PDB-7pob: An Irreversible, Promiscuous and Highly Thermostable Claisen-Cond...

ID or keywords:

Entry

Database: PDB / ID: 7pob

Title

An Irreversible, Promiscuous and Highly Thermostable Claisen-Condensation Biocatalyst Drives the Synthesis of Substituted Pyrroles

Components

8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

Keywords

SYNTHASE / Biocatalyst / cascade / chemo-enzymatic / thermophilic / promiscuous / alpha-oxoamine synthase / pyrrole

Function / homology

Function and homology information

glycine C-acetyltransferase / glycine C-acetyltransferase activity / 8-amino-7-oxononanoate synthase / 8-amino-7-oxononanoate synthase activity / biotin biosynthetic process / pyridoxal phosphate binding
Similarity search - Function

Biological species

Thermus thermophilus (bacteria)

Method

X-RAY DIFFRACTION /

SYNCHROTRON /

MOLECULAR REPLACEMENT / Resolution: 2 Å

Authors

Basle, A. / Ashley, B. / Campopiano, D. / Marles-Wright, J.

Funding support

United Kingdom, 1items

Citation

Journal: Acs Sustain Chem Eng / Year: 2023
Title: Versatile Chemo-Biocatalytic Cascade Driven by a Thermophilic and Irreversible C-C Bond-Forming alpha-Oxoamine Synthase.
Authors: Ashley, B. / Basle, A. / Sajjad, M. / El Ashram, A. / Kelis, P. / Marles-Wright, J. / Campopiano, D.J.

History

Deposition	Sep 8, 2021	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Sep 21, 2022	Provider: repository / Type: Initial release
Revision 1.1	Jan 31, 2024	Group: Data collection / Refinement description Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model / struct_ncs_dom_lim Item: _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id ..._struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id
Revision 1.2	Feb 28, 2024	Group: Database references / Category: citation / citation_author Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

Structure viewer	Molecule: MolmilJmol/JSmol

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PDBx/mmCIF format	7pob.cif.gz	607.1 KB	Display	PDBx/mmCIF format
PDB format	pdb7pob.ent.gz	486.5 KB	Display	PDB format
PDBx/mmJSON format	7pob.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/po/7pob ftp://data.pdbj.org/pub/pdb/validation_reports/po/7pob	HTTPS FTP

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Related structure data

Related structure data	7poaC 7pocC 3tqxS S: Starting model for refinement C: citing same article (ref.)
Similar structure data	Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI
Related items in Molecule of the Month	#181 - Jan 2015 Cascade and CRISPR similarity (1)

Deposited unit

A: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

B: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

C: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

D: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

hetero molecules

187 kDa, 4 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

A: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

D: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

hetero molecules

defined by author&software
93.7 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

2

B: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

C: 8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase

hetero molecules

defined by author&software
93.7 kDa, 2 polymers
Search similar-shape structures of this assembly by Omokage search (details)

Unit cell

Length a, b, c (Å)	67.100, 57.135, 189.443
Angle α, β, γ (deg.)	90.000, 92.350, 90.000
Int Tables number	4
Space group name H-M	P12₁1

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	A
2	1	B
3	2	A
4	2	C
5	3	A
6	3	D
7	4	B
8	4	C
9	5	B
10	5	D
11	6	C
12	6	D

NCS domain segments:

Beg auth comp-ID: GLY / Beg label comp-ID: GLY / End auth comp-ID: ARG / End label comp-ID: ARG / Refine code: 1 / Auth seq-ID: 2 - 396 / Label seq-ID: 27 - 421

Dom-ID	Component-ID	Ens-ID	Auth asym-ID	Label asym-ID
1	1	1	A	A
2	1	1	B	B
3	2	2	A	A
4	2	2	C	C
5	3	3	A	A
6	3	3	D	D
7	4	4	B	B
8	4	4	C	C
9	5	5	B	B
10	5	5	D	D
11	6	6	C	C
12	6	6	D	D

NCS ensembles :

ID	Details
1	Local NCS retraints between domains: 1 2
2	Local NCS retraints between domains: 3 4
3	Local NCS retraints between domains: 5 6
4	Local NCS retraints between domains: 7 8
5	Local NCS retraints between domains: 9 10
6	Local NCS retraints between domains: 11 12

