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- PDB-8uja: T33-fn10 - Designed Tetrahedral Protein Cage Using Fragment-based... -

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Basic information

Entry
Database: PDB / ID: 8uja
TitleT33-fn10 - Designed Tetrahedral Protein Cage Using Fragment-based Hydrogen Bond Networks
Components
  • T33-fn10: engineered DrsE like sulfur reductase
  • T33-fn10: engineered enoyl-CoA hydratase/isomerase
KeywordsDE NOVO PROTEIN / tetrahedral nanoparticle / designed protein / de novo interface / two-component complex / Rosetta
Function / homologyEnoyl-CoA hydratase, C-terminal / Enoyl-CoA hydratase/isomerase, conserved site / Enoyl-CoA hydratase/isomerase signature. / Enoyl-CoA hydratase/isomerase / Enoyl-CoA hydratase/isomerase / ClpP/crotonase-like domain superfamily / isomerase activity / Enoyl-CoA hydratase/isomerase
Function and homology information
Biological speciesSulfurisphaera tokodaii str. 7 (archaea)
Novosphingobium aromaticivorans DSM 12444 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 6 Å
AuthorsMeador, K. / Sawaya, M.R. / Yeates, T.O.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM129854 United States
Citation
Journal: Structure / Year: 2024
Title: A suite of designed protein cages using machine learning and protein fragment-based protocols.
Authors: Kyle Meador / Roger Castells-Graells / Roman Aguirre / Michael R Sawaya / Mark A Arbing / Trent Sherman / Chethaka Senarathne / Todd O Yeates /
Abstract: Designed protein cages and related materials provide unique opportunities for applications in biotechnology and medicine, but their creation remains challenging. Here, we apply computational ...Designed protein cages and related materials provide unique opportunities for applications in biotechnology and medicine, but their creation remains challenging. Here, we apply computational approaches to design a suite of tetrahedrally symmetric, self-assembling protein cages. For the generation of docked conformations, we emphasize a protein fragment-based approach, while for sequence design of the de novo interface, a comparison of knowledge-based and machine learning protocols highlights the power and increased experimental success achieved using ProteinMPNN. An analysis of design outcomes provides insights for improving interface design protocols, including prioritizing fragment-based motifs, balancing interface hydrophobicity and polarity, and identifying preferred polar contact patterns. In all, we report five structures for seven protein cages, along with two structures of intermediate assemblies, with the highest resolution reaching 2.0 Å using cryo-EM. This set of designed cages adds substantially to the body of available protein nanoparticles, and to methodologies for their creation.
#1: Journal: bioRxiv / Year: 2023
Title: A Suite of Designed Protein Cages Using Machine Learning Algorithms and Protein Fragment-Based Protocols
Authors: Meador, K. / Castells-Graells, R. / Aguirre, R. / Sawaya, M.R. / Arbing, M.A. / Sherman, T. / Senarathne, C. / Yeates, T.O.
History
DepositionOct 11, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 6, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 3, 2024Group: Database references / Category: citation / citation_author

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: T33-fn10: engineered DrsE like sulfur reductase
B: T33-fn10: engineered enoyl-CoA hydratase/isomerase
C: T33-fn10: engineered DrsE like sulfur reductase
D: T33-fn10: engineered enoyl-CoA hydratase/isomerase
E: T33-fn10: engineered DrsE like sulfur reductase
F: T33-fn10: engineered enoyl-CoA hydratase/isomerase
G: T33-fn10: engineered DrsE like sulfur reductase
H: T33-fn10: engineered enoyl-CoA hydratase/isomerase
I: T33-fn10: engineered DrsE like sulfur reductase
J: T33-fn10: engineered enoyl-CoA hydratase/isomerase
K: T33-fn10: engineered DrsE like sulfur reductase
L: T33-fn10: engineered enoyl-CoA hydratase/isomerase
M: T33-fn10: engineered DrsE like sulfur reductase
N: T33-fn10: engineered enoyl-CoA hydratase/isomerase
O: T33-fn10: engineered DrsE like sulfur reductase
P: T33-fn10: engineered enoyl-CoA hydratase/isomerase


