PDB-7pq9: Crystal structure of Bacillus clausii pdxR at 2.8 Angstroms resolution

Basic information

• Entry: Database: PDB / ID: 7pq9
• Title: Crystal structure of Bacillus clausii pdxR at 2.8 Angstroms resolution
• Components: PLP-dependent aminotransferase family protein
• Keywords: DNA BINDING PROTEIN / Bacillus clausii / MocR / PdxR / pyridoxal 5'-phosphate biosynthesis / transcriptional regulation / apo
• Function / homology: DI(HYDROXYETHYL)ETHER / TRIETHYLENE GLYCOL / :
• Biological species: Alkalihalobacillus clausii (bacteria)
• Method: X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.8 Å
• Authors: Vivoli Vega, M. / Isupov, M.N. / Harmer, N.

Funding support

United Kingdom, 1items

Organization	Grant number	Country
Defence Science and Technology Laboratory (DSTL)	DSTL/AGR/00210/01-R1000094053	United Kingdom

• Citation: Journal: Nucleic Acids Res / Year: 2023; Title: Structural insights into the DNA recognition mechanism by the bacterial transcription factor PdxR.; Authors: Ida Freda / Cécile Exertier / Anna Barile / Antonio Chaves-Sanjuan / Mirella Vivoli Vega / Michail N Isupov / Nicholas J Harmer / Elena Gugole / Paolo Swuec / Martino Bolognesi / Anita Scipioni / Carmelinda Savino / Martino Luigi Di Salvo / Roberto Contestabile / Beatrice Vallone / Angela Tramonti / Linda Celeste Montemiglio / ; Abstract: Specificity in protein-DNA recognition arises from the synergy of several factors that stem from the structural and chemical signatures encoded within the targeted DNA molecule. Here, we deciphered the nature of the interactions driving DNA recognition and binding by the bacterial transcription factor PdxR, a member of the MocR family responsible for the regulation of pyridoxal 5'-phosphate (PLP) biosynthesis. Single particle cryo-EM performed on the PLP-PdxR bound to its target DNA enabled the isolation of three conformers of the complex, which may be considered as snapshots of the binding process. Moreover, the resolution of an apo-PdxR crystallographic structure provided a detailed description of the transition of the effector domain to the holo-PdxR form triggered by the binding of the PLP effector molecule. Binding analyses of mutated DNA sequences using both wild type and PdxR variants revealed a central role of electrostatic interactions and of the intrinsic asymmetric bending of the DNA in allosterically guiding the holo-PdxR-DNA recognition process, from the first encounter through the fully bound state. Our results detail the structure and dynamics of the PdxR-DNA complex, clarifying the mechanism governing the DNA-binding mode of the holo-PdxR and the regulation features of the MocR family of transcription factors.

History

Deposition	Sep 16, 2021	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	Sep 28, 2022	Provider: repository / Type: Initial release
Revision 1.1	Oct 11, 2023	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond / citation / citation_author / struct_ncs_dom_lim Item: _citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _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 / _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	Jan 31, 2024	Group: Refinement description / Category: pdbx_initial_refinement_model

Assembly

Deposited unit

AAA: PLP-dependent aminotransferase family protein

BBB: PLP-dependent aminotransferase family protein

CCC: PLP-dependent aminotransferase family protein

DDD: PLP-dependent aminotransferase family protein

EEE: PLP-dependent aminotransferase family protein

FFF: PLP-dependent aminotransferase family protein

GGG: PLP-dependent aminotransferase family protein

HHH: PLP-dependent aminotransferase family protein

III: PLP-dependent aminotransferase family protein

JJJ: PLP-dependent aminotransferase family protein

KKK: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• 610 kDa, 11 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	609,690	43
Polymers	607,981	11
Non-polymers	1,709	32
Water	1,621	90

