PDB-8snf: Crystal structure of metformin hydrolase (MfmAB) from Pseudomonas...

ID or keywords:

Entry

Database: PDB / ID: 8snf

Title

Crystal structure of metformin hydrolase (MfmAB) from Pseudomonas mendocina sp. MET-2 with Ni2+2 bound

Components

metformin hydrolase subunit A
metformin hydrolase subunit B

Keywords

HYDROLASE / ureohydrolase / metformin / metalloenzyme / guanylurea

Function / homology

NICKEL (II) ION

Function and homology information

Biological species

Pseudomonas mendocina (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2.3 Å

Authors

Tassoulas, L.J. / Rankin, J.A. / Elias, M.H. / Wackett, L.P.

Funding support

United States, 1items

Citation

Journal: Proc.Natl.Acad.Sci.USA / Year: 2024
Title: Dinickel enzyme evolved to metabolize the pharmaceutical metformin and its implications for wastewater and human microbiomes.
Authors: Tassoulas, L.J. / Rankin, J.A. / Elias, M.H. / Wackett, L.P.

History

Deposition	Apr 27, 2023	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Mar 6, 2024	Provider: repository / Type: Initial release

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	8snf.cif.gz	999 KB	Display	PDBx/mmCIF format
PDB format	pdb8snf.ent.gz	638.3 KB	Display	PDB format
PDBx/mmJSON format	8snf.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	8snf_validation.pdf.gz	2.4 MB	Display	wwPDB validaton report
Full document	8snf_full_validation.pdf.gz	2.4 MB	Display
Data in XML	8snf_validation.xml.gz	70.4 KB	Display
Data in CIF	8snf_validation.cif.gz	98.2 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/sn/8snf ftp://data.pdbj.org/pub/pdb/validation_reports/sn/8snf	HTTPS FTP

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Related structure data

Related structure data	8snkC 8sp2C C: citing same article (ref.)
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

A: metformin hydrolase subunit B

B: metformin hydrolase subunit B

C: metformin hydrolase subunit B

D: metformin hydrolase subunit A

E: metformin hydrolase subunit B

F: metformin hydrolase subunit A

hetero molecules

234 kDa, 6 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: metformin hydrolase subunit B

B: metformin hydrolase subunit B

D: metformin hydrolase subunit A

hetero molecules

defined by author&software
Evidence: gel filtration
117 kDa, 3 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

C: metformin hydrolase subunit B

E: metformin hydrolase subunit B

F: metformin hydrolase subunit A

hetero molecules

defined by author&software
Evidence: gel filtration
117 kDa, 3 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	83.400, 96.400, 96.700
Angle α, β, γ (deg.)	115.500, 106.200, 101.100
Int Tables number	1
Space group name H-M	P1

#1: Protein	metformin hydrolase subunit B Mass: 38024.281 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pseudomonas mendocina (bacteria) / Strain: MET-2 / Gene: mfmB / Plasmid: pET28 / Production host: Escherichia coli BL21(DE3) (bacteria)
#2: Protein	metformin hydrolase subunit A Mass: 40723.984 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pseudomonas mendocina (bacteria) / Strain: MET-2 / Gene: mfmA / Plasmid: pET28 / Production host: Escherichia coli BL21(DE3) (bacteria)
#3: Chemical	ChemComp-NI / NICKEL (II) ION Mass: 58.693 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Ni / Feature type: SUBJECT OF INVESTIGATION
#4: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 330 / Source method: isolated from a natural source / Formula: H₂O
Has ligand of interest	Y

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Experiment

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

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Sample preparation

Crystal	Density Matthews: 2.69 Å³/Da / Density % sol: 54.29 %
Crystal grow	Temperature: 291 K / Method: vapor diffusion, hanging drop / pH: 7.5 Details: 1 uL 10 mg/mL protein + 1 uL 14.5% w/v PEG3350, 0.1 M Bis-Tris propane, 0.2 M NaNO3, pH 7.5

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

APS

/ Beamline: 23-ID-D / Wavelength: 1.0332 Å

Detector

Type: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Feb 26, 2023

Radiation

Monochromator: M / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray

Radiation wavelength

Wavelength: 1.0332 Å / Relative weight: 1

Reflection

Resolution: 2.3→80 Å / Num. obs: 99100 / % possible obs: 91.3 % / Redundancy: 2.408 % / CC_1/2: 0.998 / Net I/σ(I): 13.3

