contains friedel pairs in F_plus/minus columns and I_plus/minus columns
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop / pH: 7.5 / Details: 1.0 M ammonium sulfate, 0.1 M HEPES-Na (pH 7.5)

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: Photon Factory / Beamline: BL-17A / Wavelength: 0.98 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: May 22, 2019
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.98 Å / Relative weight: 1
ReflectionResolution: 1.15→42.41 Å / Num. obs: 60798 / % possible obs: 99.6 % / Redundancy: 6.4 % / Biso Wilson estimate: 14.88 Å2 / CC1/2: 0.999 / Rpim(I) all: 0.017 / Net I/σ(I): 17.1
Reflection shellResolution: 1.15→1.17 Å / Mean I/σ(I) obs: 2.2 / Num. unique obs: 2913 / CC1/2: 0.794 / Rpim(I) all: 0.35

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Processing

Software
NameVersionClassification
PHENIX1.19.2_4158refinement
XDSdata reduction
Aimlessdata scaling
MOLREPphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 2ZPM

2zpm


Resolution: 1.15→31.29 Å / SU ML: 0.125 / Cross valid method: FREE R-VALUE / σ(F): 0.07 / Phase error: 23.3137
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Details: The entry contains friedel pairs in F_plus/minus columns and I_plus/minus columns
RfactorNum. reflection% reflection
Rfree0.2057 5779 5.04 %
Rwork0.1962 108919 -
obs0.1967 60798 98.63 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 21.84 Å2
Refinement stepCycle: LAST / Resolution: 1.15→31.29 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1308 0 0 166 1474
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00591461
X-RAY DIFFRACTIONf_angle_d0.82192015
X-RAY DIFFRACTIONf_chiral_restr0.076227
X-RAY DIFFRACTIONf_plane_restr0.0066274
X-RAY DIFFRACTIONf_dihedral_angle_d5.3086221
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
1.15-1.160.27591830.30563582X-RAY DIFFRACTION98.12
1.16-1.180.32351600.30143758X-RAY DIFFRACTION98.99
1.18-1.190.27131760.27723602X-RAY DIFFRACTION99.08
1.19-1.210.29951800.27153619X-RAY DIFFRACTION98.73
1.21-1.220.30791990.26853674X-RAY DIFFRACTION99.33
1.22-1.240.2891950.26953615X-RAY DIFFRACTION98.99
1.24-1.260.2761950.25683707X-RAY DIFFRACTION99.26
1.26-1.280.28091820.25673662X-RAY DIFFRACTION99.12
1.28-1.30.30771790.24183616X-RAY DIFFRACTION99.22
1.3-1.320.25792080.24163655X-RAY DIFFRACTION98.82
1.32-1.340.27072560.24813573X-RAY DIFFRACTION99.07
1.34-1.360.25971950.23643683X-RAY DIFFRACTION98.95
1.36-1.390.24021980.23683503X-RAY DIFFRACTION96.99
1.39-1.420.22292250.2293627X-RAY DIFFRACTION98.85
1.42-1.450.25242100.213660X-RAY DIFFRACTION99.61
1.45-1.480.22081630.21683703X-RAY DIFFRACTION99.79
1.48-1.520.21921960.2053673X-RAY DIFFRACTION99.51
1.52-1.560.23042310.20033638X-RAY DIFFRACTION99.28
1.56-1.610.22542320.19453584X-RAY DIFFRACTION99.53
1.61-1.660.22751920.23659X-RAY DIFFRACTION99.43
1.66-1.720.18271850.1993659X-RAY DIFFRACTION98.82
1.72-1.790.24031640.20483667X-RAY DIFFRACTION98.94
1.79-1.870.20651370.20423670X-RAY DIFFRACTION98.25
1.87-1.970.19821870.18733619X-RAY DIFFRACTION97.92
1.97-2.090.17442200.19093574X-RAY DIFFRACTION97.28
2.09-2.250.2161910.18693522X-RAY DIFFRACTION95.79
2.25-2.480.21661470.19513492X-RAY DIFFRACTION94.06
2.48-2.840.20912000.20553622X-RAY DIFFRACTION98.79
2.84-3.570.17832030.18223645X-RAY DIFFRACTION99.35
3.57-31.290.17061900.16063656X-RAY DIFFRACTION99
Refinement TLS params.Method: refined / Origin x: 26.0765295291 Å / Origin y: 21.9701877421 Å / Origin z: 14.0976019364 Å
111213212223313233
T0.119760733891 Å20.0166121593798 Å20.00114078037943 Å2-0.12773860542 Å2-0.00874318656783 Å2--0.141012589071 Å2
L0.546962321156 °20.62793711151 °20.914920233234 °2-0.366853112121 °20.450197906212 °2--1.03444290037 °2
S-0.00987088762049 Å °-0.0095168537562 Å °0.0355652311658 Å °0.050359833911 Å °-0.00130316018687 Å °0.0166476729605 Å °-0.0480841563119 Å °-0.0462822216522 Å °0.00696832699605 Å °
Refinement TLS groupSelection details: all

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