1

AAA: PLP-dependent aminotransferase family protein

BBB: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• Evidence: gel filtration, Monomers A and B, C and D, E and F, G and H form dimers in the asymmetric unit, monomers I and J form dimers with their crystallographic dyad counterparts. Monomer K is the only well defined N-terminal domain out of 10 present in the xtal, not clear which monomer out of 10 it belongs to.
• 111 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	110,773	8
Polymers	110,542	2
Non-polymers	231	6
Water	36	2

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Calculated values:

Buried area	5100 Å2
ΔGint	-81 kcal/mol
Surface area	29000 Å2
Method	PISA

2

CCC: PLP-dependent aminotransferase family protein

DDD: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• 111 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	110,782	8
Polymers	110,542	2
Non-polymers	240	6
Water	36	2

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Calculated values:

Buried area	5180 Å2
ΔGint	-95 kcal/mol
Surface area	29030 Å2
Method	PISA

3

EEE: PLP-dependent aminotransferase family protein

FFF: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• 111 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	110,812	6
Polymers	110,542	2
Non-polymers	270	4
Water	36	2

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Calculated values:

Buried area	5200 Å2
ΔGint	-65 kcal/mol
Surface area	29170 Å2
Method	PISA

4

GGG: PLP-dependent aminotransferase family protein

HHH: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• 111 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	111,233	11
Polymers	110,542	2
Non-polymers	691	9
Water	36	2

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Calculated values:

Buried area	5450 Å2
ΔGint	-87 kcal/mol
Surface area	28990 Å2
Method	PISA

5

III: PLP-dependent aminotransferase family protein

hetero molecules

III: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• 111 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	110,782	8
Polymers	110,542	2
Non-polymers	240	6
Water	36	2

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	8_675	x-y+1,-y+2,-z	1

Calculated values:

Buried area	5010 Å2
ΔGint	-99 kcal/mol
Surface area	29130 Å2
Method	PISA

6

JJJ: PLP-dependent aminotransferase family protein

hetero molecules

JJJ: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• 111 kDa, 2 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	110,773	8
Polymers	110,542	2
Non-polymers	231	6
Water	36	2

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	12_565	x,x-y+1,-z+1/6	1

Calculated values:

Buried area	5010 Å2
ΔGint	-76 kcal/mol
Surface area	28590 Å2
Method	PISA

7

KKK: PLP-dependent aminotransferase family protein

hetero molecules

Summary:

• defined by author&software
• 55.3 kDa, 1 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	55,311	2
Polymers	55,271	1
Non-polymers	40	1
Water	0

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Calculated values:

Buried area	90 Å2
ΔGint	-8 kcal/mol
Surface area	5340 Å2
Method	PISA

Unit cell

Length a, b, c (Å)	250.840, 250.840, 370.300
Angle α, β, γ (deg.)	90.000, 90.000, 120.000
Int Tables number	178
Space group name H-M	P6122

Noncrystallographic symmetry (NCS)

NCS domain:

ID	Ens-ID	Details
1	1	AAA
2	1	BBB
3	2	AAA
4	2	CCC
5	3	AAA
6	3	DDD
7	4	AAA
8	4	EEE
9	5	AAA
10	5	FFF
11	6	AAA
12	6	GGG
13	7	AAA
14	7	HHH
15	8	AAA
16	8	III
17	9	AAA
18	9	JJJ
19	10	BBB
20	10	CCC
21	11	BBB
22	11	DDD
23	12	BBB
24	12	EEE
25	13	BBB
26	13	FFF
27	14	BBB
28	14	GGG
29	15	BBB
30	15	HHH
31	16	BBB
32	16	III
33	17	BBB
34	17	JJJ
35	18	CCC
36	18	DDD
37	19	CCC
38	19	EEE
39	20	CCC
40	20	FFF
41	21	CCC
42	21	GGG
43	22	CCC
44	22	HHH
45	23	CCC
46	23	III
47	24	CCC
48	24	JJJ
49	25	DDD
50	25	EEE
51	26	DDD
52	26	FFF
53	27	DDD
54	27	GGG
55	28	DDD
56	28	HHH
57	29	DDD
58	29	III
59	30	DDD
60	30	JJJ
61	31	EEE
62	31	FFF
63	32	EEE
64	32	GGG
65	33	EEE
66	33	HHH
67	34	EEE
68	34	III
69	35	EEE
70	35	JJJ
71	36	FFF
72	36	GGG
73	37	FFF
74	37	HHH
75	38	FFF
76	38	III
77	39	FFF
78	39	JJJ
79	40	GGG
80	40	HHH
81	41	GGG
82	41	III
83	42	GGG
84	42	JJJ
85	43	HHH
86	43	III
87	44	HHH
88	44	JJJ
89	45	III
90	45	JJJ

