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- PDB-8f0m: Monobody 12D5 bound to KRAS(G12D) -

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Basic information

Entry
Database: PDB / ID: 8f0m
TitleMonobody 12D5 bound to KRAS(G12D)
Components
  • Isoform 2B of GTPase KRas
  • Monobody 12D5
KeywordsSIGNALING PROTEIN/DE NOVO PROTEIN / SIGNALING PROTEIN-DE NOVO PROTEIN complex
Function / homologysmall monomeric GTPase / Ca2+ pathway / GUANOSINE-5'-DIPHOSPHATE / alpha-D-glucopyranose / 5'-GUANOSINE-DIPHOSPHATE-MONOTHIOPHOSPHATE / Isoform 2B of GTPase KRas
Function and homology information
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.44 Å
AuthorsHattori, T. / Glasser, E. / Akkapeddi, P. / Ketavarapu, G. / Teng, K.W. / Koide, A. / Koide, S.
Funding support United States, 6items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)CA194864 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA201717 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA212608 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA246457 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA225131 United States
American Cancer SocietyPF-18-180-01-TBE United States
CitationJournal: Proc.Natl.Acad.Sci.USA / Year: 2023
Title: Exploring switch II pocket conformation of KRAS(G12D) with mutant-selective monobody inhibitors.
Authors: Akkapeddi, P. / Hattori, T. / Khan, I. / Glasser, E. / Koide, A. / Ketavarapu, G. / Whaby, M. / Zuberi, M. / Teng, K.W. / Lefler, J. / Maso, L. / Bang, I. / Ostrowski, M.C. / O'Bryan, J.P. / Koide, S.
History
DepositionNov 3, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 19, 2023Provider: repository / Type: Initial release
Revision 1.1Oct 25, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Item: _pdbx_initial_refinement_model.details

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Isoform 2B of GTPase KRas
B: Monobody 12D5
C: Isoform 2B of GTPase KRas
D: Monobody 12D5
hetero molecules


Theoretical massNumber of molelcules
Total (without water)60,91810
Polymers59,5264
Non-polymers1,3916
Water1,838102
1
A: Isoform 2B of GTPase KRas
B: Monobody 12D5
hetero molecules


  • defined by author&software
  • Evidence: gel filtration, The binding of monobody 12D5 (chain B, D) to KRAS(G12D) (Chain A, C) was confirmed by gel filtration and bio-layer interferometry (BLI) experiments. Our gel filtration ...Evidence: gel filtration, The binding of monobody 12D5 (chain B, D) to KRAS(G12D) (Chain A, C) was confirmed by gel filtration and bio-layer interferometry (BLI) experiments. Our gel filtration result indicates that the monobody 12D5-KRASG12D complex does not form a dimer. Thus, the contact between two complexes in the AUS is due to the crystal contact.
  • 30.6 kDa, 2 polymers
  • Search similar-shape structures of this assembly by Omokage search (details)
Theoretical massNumber of molelcules
Total (without water)30,5916
Polymers29,7632
Non-polymers8284
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2710 Å2
ΔGint-27 kcal/mol
Surface area12100 Å2
MethodPISA
2
C: Isoform 2B of GTPase KRas
D: Monobody 12D5
hetero molecules


Theoretical massNumber of molelcules
Total (without water)30,3274
Polymers29,7632
Non-polymers5642
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2730 Å2
ΔGint-24 kcal/mol
Surface area12950 Å2
MethodPISA
Unit cell
Length a, b, c (Å)151.386, 44.733, 88.162
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number18
Space group name H-MP21221

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Components

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Protein / Antibody / Sugars , 3 types, 6 molecules ACBD

#1: Protein Isoform 2B of GTPase KRas / K-Ras 2 / Ki-Ras / c-K-ras / c-Ki-ras


Mass: 19451.809 Da / Num. of mol.: 2 / Mutation: G12D, C51S, C80L, C118S
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: KRAS, KRAS2, RASK2 / Production host: Escherichia coli (E. coli) / References: UniProt: P01116-2, small monomeric GTPase
#2: Antibody Monobody 12D5


