PDB-6i0o: Structure of human IMP dehydrogenase, isoform 2, bound to GTP

Basic information

• Entry: Database: PDB / ID: 6i0o
• Title: Structure of human IMP dehydrogenase, isoform 2, bound to GTP
• Components: Inosine-5'-monophosphate dehydrogenase 2
• Keywords: BIOSYNTHETIC PROTEIN / oxidoreductase / de novo guanine nucleotide biosynthesis

Function / homology

Function:

'de novo' XMP biosynthetic process / lymphocyte proliferation / Purine ribonucleoside monophosphate biosynthesis / IMP dehydrogenase activity / IMP dehydrogenase / GMP biosynthetic process / Azathioprine ADME / GTP biosynthetic process / peroxisomal membrane / cellular response to interleukin-4 / circadian rhythm / secretory granule lumen / ficolin-1-rich granule lumen / Potential therapeutics for SARS / nucleotide binding / Neutrophil degranulation / DNA binding / RNA binding / extracellular exosome / extracellular region / membrane / nucleus / metal ion binding / cytosol / cytoplasm

Domain/homology:

Inosine-5'-monophosphate dehydrogenase / IMP dehydrogenase / GMP reductase, conserved site / IMP dehydrogenase / GMP reductase signature. / IMP dehydrogenase/GMP reductase / IMP dehydrogenase / GMP reductase domain / IMP dehydrogenase / GMP reductase domain / Domain in cystathionine beta-synthase and other proteins. / CBS domain / CBS domain / CBS domain profile. / Aldolase class I / Aldolase-type TIM barrel / TIM Barrel / Alpha-Beta Barrel / Alpha Beta

Component:

GUANOSINE-5'-TRIPHOSPHATE / Inosine-5'-monophosphate dehydrogenase 2

• Biological species: Homo sapiens (human)
• Method: X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.623 Å
• Authors: Buey, R.M. / Fernandez-Justel, D. / Revuelta, J.L.

Funding support

Spain, 1items

Organization	Grant number	Country
	BFU2016-79237-P	Spain

• Citation: Journal: J Mol Biol / Year: 2019; Title: A Nucleotide-Dependent Conformational Switch Controls the Polymerization of Human IMP Dehydrogenases to Modulate their Catalytic Activity.; Authors: David Fernández-Justel / Rafael Núñez / Jaime Martín-Benito / David Jimeno / Adrián González-López / Eva María Soriano / José Luis Revuelta / Rubén M Buey / ; Abstract: Inosine 5'-monophosphate dehydrogenase (IMPDH) catalyzes the rate-limiting step in the de novo GTP biosynthetic pathway and plays essential roles in cell proliferation. As a clinical target, IMPDH has been studied for decades, but it has only been within the last years that we are starting to understand the complexity of the mechanisms of its physiological regulation. Here, we report structural and functional insights into how adenine and guanine nucleotides control a conformational switch that modulates the assembly of the two human IMPDH enzymes into cytoophidia and allosterically regulates their catalytic activity. In vitro reconstituted micron-length cytoophidia-like structures show catalytic activity comparable to unassembled IMPDH but, in turn, are more resistant to GTP/GDP allosteric inhibition. Therefore, IMPDH cytoophidia formation facilitates the accumulation of high levels of guanine nucleotides when the cell requires it. Finally, we demonstrate that most of the IMPDH retinopathy-associated mutations abrogate GTP/GDP-induced allosteric inhibition and alter cytoophidia dynamics.

History

Deposition	Oct 26, 2018	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Jan 30, 2019	Provider: repository / Type: Initial release
Revision 1.1	Feb 20, 2019	Group: Advisory / Data collection / Derived calculations Category: diffrn_source / pdbx_data_processing_status / pdbx_unobs_or_zero_occ_atoms / pdbx_validate_close_contact / struct_conn / struct_conn_type Item: _diffrn_source.pdbx_synchrotron_site
Revision 1.2	Mar 6, 2019	Group: Data collection / Database references / Category: citation / pdbx_database_proc Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3	Jul 10, 2019	Group: Data collection / Category: diffrn_source / Item: _diffrn_source.pdbx_synchrotron_site
Revision 1.4	Jan 24, 2024	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