NCS domain segments:

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	ARG	ARG	AAA	A	103 - 468	103 - 468
2	2	1	ASP	ASP	ARG	ARG	BBB	B	103 - 468	103 - 468
3	3	2	ASP	ASP	GLY	GLY	AAA	A	103 - 469	103 - 469
4	4	2	ASP	ASP	GLY	GLY	CCC	C	103 - 469	103 - 469
5	5	3	ASP	ASP	PRO	PRO	AAA	A	103 - 467	103 - 467
6	6	3	ASP	ASP	PRO	PRO	DDD	D	103 - 467	103 - 467
7	7	4	ASP	ASP	LEU	LEU	AAA	A	103 - 465	103 - 465
8	8	4	ASP	ASP	LEU	LEU	EEE	E	103 - 465	103 - 465
9	9	5	ASP	ASP	VAL	VAL	AAA	A	103 - 466	103 - 466
10	10	5	ASP	ASP	VAL	VAL	FFF	F	103 - 466	103 - 466
11	11	6	ASP	ASP	ARG	ARG	AAA	A	103 - 468	103 - 468
12	12	6	ASP	ASP	ARG	ARG	GGG	G	103 - 468	103 - 468
13	13	7	THR	THR	VAL	VAL	AAA	A	104 - 466	104 - 466
14	14	7	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
15	15	8	THR	THR	ARG	ARG	AAA	A	104 - 468	104 - 468
16	16	8	THR	THR	ARG	ARG	III	I	104 - 468	104 - 468
17	17	9	THR	THR	VAL	VAL	AAA	A	104 - 466	104 - 466
18	18	9	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466
19	19	10	ASP	ASP	ARG	ARG	BBB	B	103 - 468	103 - 468
20	20	10	ASP	ASP	ARG	ARG	CCC	C	103 - 468	103 - 468
21	21	11	ASP	ASP	PRO	PRO	BBB	B	103 - 467	103 - 467
22	22	11	ASP	ASP	PRO	PRO	DDD	D	103 - 467	103 - 467
23	23	12	ASP	ASP	LEU	LEU	BBB	B	103 - 465	103 - 465
24	24	12	ASP	ASP	LEU	LEU	EEE	E	103 - 465	103 - 465
25	25	13	ASP	ASP	VAL	VAL	BBB	B	103 - 466	103 - 466
26	26	13	ASP	ASP	VAL	VAL	FFF	F	103 - 466	103 - 466
27	27	14	ASP	ASP	SER	SER	BBB	B	103 - 470	103 - 470
28	28	14	ASP	ASP	SER	SER	GGG	G	103 - 470	103 - 470
29	29	15	THR	THR	VAL	VAL	BBB	B	104 - 466	104 - 466
30	30	15	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
31	31	16	THR	THR	GLY	GLY	BBB	B	104 - 469	104 - 469
32	32	16	THR	THR	GLY	GLY	III	I	104 - 469	104 - 469
33	33	17	THR	THR	VAL	VAL	BBB	B	104 - 466	104 - 466
34	34	17	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466
35	35	18	ASP	ASP	PRO	PRO	CCC	C	103 - 467	103 - 467
36	36	18	ASP	ASP	PRO	PRO	DDD	D	103 - 467	103 - 467
37	37	19	ASP	ASP	LEU	LEU	CCC	C	103 - 465	103 - 465
38	38	19	ASP	ASP	LEU	LEU	EEE	E	103 - 465	103 - 465
39	39	20	ASP	ASP	VAL	VAL	CCC	C	103 - 466	103 - 466
40	40	20	ASP	ASP	VAL	VAL	FFF	F	103 - 466	103 - 466
41	41	21	ASP	ASP	ARG	ARG	CCC	C	103 - 468	103 - 468
42	42	21	ASP	ASP	ARG	ARG	GGG	G	103 - 468	103 - 468
43	43	22	THR	THR	VAL	VAL	CCC	C	104 - 466	104 - 466
44	44	22	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
45	45	23	THR	THR	ARG	ARG	CCC	C	104 - 468	104 - 468