Reflection shell

Resolution (Å)	Mean I/σ(I) obs	Num. unique obs	CC_1/2	Diffraction-ID	% possible all
2.3-2.4	3.07	11503	0.96	1	88.6
2.4-2.5	5	10312	0.979	1	93.7

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0411	refinement
XDS	VERSION Jan 10, 2022 BUILT=20220820	data reduction
XSCALE	VERSION Jan 10, 2022 BUILT=20220820	data scaling
MOLREP	Version 11.0	phasing
Coot	0.9.8.1	model building

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT / Resolution: 2.3→19.865 Å / Cor.coef. F_o:F_c: 0.967 / Cor.coef. F_o:F_c free: 0.944 / WR_factor R_free: 0.227 / WR_factor R_work: 0.177 / SU B: 16.573 / SU ML: 0.182 / Average fsc free: 0.9534 / Average fsc work: 0.9703 / Cross valid method: THROUGHOUT / ESU R: 0.296 / ESU R Free: 0.217
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.2194	5056	5.102 %
R_work	0.1719	94042	-
all	0.174	-	-
obs	-	99098	91.623 %

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: 76.693 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-3.769 Å²	2.669 Å²	3.824 Å²
2-	-	-2.678 Å²	2.882 Å²
3-	-	-	-0.966 Å²

Refinement step

Cycle: LAST / Resolution: 2.3→19.865 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	15458	0	4	330	15792

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.007	0.012	15806
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.016	14758
X-RAY DIFFRACTION	r_angle_refined_deg	1.449	1.645	21390
X-RAY DIFFRACTION	r_angle_other_deg	0.493	1.573	33978
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.287	5	1988
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	8.088	5	126
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	17.418	10	2600
X-RAY DIFFRACTION	r_dihedral_angle_6_deg	15.517	10	728
X-RAY DIFFRACTION	r_chiral_restr	0.072	0.2	2300
X-RAY DIFFRACTION	r_gen_planes_refined	0.007	0.02	18834
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	3702
X-RAY DIFFRACTION	r_nbd_refined	0.221	0.2	3599
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.205	0.2	14315
X-RAY DIFFRACTION	r_nbtor_refined	0.184	0.2	7770
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.081	0.2	8385
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.171	0.2	545
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.075	0.2	3
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.192	0.2	4
X-RAY DIFFRACTION	r_nbd_other	0.179	0.2	18
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.195	0.2	2
X-RAY DIFFRACTION	r_mcbond_it	4.992	6.077	7982
X-RAY DIFFRACTION	r_mcbond_other	4.992	6.077	7982
X-RAY DIFFRACTION	r_mcangle_it	6.472	10.915	9960
X-RAY DIFFRACTION	r_mcangle_other	6.472	10.915	9961
X-RAY DIFFRACTION	r_scbond_it	6.167	6.662	7824
X-RAY DIFFRACTION	r_scbond_other	6.167	6.663	7825
X-RAY DIFFRACTION	r_scangle_it	8.718	11.998	11430
X-RAY DIFFRACTION	r_scangle_other	8.718	11.998	11431
X-RAY DIFFRACTION	r_lrange_it	10.232	56.663	17925
X-RAY DIFFRACTION	r_lrange_other	10.237	56.652	17887

LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Total num. of bins used: 20