Assembly

Deposited unit

A: Inosine-5'-monophosphate dehydrogenase 2

B: Inosine-5'-monophosphate dehydrogenase 2

hetero molecules

Summary:

• 118 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	117,951	34
Polymers	112,314	2
Non-polymers	5,637	32
Water	1,603	89

1

A: Inosine-5'-monophosphate dehydrogenase 2

B: Inosine-5'-monophosphate dehydrogenase 2

hetero molecules

A: Inosine-5'-monophosphate dehydrogenase 2

B: Inosine-5'-monophosphate dehydrogenase 2

hetero molecules

A: Inosine-5'-monophosphate dehydrogenase 2

B: Inosine-5'-monophosphate dehydrogenase 2

hetero molecules

A: Inosine-5'-monophosphate dehydrogenase 2

B: Inosine-5'-monophosphate dehydrogenase 2

hetero molecules

Summary:

• defined by author&software
• Evidence: SAXS
• 472 kDa, 8 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	471,804	136
Polymers	449,257	8
Non-polymers	22,547	128
Water	144	8

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	2_445	-x-1,-y-1,z	1
crystal symmetry operation	3_455	-y-1,x,z	1
crystal symmetry operation	4_545	y,-x-1,z	1

Calculated values:

Buried area	67990 Å2
ΔGint	-1658 kcal/mol
Surface area	126460 Å2
Method	PISA

Unit cell

Length a, b, c (Å)	134.119, 134.119, 325.703
Angle α, β, γ (deg.)	90.00, 90.00, 90.00
Int Tables number	97
Space group name H-M	I422

Components

#1: Protein

A B Inosine-5'-monophosphate dehydrogenase 2 / IMPDH 2 / IMPDH-II

Details:

Mass: 56157.137 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: IMPDH2, IMPD2 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P12268, IMP dehydrogenase

#2: Chemical

A-601 A-602 A-603 B-601 B-602 B-603
ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE

Details:

Mass: 523.180 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C₁₀H₁₆N₅O₁₄P₃ / Comment: GTP, energy-carrying molecule*YM

#3: Chemical

A-604 A-605 A-606 A-607 A-608 A-609 A-610 A-611 A-612 A-613 B-604 B-605 B-606 B-607 B-608 B-609 B-610 B-611 B-612 B-613 ...
ChemComp-SO4 / SULFATE ION

Details:

Mass: 96.063 Da / Num. of mol.: 26 / Source method: obtained synthetically / Formula: SO₄

#4: Water

ChemComp-HOH / water

Details:

Mass: 18.015 Da / Num. of mol.: 89 / Source method: isolated from a natural source / Formula: H₂O

Experimental details

Experiment

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

Sample preparation

• Crystal: Density Matthews: 3.26 ų/Da / Density % sol: 62.27 %
• Crystal grow: Temperature: 295 K / Method: vapor diffusion, sitting drop; Details: 25% (w/v) PEG-1500 0.1 M Buffer MIB (malonic acid, imadazole, boric acid), pH 9.0

Data collection

• Diffraction: Mean temperature: 100 K / Serial crystal experiment: N
• Diffraction source: Source: SYNCHROTRON / Site: Diamond / Beamline: I03 / Wavelength: 0.99987 Å
• Detector: Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Sep 21, 2015
• Radiation: Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 0.99987 Å / Relative weight: 1
• Reflection: Resolution: 2.623→81.953 Å / Num. obs: 44889 / % possible obs: 92.6 % / Redundancy: 6.5 % / CC_1/2: 0.997 / R_merge(I) obs: 0.142 / R_pim(I) all: 0.06 / R_rim(I) all: 0.155 / Net I/σ(I): 10.6
• Reflection shell: Resolution: 2.623→2.827 Å / Redundancy: 7 % / R_merge(I) obs: 1.564 / Mean I/σ(I) obs: 1.5 / Num. unique obs: 353 / CC_1/2: 0.408 / R_pim(I) all: 0.635 / R_rim(I) all: 1.689 / % possible all: 47