46	46	23	THR	THR	ARG	ARG	III	I	104 - 468	104 - 468
47	47	24	THR	THR	VAL	VAL	CCC	C	104 - 466	104 - 466
48	48	24	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466
49	49	25	ASP	ASP	LEU	LEU	DDD	D	103 - 465	103 - 465
50	50	25	ASP	ASP	LEU	LEU	EEE	E	103 - 465	103 - 465
51	51	26	ASP	ASP	VAL	VAL	DDD	D	103 - 466	103 - 466
52	52	26	ASP	ASP	VAL	VAL	FFF	F	103 - 466	103 - 466
53	53	27	ASP	ASP	PRO	PRO	DDD	D	103 - 467	103 - 467
54	54	27	ASP	ASP	PRO	PRO	GGG	G	103 - 467	103 - 467
55	55	28	THR	THR	VAL	VAL	DDD	D	104 - 466	104 - 466
56	56	28	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
57	57	29	THR	THR	PRO	PRO	DDD	D	104 - 467	104 - 467
58	58	29	THR	THR	PRO	PRO	III	I	104 - 467	104 - 467
59	59	30	THR	THR	VAL	VAL	DDD	D	104 - 466	104 - 466
60	60	30	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466
61	61	31	ASP	ASP	LEU	LEU	EEE	E	103 - 465	103 - 465
62	62	31	ASP	ASP	LEU	LEU	FFF	F	103 - 465	103 - 465
63	63	32	ASP	ASP	LEU	LEU	EEE	E	103 - 465	103 - 465
64	64	32	ASP	ASP	LEU	LEU	GGG	G	103 - 465	103 - 465
65	65	33	THR	THR	LEU	LEU	EEE	E	104 - 465	104 - 465
66	66	33	THR	THR	LEU	LEU	HHH	H	104 - 465	104 - 465
67	67	34	THR	THR	LEU	LEU	EEE	E	104 - 465	104 - 465
68	68	34	THR	THR	LEU	LEU	III	I	104 - 465	104 - 465
69	69	35	THR	THR	LEU	LEU	EEE	E	104 - 465	104 - 465
70	70	35	THR	THR	LEU	LEU	JJJ	J	104 - 465	104 - 465
71	71	36	ASP	ASP	VAL	VAL	FFF	F	103 - 466	103 - 466
72	72	36	ASP	ASP	VAL	VAL	GGG	G	103 - 466	103 - 466
73	73	37	THR	THR	VAL	VAL	FFF	F	104 - 466	104 - 466
74	74	37	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
75	75	38	THR	THR	VAL	VAL	FFF	F	104 - 466	104 - 466
76	76	38	THR	THR	VAL	VAL	III	I	104 - 466	104 - 466
77	77	39	THR	THR	VAL	VAL	FFF	F	104 - 466	104 - 466
78	78	39	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466
79	79	40	THR	THR	VAL	VAL	GGG	G	104 - 466	104 - 466
80	80	40	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
81	81	41	THR	THR	GLY	GLY	GGG	G	104 - 469	104 - 469
82	82	41	THR	THR	GLY	GLY	III	I	104 - 469	104 - 469
83	83	42	THR	THR	VAL	VAL	GGG	G	104 - 466	104 - 466
84	84	42	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466
85	85	43	THR	THR	VAL	VAL	HHH	H	104 - 466	104 - 466
86	86	43	THR	THR	VAL	VAL	III	I	104 - 466	104 - 466
87	87	44	THR	THR	PRO	PRO	HHH	H	104 - 467	104 - 467
88	88	44	THR	THR	PRO	PRO	JJJ	J	104 - 467	104 - 467
89	89	45	THR	THR	VAL	VAL	III	I	104 - 466	104 - 466
90	90	45	THR	THR	VAL	VAL	JJJ	J	104 - 466	104 - 466