Mass: 10311.420 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#5: Sugar ChemComp-GLC / alpha-D-glucopyranose / alpha-D-glucose / D-glucose / glucose / Glucose


Type: D-saccharide, alpha linking / Mass: 180.156 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C6H12O6
IdentifierTypeProgram
DGlcpaCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
a-D-glucopyranoseCOMMON NAMEGMML 1.0
a-D-GlcpIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 4 types, 106 molecules

#3: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#4: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE / Guanosine diphosphate


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: GDP, energy-carrying molecule*YM
#6: Chemical ChemComp-GSP / 5'-GUANOSINE-DIPHOSPHATE-MONOTHIOPHOSPHATE


Mass: 539.246 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: C10H16N5O13P3S / Feature type: SUBJECT OF INVESTIGATION
#7: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 102 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY

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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: 2.56 Å3/Da / Density % sol: 52.02 %
Crystal growTemperature: 291 K / Method: vapor diffusion, sitting drop
Details: 0.2 M sodium sulfate decahydrate, 20% w/v PEG3350, 0.05% w/v benzamidine hydrochloride

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 19-ID / Wavelength: 0.97918 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Sep 24, 2022
RadiationMonochromator: Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97918 Å / Relative weight: 1
ReflectionResolution: 2.44→50 Å / Num. obs: 23370 / % possible obs: 98.4 % / Redundancy: 3.3 % / CC1/2: 0.939 / CC star: 0.984 / Rmerge(I) obs: 0.091 / Rpim(I) all: 0.061 / Rrim(I) all: 0.11 / Χ2: 1.043 / Net I/σ(I): 12.4
Reflection shellResolution: 2.44→2.48 Å / Redundancy: 3.2 % / Rmerge(I) obs: 0.308 / Mean I/σ(I) obs: 5 / Num. unique obs: 1168 / CC1/2: 0.905 / CC star: 0.975 / Rpim(I) all: 0.207 / Rrim(I) all: 0.373 / Χ2: 1.024 / % possible all: 99.7

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Processing

Software
NameVersionClassification
PHENIX(1.19.2_4158: 000)refinement
PDB-REDOrefinement
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB entries 7L0G & 5VPZ

7l0g

5vpz


Resolution: 2.44→42.9 Å / SU ML: 0.23 / Cross valid method: THROUGHOUT / σ(F): 1.36 / Phase error: 25.14 / Stereochemistry target values: ML
RfactorNum. reflection% reflectionSelection details
Rfree0.245 1189 5.1 %RANDOM
Rwork0.2009 ---
obs0.2031 23331 96.6 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.44→42.9 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4019 0 86 102 4207
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0034184
X-RAY DIFFRACTIONf_angle_d0.665699
X-RAY DIFFRACTIONf_dihedral_angle_d9.949601
X-RAY DIFFRACTIONf_chiral_restr0.047659
X-RAY DIFFRACTIONf_plane_restr0.007715
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.44-2.510.31341350.25852360X-RAY DIFFRACTION84
2.51-2.640.2881570.23572748X-RAY DIFFRACTION98
2.64-2.810.28641450.23582792X-RAY DIFFRACTION98
2.81-3.030.27081480.2442796X-RAY DIFFRACTION99
3.03-3.330.27771250.20632813X-RAY DIFFRACTION99
3.33-3.810.24021640.19752839X-RAY DIFFRACTION99
3.81-4.80.21521440.16482841X-RAY DIFFRACTION98
4.8-42.90.21641710.19012953X-RAY DIFFRACTION97
Refinement TLS params.Method: refined / Origin x: 22.1947 Å / Origin y: 29.3203 Å / Origin z: 36.6075 Å
111213212223313233
T0.2175 Å2-0.0022 Å2-0.0004 Å2-0.1162 Å2-0.0349 Å2--0.2407 Å2
L1.4052 °2-0.1697 °20.459 °2-0.0917 °20.0822 °2--1.3038 °2
S-0.0655 Å °-0.0289 Å °0.1927 Å °-0.0288 Å °0.0479 Å °-0.0323 Å °-0.0525 Å °-0.0937 Å °0.0001 Å °
Refinement TLS groupSelection details: all

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