Theoretical massNumber of molelcules
Total (without water)318,35216
Polymers318,35216
Non-polymers00
Water0
1
A: T33-fn10: engineered DrsE like sulfur reductase
B: T33-fn10: engineered enoyl-CoA hydratase/isomerase
C: T33-fn10: engineered DrsE like sulfur reductase
D: T33-fn10: engineered enoyl-CoA hydratase/isomerase
E: T33-fn10: engineered DrsE like sulfur reductase
F: T33-fn10: engineered enoyl-CoA hydratase/isomerase
G: T33-fn10: engineered DrsE like sulfur reductase
H: T33-fn10: engineered enoyl-CoA hydratase/isomerase

A: T33-fn10: engineered DrsE like sulfur reductase
B: T33-fn10: engineered enoyl-CoA hydratase/isomerase
C: T33-fn10: engineered DrsE like sulfur reductase
D: T33-fn10: engineered enoyl-CoA hydratase/isomerase
E: T33-fn10: engineered DrsE like sulfur reductase
F: T33-fn10: engineered enoyl-CoA hydratase/isomerase
G: T33-fn10: engineered DrsE like sulfur reductase
H: T33-fn10: engineered enoyl-CoA hydratase/isomerase

A: T33-fn10: engineered DrsE like sulfur reductase
B: T33-fn10: engineered enoyl-CoA hydratase/isomerase
C: T33-fn10: engineered DrsE like sulfur reductase
D: T33-fn10: engineered enoyl-CoA hydratase/isomerase
E: T33-fn10: engineered DrsE like sulfur reductase
F: T33-fn10: engineered enoyl-CoA hydratase/isomerase
G: T33-fn10: engineered DrsE like sulfur reductase
H: T33-fn10: engineered enoyl-CoA hydratase/isomerase


Theoretical massNumber of molelcules
Total (without water)477,52824
Polymers477,52824
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation5_555z,x,y1
crystal symmetry operation9_555y,z,x1
Buried area80070 Å2
ΔGint-628 kcal/mol
Surface area147530 Å2
MethodPISA
2
I: T33-fn10: engineered DrsE like sulfur reductase
J: T33-fn10: engineered enoyl-CoA hydratase/isomerase
K: T33-fn10: engineered DrsE like sulfur reductase
L: T33-fn10: engineered enoyl-CoA hydratase/isomerase
M: T33-fn10: engineered DrsE like sulfur reductase
N: T33-fn10: engineered enoyl-CoA hydratase/isomerase
O: T33-fn10: engineered DrsE like sulfur reductase
P: T33-fn10: engineered enoyl-CoA hydratase/isomerase

I: T33-fn10: engineered DrsE like sulfur reductase
J: T33-fn10: engineered enoyl-CoA hydratase/isomerase
K: T33-fn10: engineered DrsE like sulfur reductase
L: T33-fn10: engineered enoyl-CoA hydratase/isomerase
M: T33-fn10: engineered DrsE like sulfur reductase
N: T33-fn10: engineered enoyl-CoA hydratase/isomerase
O: T33-fn10: engineered DrsE like sulfur reductase
P: T33-fn10: engineered enoyl-CoA hydratase/isomerase

I: T33-fn10: engineered DrsE like sulfur reductase
J: T33-fn10: engineered enoyl-CoA hydratase/isomerase
K: T33-fn10: engineered DrsE like sulfur reductase
L: T33-fn10: engineered enoyl-CoA hydratase/isomerase
M: T33-fn10: engineered DrsE like sulfur reductase
N: T33-fn10: engineered enoyl-CoA hydratase/isomerase
O: T33-fn10: engineered DrsE like sulfur reductase
P: T33-fn10: engineered enoyl-CoA hydratase/isomerase


Theoretical massNumber of molelcules
Total (without water)477,52824
Polymers477,52824
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation5_555z,x,y1
crystal symmetry operation9_555y,z,x1
Buried area80480 Å2
ΔGint-638 kcal/mol
Surface area147400 Å2
MethodPISA
Unit cell
Length a, b, c (Å)225.630, 225.630, 225.630
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number198
Space group name H-MP213
Space group name HallP2ac2ab3
Symmetry operation#1: x,y,z
#2: z,x,y
#3: y,z,x
#4: -y+1/2,-z,x+1/2
#5: z+1/2,-x+1/2,-y
#6: -y,z+1/2,-x+1/2
#7: -z+1/2,-x,y+1/2
#8: -z,x+1/2,-y+1/2
#9: y+1/2,-z+1/2,-x
#10: x+1/2,-y+1/2,-z
#11: -x,y+1/2,-z+1/2
#12: -x+1/2,-y,z+1/2