Processing

Software

Name	Version	Classification
PHENIX	(1.12rc1_2801: ???)	refinement
XDS		data reduction
STARANISO		data scaling
PHASER		phasing

Refinement

Method to determine structure: MOLECULAR REPLACEMENT
Starting model: 4Z87

4z87

Resolution: 2.623→81.953 Å / SU ML: 0.32 / Cross valid method: FREE R-VALUE / σ(F): 1.35 / Phase error: 28.37 / Stereochemistry target values: ML

	Rfactor	Num. reflection	% reflection
Rfree	0.2437	1844	5.06 %
Rwork	0.2193	-	-
obs	0.2206	36415	81.07 %

• Solvent computation: Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL

Refinement step

Cycle: LAST / Resolution: 2.623→81.953 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	6340	0	322	89	6751

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.003	6720
X-RAY DIFFRACTION	f_angle_d	0.615	9177
X-RAY DIFFRACTION	f_dihedral_angle_d	14.245	3887
X-RAY DIFFRACTION	f_chiral_restr	0.043	1063
X-RAY DIFFRACTION	f_plane_restr	0.003	1128

LS refinement shell

Resolution (Å)	Rfactor Rfree	Num. reflection Rfree	Rfactor Rwork	Num. reflection Rwork	Refine-ID	% reflection obs (%)
2.623-2.6939	0.2804	11	0.2996	196	X-RAY DIFFRACTION	6
2.6939-2.7732	0.3139	38	0.3032	702	X-RAY DIFFRACTION	22
2.7732-2.8627	0.3346	91	0.3118	1508	X-RAY DIFFRACTION	47
2.8627-2.965	0.3409	136	0.3034	2472	X-RAY DIFFRACTION	77
2.965-3.0838	0.3448	189	0.291	3196	X-RAY DIFFRACTION	99
3.0838-3.2241	0.2894	165	0.2833	3245	X-RAY DIFFRACTION	100
3.2241-3.3941	0.2638	178	0.2627	3248	X-RAY DIFFRACTION	100
3.3941-3.6068	0.2307	158	0.2272	3281	X-RAY DIFFRACTION	100
3.6068-3.8852	0.2331	185	0.2131	3256	X-RAY DIFFRACTION	100
3.8852-4.2762	0.2087	168	0.189	3280	X-RAY DIFFRACTION	100
4.2762-4.8949	0.2011	156	0.169	3322	X-RAY DIFFRACTION	100
4.8949-6.1668	0.2279	164	0.2097	3365	X-RAY DIFFRACTION	100
6.1668-81.9928	0.2498	205	0.217	3500	X-RAY DIFFRACTION	100

