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

ID or keywords:

Entry

Database: PDB / ID: 7poc

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: 2.6 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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Download

PDBx/mmCIF format	7poc.cif.gz	595.2 KB	Display	PDBx/mmCIF format
PDB format	pdb7poc.ent.gz	481.8 KB	Display	PDB format
PDBx/mmJSON format	7poc.json.gz		Tree view	PDBx/mmJSON format
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Validation report

Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/po/7poc ftp://data.pdbj.org/pub/pdb/validation_reports/po/7poc	HTTPS FTP

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Related structure data

Related structure data	7poaC 7pobC 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)	57.040, 134.790, 184.020
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	19
Space group name H-M	P2₁2₁2₁

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:

Refine code: 1

Dom-ID	Component-ID	Ens-ID	Beg auth comp-ID	Beg label comp-ID	End auth comp-ID	End label comp-ID	Auth asym-ID	Label asym-ID	Auth seq-ID	Label seq-ID
1	1	1	ASP	ASP	ILE	ILE	A	A	5 - 394	30 - 419
2	1	1	ASP	ASP	ILE	ILE	B	B	5 - 394	30 - 419
3	2	2	ASP	ASP	ARG	ARG	A	A	5 - 396	30 - 421
4	2	2	ASP	ASP	ARG	ARG	C	C	5 - 396	30 - 421
5	3	3	ASP	ASP	ILE	ILE	A	A	5 - 394	30 - 419
6	3	3	ASP	ASP	ILE	ILE	D	D	5 - 394	30 - 419
7	4	4	ASP	ASP	ILE	ILE	B	B	5 - 394	30 - 419
8	4	4	ASP	ASP	ILE	ILE	C	C	5 - 394	30 - 419
9	5	5	SER	SER	ILE	ILE	B	B	3 - 395	28 - 420
10	5	5	SER	SER	ILE	ILE	D	D	3 - 395	28 - 420
11	6	6	ASP	ASP	ILE	ILE	C	C	5 - 394	30 - 419
12	6	6	ASP	ASP	ILE	ILE	D	D	5 - 394	30 - 419

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: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 49 / 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 grow	Temperature: 293.15 K / Method: vapor diffusion, sitting drop Details: 30 mM magnesium chloride, 30 mM calcium chloride, 100 mM Tris/Bicine pH 8.5, 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 PILATUS3 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: 2.6→54.483 Å / Num. obs: 44495 / % possible obs: 99.7 % / Redundancy: 21.8 % / CC_1/2: 0.999 / R_pim(I) all: 0.063 / R_rim(I) all: 0.215 / Net I/σ(I): 11.6

Reflection shell

Resolution (Å)	Redundancy (%)	Num. unique obs	CC_1/2	R_pim(I) all	R_rim(I) all	Diffraction-ID
9.73-54.48	24	976	0.999	0.013	0.049	1
2.6-2.7	21.6	4583	0.775	0.703	2.368	1

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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
xia2		data scaling
autoPROC		data reduction

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 3TQX

3tqx

Resolution: 2.6→54.483 Å / Cor.coef. F_o:F_c: 0.935 / Cor.coef. F_o:F_c free: 0.9 / WR_factor R_free: 0.279 / WR_factor R_work: 0.216 / SU B: 21.83 / SU ML: 0.437 / Average fsc free: 0.8069 / Average fsc work: 0.8427 / Cross valid method: FREE R-VALUE / ESU R Free: 0.444
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.3107	2229	5.017 %
R_work	0.2423	42198	-
all	0.246	-	-
obs	-	44427	99.655 %

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: 59.424 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-3.188 Å²	0 Å²	0 Å²
2-	-	-2.166 Å²	0 Å²
3-	-	-	5.353 Å²

Refinement step

Cycle: LAST / Resolution: 2.6→54.483 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	12178	0	60	49	12287