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
7	Local NCS retraints between domains: 13 14
8	Local NCS retraints between domains: 15 16
9	Local NCS retraints between domains: 17 18
10	Local NCS retraints between domains: 19 20
11	Local NCS retraints between domains: 21 22
12	Local NCS retraints between domains: 23 24
13	Local NCS retraints between domains: 25 26
14	Local NCS retraints between domains: 27 28
15	Local NCS retraints between domains: 29 30
16	Local NCS retraints between domains: 31 32
17	Local NCS retraints between domains: 33 34
18	Local NCS retraints between domains: 35 36
19	Local NCS retraints between domains: 37 38
20	Local NCS retraints between domains: 39 40
21	Local NCS retraints between domains: 41 42
22	Local NCS retraints between domains: 43 44
23	Local NCS retraints between domains: 45 46
24	Local NCS retraints between domains: 47 48
25	Local NCS retraints between domains: 49 50
26	Local NCS retraints between domains: 51 52
27	Local NCS retraints between domains: 53 54
28	Local NCS retraints between domains: 55 56
29	Local NCS retraints between domains: 57 58
30	Local NCS retraints between domains: 59 60
31	Local NCS retraints between domains: 61 62
32	Local NCS retraints between domains: 63 64
33	Local NCS retraints between domains: 65 66
34	Local NCS retraints between domains: 67 68
35	Local NCS retraints between domains: 69 70
36	Local NCS retraints between domains: 71 72
37	Local NCS retraints between domains: 73 74
38	Local NCS retraints between domains: 75 76
39	Local NCS retraints between domains: 77 78
40	Local NCS retraints between domains: 79 80
41	Local NCS retraints between domains: 81 82
42	Local NCS retraints between domains: 83 84
43	Local NCS retraints between domains: 85 86
44	Local NCS retraints between domains: 87 88
45	Local NCS retraints between domains: 89 90

Components

Protein , 1 types, 11 molecules AAA BBB CCC DDD EEE FFF GGG HHH III JJJ KKK

#1: Protein

AAA BBB CCC DDD EEE FFF GGG HHH III JJJ KKK
PLP-dependent aminotransferase family protein

Details:

Mass: 55271.004 Da / Num. of mol.: 11
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Alkalihalobacillus clausii (bacteria) / Gene: HJ578_10545 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A7Y3RUA1

Non-polymers , 7 types, 122 molecules

#2: Chemical

AAA-501 AAA-502 BBB-501 BBB-502 CCC-501 CCC-502 CCC-503 DDD-501 DDD-502 DDD-503 EEE-502 EEE-503 FFF-501 GGG-503 GGG-504 GGG-505 HHH-501 HHH-502 HHH-503 III-501 ...
ChemComp-CA / CALCIUM ION

Details:

Mass: 40.078 Da / Num. of mol.: 25 / Source method: obtained synthetically / Formula: Ca

#3: Chemical

AAA-503 AAA-504 JJJ-503 ChemComp-CL / CHLORIDE ION

Details:

Mass: 35.453 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Cl

#4: Chemical

EEE-501 ChemComp-PGE / TRIETHYLENE GLYCOL

Details:

Mass: 150.173 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₆H₁₄O₄

#5: Chemical

GGG-501 ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL

Details:

Mass: 62.068 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₂H₆O₂

#6: Chemical

GGG-502 ChemComp-PEG / DI(HYDROXYETHYL)ETHER

Details:

Mass: 106.120 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₄H₁₀O₃

#7: Chemical

HHH-504 ChemComp-P6G / HEXAETHYLENE GLYCOL / POLYETHYLENE GLYCOL PEG400

Details:

Mass: 282.331 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₁₂H₂₆O₇ / Comment: precipitant*YM

#8: Water

ChemComp-HOH / water

Details:

Mass: 18.015 Da / Num. of mol.: 90 / Source method: isolated from a natural source / Formula: H₂O

Details

• Has ligand of interest: N

Experimental details

Experiment

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

Sample preparation

• Crystal: Density Matthews: 2.81 ų/Da / Density % sol: 56.24 %
• Crystal grow: Temperature: 291 K / Method: microbatch; Details: 33% (vol/vol) PEG400, 200 mM calcium acetate, and 200 mM imidazole, pH7.5.

Data collection

• Diffraction: Mean temperature: 100 K / Serial crystal experiment: N
• Diffraction source: Source: SYNCHROTRON / Site: Diamond / Beamline: I03 / Wavelength: 0.97625 Å
• Detector: Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Sep 27, 2015
• Radiation: Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 0.97625 Å / Relative weight: 1
• Reflection: Resolution: 2.8→125.42 Å / Num. obs: 164628 / % possible obs: 98.6 % / Redundancy: 25.1 % / CC_1/2: 0.999 / Net I/σ(I): 15.5
• Reflection shell: Resolution: 2.8→2.87 Å / Num. unique obs: 12078 / CC_1/2: 0.279

Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
XDS		data reduction
XSCALE		data scaling
MoRDa		phasing
PARROT		phasing
DM		phasing
Coot		model building

Refinement

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

4mgr

Resolution: 2.8→125.735 Å / Cor.coef. F_o:F_c: 0.954 / Cor.coef. F_o:F_c free: 0.938 / WR_factor R_free: 0.234 / WR_factor R_work: 0.189 / SU B: 17.562 / SU ML: 0.314 / Average fsc free: 0.8193 / Average fsc work: 0.8315 / Cross valid method: FREE R-VALUE / ESU R: 0.576 / ESU R Free: 0.322 / Details: Hydrogens have not been used

	Rfactor	Num. reflection	% reflection
Rfree	0.2537	8331	5.061 %
Rwork	0.2094	156297	-
all	0.212	-	-
obs	-	164628	98.185 %

• Solvent computation: Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: BABINET MODEL PLUS MASK

Displacement parameters

B_iso mean: 111.655 Ų

	Baniso -1	Baniso -2	Baniso -3
1-	0.324 Å2	0.162 Å2	0 Å2
2-	-	0.324 Å2	-0 Å2
3-	-	-	-1.052 Å2

Refinement step

Cycle: LAST / Resolution: 2.8→125.735 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	30571	0	68	90	30729

