PDB-8cm0: Rhs2-CT endonuclease toxin in complex with cognate immunity prote...

ID or keywords:

Entry

Database: PDB / ID: 8cm0

Title

Rhs2-CT endonuclease toxin in complex with cognate immunity protein RhsI2

Components

Rhs2CT
RhsI2

Keywords

ANTIMICROBIAL PROTEIN / T6SS / effector-immunity compex / HNH endonuclease / DNAse / S. marcescens DB10

Function / homology

ACETATE ION

Function and homology information

Biological species

Serratia marcescens (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.17 Å

Authors

Pankov, G. / Hunter, W.N. / Coulthurst, S.J.

Funding support

United Kingdom, 2items

Citation

Journal: To Be Published
Title: Rhs2-CT endonuclease toxin in complex with cognate immunity protein RhsI2
Authors: Pankov, G. / Coulthurst, C. / Hunter, W.N.

History

Deposition	Feb 17, 2023	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Aug 28, 2024	Provider: repository / Type: Initial release

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8cm0.cif.gz	76.4 KB	Display	PDBx/mmCIF format
PDB format	pdb8cm0.ent.gz		Display	PDB format
PDBx/mmJSON format	8cm0.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/cm/8cm0 ftp://data.pdbj.org/pub/pdb/validation_reports/cm/8cm0	HTTPS FTP

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Related structure data

Similar structure data	Similarity search - Function & homologyF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

BBB: RhsI2

AAA: Rhs2CT

hetero molecules

36.7 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

Idetical with deposited unit
defined by author&software
Evidence: gel filtration, Rhs2CT-RhsI2 copurify together in SEC chromatography with the elutionvolume consistent with the mass of the heterodimer, RhsI2 protects susceptible cells from Rhs2CT ...Evidence: gel filtration, Rhs2CT-RhsI2 copurify together in SEC chromatography with the elutionvolume consistent with the mass of the heterodimer, RhsI2 protects susceptible cells from Rhs2CT intoxication in competition assays

Unit cell

Length a, b, c (Å)	37.006, 81.931, 101.371
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	19
Space group name H-M	P2₁2₁2₁

#1: Protein	RhsI2 Mass: 18682.303 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Serratia marcescens (bacteria) / Production host: Escherichia coli BL21(DE3) (bacteria)
#2: Protein	Rhs2CT Mass: 17797.018 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Serratia marcescens (bacteria) / Production host: Escherichia coli BL21(DE3) (bacteria)
#3: Chemical	ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL Mass: 62.068 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₂H₆O₂
#4: Chemical	ChemComp-ACT / ACETATE ION Mass: 59.044 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C₂H₃O₂
#5: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 131 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	N

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Experiment

Experiment	Method: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

Crystal	Density Matthews: 2.13 Å³/Da / Density % sol: 42.41 %
Crystal grow	Temperature: 296 K / Method: vapor diffusion, sitting drop Details: Crystals were grown in a condition containing 0.2 uL of 6.5 mg/mL Rhs2CT-RhsI2 (in 20 mM MES & 20 mM imidazole pH 6.0, 150 mM NaCl, 1 mM MgCl2, 0.5 mM TCEP) and 0.2 uL of reservoir solution ...Details: Crystals were grown in a condition containing 0.2 uL of 6.5 mg/mL Rhs2CT-RhsI2 (in 20 mM MES & 20 mM imidazole pH 6.0, 150 mM NaCl, 1 mM MgCl2, 0.5 mM TCEP) and 0.2 uL of reservoir solution (0.2 M ammonium acetate, 0.1 M sodium acetate pH 5.0, 20% PEG4K).

Crystal

Density Matthews: 2.13 Å³/Da / Density % sol: 42.41 %

Crystal grow

Temperature: 296 K / Method: vapor diffusion, sitting drop
Details: Crystals were grown in a condition containing 0.2 uL of 6.5 mg/mL Rhs2CT-RhsI2 (in 20 mM MES & 20 mM imidazole pH 6.0, 150 mM NaCl, 1 mM MgCl2, 0.5 mM TCEP) and 0.2 uL of reservoir solution ...