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.007	0.013	12482
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.015	12205
X-RAY DIFFRACTION	r_angle_refined_deg	1.575	1.641	16894
X-RAY DIFFRACTION	r_angle_other_deg	1.195	1.579	28048
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.651	5	1566
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	29.051	21.056	644
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	20.079	15	2148
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	21.628	15	102
X-RAY DIFFRACTION	r_chiral_restr	0.067	0.2	1566
X-RAY DIFFRACTION	r_gen_planes_refined	0.006	0.02	14012
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	2813
X-RAY DIFFRACTION	r_nbd_refined	0.217	0.2	2596
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.187	0.2	11516
X-RAY DIFFRACTION	r_nbtor_refined	0.16	0.2	5868
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.08	0.2	6447
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.164	0.2	292
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.033	0.2	2
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.224	0.2	34
X-RAY DIFFRACTION	r_nbd_other	0.314	0.2	90
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.242	0.2	3
X-RAY DIFFRACTION	r_mcbond_it	5.261	6.098	6276
X-RAY DIFFRACTION	r_mcbond_other	5.256	6.097	6275
X-RAY DIFFRACTION	r_mcangle_it	7.72	9.143	7838
X-RAY DIFFRACTION	r_mcangle_other	7.72	9.144	7839
X-RAY DIFFRACTION	r_scbond_it	4.877	6.518	6206
X-RAY DIFFRACTION	r_scbond_other	4.877	6.519	6203
X-RAY DIFFRACTION	r_scangle_it	7.425	9.59	9056
X-RAY DIFFRACTION	r_scangle_other	7.424	9.59	9057
X-RAY DIFFRACTION	r_lrange_it	10.282	71.326	13431
X-RAY DIFFRACTION	r_lrange_other	10.282	71.323	13431
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.076	0.05	11970
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.07	0.05	12284
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.087	0.05	11933
X-RAY DIFFRACTION	r_ncsr_local_group_4	0.084	0.05	11923
X-RAY DIFFRACTION	r_ncsr_local_group_5	0.079	0.05	12080
X-RAY DIFFRACTION	r_ncsr_local_group_6	0.086	0.05	11905

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	0.07632	0.05008
1	2	B	X-RAY DIFFRACTION	Local ncs	0.07632	0.05008
2	3	A	X-RAY DIFFRACTION	Local ncs	0.06996	0.05009
2	4	C	X-RAY DIFFRACTION	Local ncs	0.06996	0.05009
3	5	A	X-RAY DIFFRACTION	Local ncs	0.08678	0.05008
3	6	D	X-RAY DIFFRACTION	Local ncs	0.08678	0.05008
4	7	B	X-RAY DIFFRACTION	Local ncs	0.08379	0.05008
4	8	C	X-RAY DIFFRACTION	Local ncs	0.08379	0.05008
5	9	B	X-RAY DIFFRACTION	Local ncs	0.0792	0.05008
5	10	D	X-RAY DIFFRACTION	Local ncs	0.0792	0.05008
6	11	C	X-RAY DIFFRACTION	Local ncs	0.08621	0.05008
6	12	D	X-RAY DIFFRACTION	Local ncs	0.08621	0.05008

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
2.6-2.667	0.451	140	0.372	3060	0.375	3201	0.529	0.562	99.9688	0.348
2.667-2.74	0.396	177	0.327	3003	0.33	3180	0.664	0.712	100	0.303
2.74-2.82	0.369	162	0.313	2888	0.316	3056	0.706	0.754	99.8037	0.284
2.82-2.906	0.364	152	0.303	2835	0.306	2991	0.753	0.779	99.8663	0.273
2.906-3.001	0.398	155	0.294	2777	0.3	2934	0.75	0.788	99.9318	0.263
3.001-3.106	0.339	142	0.277	2631	0.28	2778	0.808	0.847	99.82	0.243
3.106-3.223	0.374	132	0.268	2575	0.274	2711	0.788	0.857	99.8525	0.236
3.223-3.354	0.31	126	0.233	2523	0.236	2652	0.837	0.899	99.8869	0.203
3.354-3.503	0.334	143	0.237	2339	0.243	2489	0.853	0.896	99.7188	0.206
3.503-3.673	0.289	119	0.217	2283	0.221	2413	0.884	0.915	99.5441	0.188
3.673-3.87	0.31	118	0.219	2166	0.223	2296	0.869	0.913	99.4773	0.191
3.87-4.104	0.276	119	0.204	2045	0.208	2184	0.911	0.931	99.0842	0.177
4.104-4.386	0.277	89	0.217	1964	0.22	2071	0.899	0.919	99.1309	0.194
4.386-4.734	0.272	87	0.211	1819	0.214	1926	0.92	0.937	98.9616	0.194
4.734-5.182	0.281	91	0.206	1682	0.209	1784	0.922	0.947	99.3834	0.193
5.182-5.787	0.265	71	0.229	1524	0.231	1603	0.908	0.926	99.5009	0.213
5.787-6.67	0.285	80	0.229	1361	0.232	1448	0.915	0.929	99.5166	0.214
6.67-8.139	0.324	55	0.24	1196	0.244	1254	0.906	0.926	99.7608	0.235
8.139-11.384	0.223	38	0.206	952	0.207	995	0.965	0.975	99.4975	0.214
11.384-54.483	0.271	33	0.284	574	0.283	608	0.957	0.953	99.8355	0.306

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