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.008	0.012	31374
X-RAY DIFFRACTION	r_angle_refined_deg	1.706	1.637	42471
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	7.525	5	3730
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	33.16	21.456	1744
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	17.398	15	5498
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	25.421	15	230
X-RAY DIFFRACTION	r_chiral_restr	0.128	0.2	3930
X-RAY DIFFRACTION	r_gen_planes_refined	0.009	0.02	23767
X-RAY DIFFRACTION	r_nbd_refined	0.227	0.2	13432
X-RAY DIFFRACTION	r_nbtor_refined	0.32	0.2	21065
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.151	0.2	839
X-RAY DIFFRACTION	r_metal_ion_refined	0.158	0.2	6
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.298	0.2	190
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.242	0.2	6
X-RAY DIFFRACTION	r_mcbond_it	13.007	10.798	14957
X-RAY DIFFRACTION	r_mcangle_it	17.951	16.171	18672
X-RAY DIFFRACTION	r_scbond_it	14.601	11.314	16410
X-RAY DIFFRACTION	r_scangle_it	19.728	16.692	23796
X-RAY DIFFRACTION	r_lrange_it	25.167	203.465	128219
X-RAY DIFFRACTION	r_ncsr_local_group_1	0.094	0.05	12026
X-RAY DIFFRACTION	r_ncsr_local_group_2	0.097	0.05	12037
X-RAY DIFFRACTION	r_ncsr_local_group_3	0.084	0.05	12071
X-RAY DIFFRACTION	r_ncsr_local_group_4	0.083	0.05	12046
X-RAY DIFFRACTION	r_ncsr_local_group_5	0.082	0.05	12002
X-RAY DIFFRACTION	r_ncsr_local_group_6	0.093	0.05	12011
X-RAY DIFFRACTION	r_ncsr_local_group_7	0.088	0.05	11945
X-RAY DIFFRACTION	r_ncsr_local_group_8	0.09	0.05	11943
X-RAY DIFFRACTION	r_ncsr_local_group_9	0.08	0.05	12022
X-RAY DIFFRACTION	r_ncsr_local_group_10	0.094	0.05	11993
X-RAY DIFFRACTION	r_ncsr_local_group_11	0.086	0.05	12025
X-RAY DIFFRACTION	r_ncsr_local_group_12	0.091	0.05	11957
X-RAY DIFFRACTION	r_ncsr_local_group_13	0.077	0.05	12041
X-RAY DIFFRACTION	r_ncsr_local_group_14	0.09	0.05	12003
X-RAY DIFFRACTION	r_ncsr_local_group_15	0.088	0.05	11914
X-RAY DIFFRACTION	r_ncsr_local_group_16	0.083	0.05	12067
X-RAY DIFFRACTION	r_ncsr_local_group_17	0.077	0.05	12022
X-RAY DIFFRACTION	r_ncsr_local_group_18	0.09	0.05	12067
X-RAY DIFFRACTION	r_ncsr_local_group_19	0.086	0.05	12018
X-RAY DIFFRACTION	r_ncsr_local_group_20	0.086	0.05	12022
X-RAY DIFFRACTION	r_ncsr_local_group_21	0.084	0.05	12195
X-RAY DIFFRACTION	r_ncsr_local_group_22	0.093	0.05	11946
X-RAY DIFFRACTION	r_ncsr_local_group_23	0.093	0.05	11942
X-RAY DIFFRACTION	r_ncsr_local_group_24	0.087	0.05	12005
X-RAY DIFFRACTION	r_ncsr_local_group_25	0.072	0.05	12149
X-RAY DIFFRACTION	r_ncsr_local_group_26	0.079	0.05	12096
X-RAY DIFFRACTION	r_ncsr_local_group_27	0.082	0.05	12103
X-RAY DIFFRACTION	r_ncsr_local_group_28	0.081	0.05	12025
X-RAY DIFFRACTION	r_ncsr_local_group_29	0.083	0.05	11979
X-RAY DIFFRACTION	r_ncsr_local_group_30	0.076	0.05	12044
X-RAY DIFFRACTION	r_ncsr_local_group_31	0.079	0.05	12084
X-RAY DIFFRACTION	r_ncsr_local_group_32	0.086	0.05	12003
X-RAY DIFFRACTION	r_ncsr_local_group_33	0.08	0.05	11999
X-RAY DIFFRACTION	r_ncsr_local_group_34	0.076	0.05	12024
X-RAY DIFFRACTION	r_ncsr_local_group_35	0.073	0.05	12058
X-RAY DIFFRACTION	r_ncsr_local_group_36	0.081	0.05	12004
X-RAY DIFFRACTION	r_ncsr_local_group_37	0.081	0.05	12004
X-RAY DIFFRACTION	r_ncsr_local_group_38	0.077	0.05	12026
X-RAY DIFFRACTION	r_ncsr_local_group_39	0.069	0.05	12097
X-RAY DIFFRACTION	r_ncsr_local_group_40	0.084	0.05	12002
X-RAY DIFFRACTION	r_ncsr_local_group_41	0.091	0.05	11932
X-RAY DIFFRACTION	r_ncsr_local_group_42	0.078	0.05	11989
X-RAY DIFFRACTION	r_ncsr_local_group_43	0.085	0.05	11996
X-RAY DIFFRACTION	r_ncsr_local_group_44	0.076	0.05	12072
X-RAY DIFFRACTION	r_ncsr_local_group_45	0.074	0.05	12069