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Components

#1: Protein
T33-fn10: engineered DrsE like sulfur reductase


Mass: 12122.230 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Sulfurisphaera tokodaii str. 7 (archaea)
Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): LOBSTR
#2: Protein
T33-fn10: engineered enoyl-CoA hydratase/isomerase


Mass: 27671.732 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Novosphingobium aromaticivorans DSM 12444 (bacteria)
Gene: Saro_3457 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): LOBSTR / References: UniProt: A4XEF6

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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: 3.01 Å3/Da / Density % sol: 59.09 %
Crystal growTemperature: 294 K / Method: vapor diffusion, hanging drop / pH: 7.5
Details: 100 mM HEPES pH 7.5, 10% PEG 8000, 10% Ethylene glycol

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-C / Wavelength: 0.97918 Å
DetectorType: DECTRIS EIGER2 X 16M / Detector: PIXEL / Date: Apr 1, 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: 6→92.11 Å / Num. obs: 18590 / % possible obs: 100 % / Redundancy: 10.6 % / Biso Wilson estimate: 356.46 Å2 / CC1/2: 0.997 / Rmerge(I) obs: 0.174 / Rrim(I) all: 0.182 / Net I/σ(I): 9.17
Reflection shell
Resolution (Å)Rmerge(I) obsNum. unique obsCC1/2Rrim(I) allDiffraction-ID
6-6.152.17413680.3832.2851
6.15-6.321.87313570.4971.9651
6.32-6.511.54113140.571.6181
6.51-6.711.18812300.7061.2481
6.71-6.930.87812550.8030.9251
6.93-7.170.75611480.8650.8021
7.17-7.440.67911640.8910.7121
7.44-7.740.45811160.950.4791
7.74-8.090.28610370.9810.2991
8.09-8.480.25610080.980.2681
8.48-8.940.1739620.9910.1811
8.94-9.490.1399210.9930.1461
9.49-10.140.1428320.9930.1491
10.14-10.950.1228290.9940.1281
10.95-120.1127150.9960.1181
12-13.410.1066600.9950.1121
13.41-15.490.0955830.9960.1011
15.49-18.970.0864950.9970.0911
18.97-26.830.083840.9970.0841
26.83-92.110.0642120.9980.0681

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Processing

Software
NameVersionClassification
PHENIX1.20.1_4487refinement
XDS20220110data reduction
XDS20220110data scaling
PHASER2.8.3phasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 6→92.11 Å / SU ML: 0.8147 / Cross valid method: FREE R-VALUE / σ(F): 1.37 / Phase error: 31.5139
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.2481 986 10.02 %
Rwork0.2145 8856 -
obs0.218 9842 99.98 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 400.92 Å2
Refinement stepCycle: LAST / Resolution: 6→92.11 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms21720 0 0 0 21720
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.009522000
X-RAY DIFFRACTIONf_angle_d1.019829744
X-RAY DIFFRACTIONf_chiral_restr0.06963520
X-RAY DIFFRACTIONf_plane_restr0.00373888
X-RAY DIFFRACTIONf_dihedral_angle_d11.06398144
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
6-6.310.37551380.33111245X-RAY DIFFRACTION99.93
6.32-6.710.37781400.30611256X-RAY DIFFRACTION100
6.72-7.230.33641380.29761246X-RAY DIFFRACTION100
7.23-7.960.3011390.25381249X-RAY DIFFRACTION100
7.96-9.110.26571410.21291262X-RAY DIFFRACTION100
9.11-11.470.18971410.17711275X-RAY DIFFRACTION100
11.48-92.110.21871490.18731323X-RAY DIFFRACTION99.93
Refinement TLS params.Method: refined / Origin x: 80.4415769715 Å / Origin y: 45.1882430276 Å / Origin z: 43.114529597 Å
111213212223313233
T3.30855199992 Å2-0.341001095094 Å20.38838674587 Å2-3.73958081664 Å2-0.0525756497287 Å2--3.81364020357 Å2
L0.115111896069 °20.201542198764 °20.524572851051 °2-0.816843190644 °20.762637442109 °2--0.731392720804 °2
S0.271322305259 Å °0.238394417968 Å °-0.288857995908 Å °0.0921758269131 Å °0.135215407006 Å °-0.603362855248 Å °0.219953339762 Å °0.223920822367 Å °-0.0432359620976 Å °
Refinement TLS groupSelection details: all

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