#1: Protein	8-amino-7-oxononanoate synthase/2-amino-3-ketobutyrate coenzyme A ligase / AONS/AKB ligase / 7-keto-8-amino-pelargonic acid synthase / 7-KAP synthase / KAPA synthase / 8- ...AONS/AKB ligase / 7-keto-8-amino-pelargonic acid synthase / 7-KAP synthase / KAPA synthase / 8-amino-7-ketopelargonate synthase / Alpha-oxoamine synthase / Glycine acetyltransferase Mass: 46585.145 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Thermus thermophilus (bacteria) / Gene: TTHA1582 / Production host: Escherichia coli (E. coli) References: UniProt: Q5SHZ8, glycine C-acetyltransferase, 8-amino-7-oxononanoate synthase
#2: Chemical	ChemComp-PLP / PYRIDOXAL-5'-PHOSPHATE / VITAMIN B6 Phosphate Mass: 247.142 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C₈H₁₀NO₆P / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical	ChemComp-NA / SODIUM ION Mass: 22.990 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Na
#4: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 493 / 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: 1.95 Å³/Da / Density % sol: 36.83 %
Crystal grow	Temperature: 293.15 K / Method: vapor diffusion, sitting drop Details: 30 mM magnesium chloride, 30 mM calcium chloride, 100 mM Hepes/MOPS pH 7.5, 12.5 % (v/v) MPD, 12.5 % (w/v) PEG 1000 and 12.5 % (w/v) PEG 3350

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Data collection

Diffraction

Mean temperature: 100 K / Serial crystal experiment: N

Diffraction source

Source:

SYNCHROTRON / Site:

Diamond

/ Beamline: I03 / Wavelength: 0.9796 Å

Detector

Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: May 5, 2021

Radiation

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

Radiation wavelength

Wavelength: 0.9796 Å / Relative weight: 1

Reflection

Resolution: 2→94.642 Å / Num. obs: 97294 / % possible obs: 99.9 % / Redundancy: 12.5 % / CC_1/2: 0.999 / R_merge(I) obs: 0.166 / R_pim(I) all: 0.071 / R_rim(I) all: 0.18 / Net I/σ(I): 11

Reflection shell

Diffraction-ID: 1

Resolution (Å)	Redundancy (%)	R_merge(I) obs	Num. unique obs	CC_1/2	R_pim(I) all	R_rim(I) all
10.95-94.64	12.2	0.034	653	1	0.014	0.037
2-2.03	13	1.544	4797	0.739	0.642	1.674

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
REFMAC	5.8.0267	refinement
Aimless		data scaling
pointless		data scaling
xia2		data scaling
PHASER		phasing
Coot		model building
BUCCANEER		model building
MolProbity		model building
autoPROC		data reduction

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 3TQX

3tqx

Resolution: 2→94.642 Å / Cor.coef. F_o:F_c: 0.954 / Cor.coef. F_o:F_c free: 0.911 / WR_factor R_free: 0.237 / WR_factor R_work: 0.185 / SU B: 8.328 / SU ML: 0.206 / Average fsc free: 0.809 / Average fsc work: 0.8188 / Cross valid method: FREE R-VALUE / ESU R: 0.24 / ESU R Free: 0.198
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.2614	4850	4.988 %
R_work	0.2114	92378	-
all	0.214	-	-
obs	-	97228	99.745 %

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: 35.896 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-3.061 Å²	0 Å²	-1.169 Å²
2-	-	-2.852 Å²	-0 Å²
3-	-	-	5.988 Å²

Refinement step

Cycle: LAST / Resolution: 2→94.642 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	12244	0	61	493	12798

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.01	0.013	12548
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.017	12273
X-RAY DIFFRACTION	r_angle_refined_deg	1.688	1.64	16972
X-RAY DIFFRACTION	r_angle_other_deg	1.354	1.579	28198
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.85	5	1576
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	29.396	20.988	648
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	16.131	15	2160
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	18.777	15	104
X-RAY DIFFRACTION	r_chiral_restr	0.084	0.2	1572
X-RAY DIFFRACTION	r_gen_planes_refined	0.008	0.02	14080
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	2829
X-RAY DIFFRACTION	r_nbd_refined	0.206	0.2	2608
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.183	0.2	11564
X-RAY DIFFRACTION	r_nbtor_refined	0.162	0.2	6109
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.082	0.2	5972
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.158	0.2	573
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.191	0.2	4
X-RAY DIFFRACTION	r_metal_ion_refined	0.323	0.2	1
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.192	0.2	29
X-RAY DIFFRACTION	r_nbd_other	0.248	0.2	68
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.268	0.2	12
X-RAY DIFFRACTION	r_symmetry_metal_ion_refined	0.076	0.2	1
X-RAY DIFFRACTION	r_mcbond_it	3.034	3.544	6316
X-RAY DIFFRACTION	r_mcbond_other	3.031	3.544	6315
X-RAY DIFFRACTION	r_mcangle_it	4.126	5.304	7888
X-RAY DIFFRACTION	r_mcangle_other	4.126	5.304	7889
X-RAY DIFFRACTION	r_scbond_it	3.992	3.996	6232
X-RAY DIFFRACTION	r_scbond_other	3.993	3.996	6229
X-RAY DIFFRACTION	r_scangle_it	5.901	5.811	9084
X-RAY DIFFRACTION	r_scangle_other	5.901	5.811	9085
X-RAY DIFFRACTION	r_lrange_it	7.187	41.862	13880
X-RAY DIFFRACTION	r_lrange_other	7.186	41.821	13810
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.034	0.05	12716
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.08	0.05	12311
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.076	0.05	12320
X-RAY DIFFRACTION	r_ncsr_local_group_4	0.08	0.05	12329
X-RAY DIFFRACTION	r_ncsr_local_group_5	0.077	0.05	12331
X-RAY DIFFRACTION	r_ncsr_local_group_6	0.05	0.05	12633