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SYNCHROTRON / Site: ESRF / Beamline: ID23-1 / Wavelength: 0.8731 Å
Detector	Type: DECTRIS PILATUS3 X 2M / Detector: PIXEL / Date: Jun 9, 2022
Radiation	Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 0.8731 Å / Relative weight: 1
Reflection	Resolution: 2.17→37.98 Å / Num. obs: 17007 / % possible obs: 100 % / Redundancy: 6 % / CC_1/2: 0.993 / R_merge(I) obs: 0.157 / R_pim(I) all: 0.07 / Net I/σ(I): 8.7
Reflection shell	Resolution: 2.17→2.24 Å / R_merge(I) obs: 1.511 / Num. unique obs: 1408 / CC_1/2: 0.542 / R_pim(I) all: 0.661 / % possible all: 100

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
XDS		data reduction
xia2		data scaling
PHASES		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 2.17→37.98 Å / Cor.coef. F_o:F_c: 0.95 / Cor.coef. F_o:F_c free: 0.916 / SU B: 6.765 / SU ML: 0.171 / Cross valid method: FREE R-VALUE / ESU R: 0.253 / ESU R Free: 0.212
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.2526	842	4.968 %
R_work	0.195	16107	-
all	0.198	-	-
obs	-	16949	99.906 %

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK BULK SOLVENT

Displacement parameters

B_iso mean: 34.743 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	0.916 Å²	-0 Å²	-0 Å²
2-	-	-1.073 Å²	-0 Å²
3-	-	-	0.157 Å²

Refinement step

Cycle: LAST / Resolution: 2.17→37.98 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	2185	0	16	131	2332

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.007	0.013	2251
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.015	2129
X-RAY DIFFRACTION	r_angle_refined_deg	1.477	1.645	3044
X-RAY DIFFRACTION	r_angle_other_deg	1.309	1.583	4886
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	6.934	5	270
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	34.427	21.545	123
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	18.868	15	378
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	18.442	15	17
X-RAY DIFFRACTION	r_chiral_restr	0.075	0.2	285
X-RAY DIFFRACTION	r_gen_planes_refined	0.007	0.02	2543
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	547
X-RAY DIFFRACTION	r_nbd_refined	0.206	0.2	440
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.196	0.2	1844
X-RAY DIFFRACTION	r_nbtor_refined	0.168	0.2	1071
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.08	0.2	1054
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.146	0.2	105
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.139	0.2	15
X-RAY DIFFRACTION	r_nbd_other	0.217	0.2	39
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.138	0.2	5
X-RAY DIFFRACTION	r_mcbond_it	2.792	3.345	1092
X-RAY DIFFRACTION	r_mcbond_other	2.79	3.339	1088
X-RAY DIFFRACTION	r_mcangle_it	4.349	4.989	1356
X-RAY DIFFRACTION	r_mcangle_other	4.347	4.993	1357
X-RAY DIFFRACTION	r_scbond_it	3.676	3.868	1159
X-RAY DIFFRACTION	r_scbond_other	3.675	3.869	1160
X-RAY DIFFRACTION	r_scangle_it	6.025	5.579	1688
X-RAY DIFFRACTION	r_scangle_other	6.024	5.58	1689
X-RAY DIFFRACTION	r_lrange_it	8.126	38.56	2520
X-RAY DIFFRACTION	r_lrange_other	8.125	38.568	2521

LS refinement shell

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Refine-ID	% reflection obs (%)
2.17-2.226	0.329	61	0.27	1141	X-RAY DIFFRACTION	100
2.226-2.287	0.352	66	0.258	1136	X-RAY DIFFRACTION	99.6683
2.287-2.353	0.278	38	0.233	1123	X-RAY DIFFRACTION	99.6567
2.353-2.426	0.303	59	0.208	1074	X-RAY DIFFRACTION	100
2.426-2.505	0.277	61	0.198	1051	X-RAY DIFFRACTION	99.9101
2.505-2.593	0.287	45	0.198	1001	X-RAY DIFFRACTION	100
2.593-2.691	0.287	47	0.208	983	X-RAY DIFFRACTION	100
2.691-2.8	0.32	52	0.203	938	X-RAY DIFFRACTION	99.8991
2.8-2.925	0.299	46	0.202	930	X-RAY DIFFRACTION	100
2.925-3.067	0.315	43	0.195	884	X-RAY DIFFRACTION	100
3.067-3.233	0.238	47	0.199	812	X-RAY DIFFRACTION	100
3.233-3.428	0.246	41	0.194	792	X-RAY DIFFRACTION	99.8801
3.428-3.664	0.198	41	0.17	749	X-RAY DIFFRACTION	99.8736
3.664-3.957	0.245	44	0.174	685	X-RAY DIFFRACTION	100
3.957-4.332	0.235	26	0.168	650	X-RAY DIFFRACTION	100
4.332-4.841	0.242	30	0.181	602	X-RAY DIFFRACTION	100
4.841-5.585	0.181	26	0.181	526	X-RAY DIFFRACTION	100
5.585-6.827	0.282	29	0.2	450	X-RAY DIFFRACTION	100
6.827-9.599	0.167	24	0.157	359	X-RAY DIFFRACTION	100
9.599-37.98	0.187	16	0.231	221	X-RAY DIFFRACTION	98.75

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