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	R_factor all	Num. reflection all	Fsc free	Fsc work	% reflection obs (%)	WR_factor R_work
2.3-2.359	0.438	315	0.369	6433	0.372	7882	0.914	0.931	85.6128	0.314
2.359-2.423	0.328	324	0.243	6858	0.247	7669	0.93	0.95	93.6498	0.206
2.423-2.492	0.313	331	0.232	6774	0.236	7519	0.937	0.957	94.4939	0.2
2.492-2.567	0.287	306	0.249	6455	0.251	7278	0.917	0.947	92.8964	0.22
2.567-2.65	0.246	300	0.219	6239	0.22	7033	0.952	0.969	92.976	0.192
2.65-2.741	0.247	291	0.232	5867	0.232	6842	0.928	0.961	90.0029	0.21
2.741-2.843	0.272	302	0.198	5777	0.201	6636	0.95	0.976	91.6064	0.178
2.843-2.956	0.261	303	0.189	5571	0.193	6331	0.958	0.979	92.7815	0.176
2.956-3.084	0.267	291	0.193	5328	0.196	6123	0.953	0.978	91.7687	0.182
3.084-3.231	0.257	271	0.192	5066	0.196	5813	0.96	0.978	91.8115	0.185
3.231-3.401	0.227	303	0.177	4719	0.18	5556	0.967	0.982	90.3888	0.175
3.401-3.601	0.219	277	0.173	4429	0.176	5280	0.969	0.982	89.1288	0.173
3.601-3.84	0.285	226	0.223	3989	0.227	4947	0.946	0.972	85.2032	0.223
3.84-4.135	0.188	229	0.13	3925	0.133	4619	0.978	0.99	89.9329	0.144
4.135-4.51	0.172	207	0.115	3817	0.118	4295	0.982	0.992	93.6903	0.135
4.51-5.01	0.156	175	0.109	3496	0.111	3866	0.986	0.993	94.956	0.132
5.01-5.725	0.154	175	0.129	3093	0.131	3456	0.985	0.991	94.5602	0.152
5.725-6.871	0.202	158	0.137	2712	0.14	2958	0.978	0.989	97.025	0.164
6.871-9.183	0.171	158	0.129	2107	0.132	2347	0.983	0.99	96.5062	0.163
9.183-19.865	0.157	114	0.153	1387	0.153	1542	0.984	0.987	97.3411	0.199

Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

ID	L11 (°²)	L12 (°²)	L13 (°²)	L22 (°²)	L23 (°²)	L33 (°²)	S11 (Å °)	S12 (Å °)	S13 (Å °)	S21 (Å °)	S22 (Å °)	S23 (Å °)	S31 (Å °)	S32 (Å °)	S33 (Å °)	T11 (Å²)	T12 (Å²)	T13 (Å²)	T22 (Å²)	T23 (Å²)	T33 (Å²)	Origin x (Å)	Origin y (Å)	Origin z (Å)
1	0.3663	-0.2509	-0.0695	0.5687	-0.172	0.2618	-0.0523	-0.1007	0.0391	0.1001	0.0241	-0.0291	-0.0107	0.0457	0.0281	0.1868	-0.0298	-0.0795	0.2153	-0.0675	0.0793	-0.8198	-2.7594	2.4634
2	0.1781	-0.0045	0.2472	0.9866	-0.0545	0.5027	0.0194	0.0751	-0.0717	0.1279	-0.1261	0.2347	0.001	0.1192	0.1067	0.0969	-0.0222	0.0264	0.0511	-0.0515	0.3623	-10.7833	-35.358	-16.193
3	0.1483	0.2507	0.0018	0.6479	-0.3578	1.0486	0.0638	-0.0832	0.0116	-0.0046	-0.0936	0.0929	0.0707	0.2014	0.0298	0.1272	-0.0016	-0.0297	0.2825	0.0952	0.0665	-42.059	-17.0716	13.3306
4	0.4446	0.0128	-0.0272	0.1979	0.1003	0.1216	0.0423	0.1667	0.0518	0.0307	0.0005	-0.08	0.0251	-0.0203	-0.0429	0.1841	-0.0185	-0.023	0.1885	-0.0155	0.0822	-11.2648	0.5691	-33.4608
5	0.5365	0.1735	-0.3353	0.3587	-0.1257	0.2565	-0.0845	0.1262	-0.0542	0.0003	0.0425	0.0068	0.0427	-0.0638	0.042	0.1442	-0.0393	-0.0801	0.1979	-0.0724	0.1067	-50.0017	-15.0371	-24.667
6	0.4504	-0.0097	-0.0205	0.1207	0.016	0.1823	0.0781	-0.0101	0.1948	0.051	0.0168	0.0436	0.0401	-0.0141	-0.0949	0.1989	-0.0007	0.0077	0.0921	-0.0397	0.1851	-37.3783	16.2932	-8.1679

Refinement TLS group

Selection: ALL

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