Refine LS restraints NCS

Ens-ID	Dom-ID	Auth asym-ID	Refine-ID	Type	Rms dev position (Å)	Weight position
1	1	A	X-RAY DIFFRACTION	Local ncs	0.0337	0.05009
1	2	B	X-RAY DIFFRACTION	Local ncs	0.0337	0.05009
2	3	A	X-RAY DIFFRACTION	Local ncs	0.07966	0.05007
2	4	C	X-RAY DIFFRACTION	Local ncs	0.07966	0.05007
3	5	A	X-RAY DIFFRACTION	Local ncs	0.07587	0.05007
3	6	D	X-RAY DIFFRACTION	Local ncs	0.07587	0.05007
4	7	B	X-RAY DIFFRACTION	Local ncs	0.07984	0.05007
4	8	C	X-RAY DIFFRACTION	Local ncs	0.07984	0.05007
5	9	B	X-RAY DIFFRACTION	Local ncs	0.07719	0.05007
5	10	D	X-RAY DIFFRACTION	Local ncs	0.07719	0.05007
6	11	C	X-RAY DIFFRACTION	Local ncs	0.04997	0.05009
6	12	D	X-RAY DIFFRACTION	Local ncs	0.04997	0.05009

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
2-2.052	0.355	349	0.354	6771	0.355	7163	0.453	0.458	99.3997	0.34
2.052-2.108	0.35	300	0.342	6653	0.343	6991	0.525	0.54	99.4564	0.326
2.108-2.169	0.33	289	0.309	6432	0.31	6752	0.662	0.677	99.5409	0.287
2.169-2.236	0.305	364	0.288	6188	0.289	6574	0.753	0.739	99.6654	0.261
2.236-2.309	0.3	323	0.261	6078	0.262	6419	0.789	0.81	99.7196	0.232
2.309-2.39	0.32	323	0.233	5886	0.237	6228	0.815	0.858	99.6949	0.205
2.39-2.48	0.31	283	0.242	5654	0.245	5949	0.832	0.85	99.7983	0.214
2.48-2.582	0.287	286	0.222	5466	0.225	5761	0.863	0.885	99.8438	0.192
2.582-2.696	0.287	274	0.219	5214	0.223	5491	0.868	0.897	99.9454	0.19
2.696-2.828	0.276	287	0.215	5033	0.219	5321	0.896	0.906	99.9812	0.185
2.828-2.981	0.269	286	0.198	4693	0.202	4981	0.91	0.925	99.9599	0.172
2.981-3.161	0.26	275	0.209	4557	0.212	4835	0.909	0.916	99.938	0.185
3.161-3.379	0.242	213	0.187	4250	0.189	4469	0.924	0.939	99.8657	0.167
3.379-3.65	0.221	191	0.185	3961	0.187	4157	0.938	0.946	99.8797	0.17
3.65-3.997	0.215	200	0.167	3656	0.17	3863	0.947	0.959	99.8188	0.155
3.997-4.468	0.228	157	0.149	3337	0.152	3503	0.941	0.967	99.7431	0.141
4.468-5.157	0.199	148	0.162	2974	0.163	3126	0.962	0.968	99.872	0.158
5.157-6.312	0.208	142	0.189	2492	0.19	2638	0.95	0.952	99.8484	0.185
6.312-8.907	0.217	98	0.183	1948	0.185	2052	0.954	0.96	99.7076	0.181
8.907-94.642	0.311	61	0.227	1135	0.231	1202	0.871	0.939	99.5008	0.228

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