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	2.554	-0.3972	-0.5409	0.6609	-0.579	0.6503	-0.0974	0.1285	-0.1427	-0.0445	0.0025	-0.1487	0.0049	-0.0595	-0	0.6917	-0.0711	0.0202	0.6528	0.0047	0.4507	-46.1679	-46.0051	-23.2293
2	0.151	0.0693	0.1786	0.3314	0.2289	0.2802	0.0836	0.0168	-0.0085	0.5688	-0.1923	-0.871	0.6228	0.932	0.0041	0.7415	-0.0072	0.0874	1.1255	0.1727	0.9485	-18.9637	-31.5522	-45.3425
3	0.738	0.1061	0.0339	0.8486	0.2249	0.2263	-0.0952	0.1425	-0.0176	0.0189	-0.1734	-0.2992	-0.1379	0.3487	0.0074	0.8274	0.01	0.1216	1.1027	0.1356	0.9738	-17.8847	-25.7854	-49.403
4	1.2066	-0.0752	0.1126	0.6784	-0.3995	0.826	-0.1092	-0.3639	0.7592	0.2945	-0.2213	-0.7033	-0.3735	0.3891	-0.1949	0.9265	-0.2659	0.0476	0.9838	0.2383	1.0034	-20.7269	-17.0712	-37.7138
5	1.3172	0.7105	-0.2768	0.7428	-0.7935	0.9224	-0.2549	0.0708	-0.0047	0.3687	0.1775	-0.0606	-0.239	-0.0313	-0.0019	0.6903	-0.0667	0.0427	0.7483	0.0914	0.572	-26.6636	-35.2087	-31.6162
6	1.4362	-1.0446	0.0313	1.6831	-0.8632	1.8212	-0.0986	0.0971	-0.2393	0.0599	-0.0155	-0.7714	0.1889	0.2495	-0	0.6387	-0.0392	0.0027	0.6301	0.0256	0.5781	-33.2629	-57.0818	-17.4728
7	0.8384	0.6863	-0.3427	0.5978	-0.3428	0.1758	0.0004	0.195	0.2188	-0.069	-0.0525	0.0863	-0.1801	-0.0335	-0	0.7203	-0.0337	-0.1145	0.7168	-0.0358	0.474	-46.043	-52.9328	-14.0808
8	0.0983	-0.173	0.2716	0.1644	-0.2277	0.4785	0.0991	0.3061	0.3523	-0.4593	-0.0419	-0.0581	0.1595	0.0779	-0.0328	0.8352	-0.0358	-0.0077	0.7486	-0.0011	0.6066	-47.8683	-48.353	-32.9287
9	1.0505	-0.7204	0.0776	1.4123	-0.9256	1.2542	0.0394	0.3307	0.2019	-0.0908	-0.0007	0.0729	-0.4773	-0.0614	-0.0002	0.5842	-0.0023	0.0178	0.5836	0.0614	0.3785	-57.1007	-43.651	-19.1971
10	1.1962	0.1786	0.1213	2.5642	0.6933	0.7821	0.1104	-0.0957	-0.0017	0.1959	-0.126	0.1537	0.063	-0.136	0	0.406	-0.0403	0.0521	0.3723	0.0277	0.4617	-52.3051	-36.2193	-68.9129
11	1.1083	-0.3482	-0.4907	0.3661	0.1517	1.1225	-0.3712	-0.866	0.0101	0.4494	0.4197	-0.0255	0.3654	-0.584	0.0039	0.9125	-0.0447	0.1433	0.7749	-0.1571	0.7781	-43.4946	-8.5144	-44.7146
12	1.7208	1.4317	-0.0403	2.9963	0.7567	1.0658	0.1445	-0.1622	0.2238	0.1874	-0.1308	-0.0629	-0.0072	0.0468	0.0046	0.5557	-0.0432	0.0605	0.49	-0.0054	0.5347	-43.6763	-15.3942	-61.2567
13	2.4275	0.4829	0.2858	1.5974	0.2475	1.5583	0.0467	-0.0413	0.0237	0.1413	0.02	0.1006	0.0402	0.0394	0.0012	0.3501	-0.0465	0.0638	0.2897	0.0252	0.4753	-58.1454	-40.7564	-74.9317

Refinement TLS group

ID	Refine-ID	Refine TLS-ID	Selection details
1	X-RAY DIFFRACTION	1	chain 'A' and (resid 11 through 110 )
2	X-RAY DIFFRACTION	2	chain 'A' and (resid 111 through 137 )
3	X-RAY DIFFRACTION	3	chain 'A' and (resid 138 through 174 )
4	X-RAY DIFFRACTION	4	chain 'A' and (resid 175 through 204 )
5	X-RAY DIFFRACTION	5	chain 'A' and (resid 205 through 266 )
6	X-RAY DIFFRACTION	6	chain 'A' and (resid 267 through 342 )
7	X-RAY DIFFRACTION	7	chain 'A' and (resid 343 through 369 )
8	X-RAY DIFFRACTION	8	chain 'A' and (resid 370 through 452 )
9	X-RAY DIFFRACTION	9	chain 'A' and (resid 453 through 499 )
10	X-RAY DIFFRACTION	10	chain 'B' and (resid 11 through 126 )
11	X-RAY DIFFRACTION	11	chain 'B' and (resid 127 through 174 )
12	X-RAY DIFFRACTION	12	chain 'B' and (resid 175 through 266 )
13	X-RAY DIFFRACTION	13	chain 'B' and (resid 267 through 497 )