Refine LS restraints NCS

Ens-ID	Dom-ID	Auth asym-ID	Refine-ID	Type	Rms dev position (Å)	Weight position
1	1	AAA	X-RAY DIFFRACTION	Local ncs	0.09418	0.05009
1	2	BBB	X-RAY DIFFRACTION	Local ncs	0.09418	0.05009
2	3	AAA	X-RAY DIFFRACTION	Local ncs	0.09672	0.05009
2	4	CCC	X-RAY DIFFRACTION	Local ncs	0.09672	0.05009
3	5	AAA	X-RAY DIFFRACTION	Local ncs	0.0838	0.0501
3	6	DDD	X-RAY DIFFRACTION	Local ncs	0.0838	0.0501
4	7	AAA	X-RAY DIFFRACTION	Local ncs	0.08312	0.0501
4	8	EEE	X-RAY DIFFRACTION	Local ncs	0.08312	0.0501
5	9	AAA	X-RAY DIFFRACTION	Local ncs	0.08243	0.05009
5	10	FFF	X-RAY DIFFRACTION	Local ncs	0.08243	0.05009
6	11	AAA	X-RAY DIFFRACTION	Local ncs	0.09288	0.05009
6	12	GGG	X-RAY DIFFRACTION	Local ncs	0.09288	0.05009
7	13	AAA	X-RAY DIFFRACTION	Local ncs	0.08831	0.05009
7	14	HHH	X-RAY DIFFRACTION	Local ncs	0.08831	0.05009
8	15	AAA	X-RAY DIFFRACTION	Local ncs	0.0898	0.05009
8	16	III	X-RAY DIFFRACTION	Local ncs	0.0898	0.05009
9	17	AAA	X-RAY DIFFRACTION	Local ncs	0.07975	0.05009
9	18	JJJ	X-RAY DIFFRACTION	Local ncs	0.07975	0.05009
10	19	BBB	X-RAY DIFFRACTION	Local ncs	0.09416	0.05009
10	20	CCC	X-RAY DIFFRACTION	Local ncs	0.09416	0.05009
11	21	BBB	X-RAY DIFFRACTION	Local ncs	0.08578	0.05009
11	22	DDD	X-RAY DIFFRACTION	Local ncs	0.08578	0.05009
12	23	BBB	X-RAY DIFFRACTION	Local ncs	0.09074	0.05009
12	24	EEE	X-RAY DIFFRACTION	Local ncs	0.09074	0.05009
13	25	BBB	X-RAY DIFFRACTION	Local ncs	0.07733	0.05009
13	26	FFF	X-RAY DIFFRACTION	Local ncs	0.07733	0.05009
14	27	BBB	X-RAY DIFFRACTION	Local ncs	0.08977	0.05009
14	28	GGG	X-RAY DIFFRACTION	Local ncs	0.08977	0.05009
15	29	BBB	X-RAY DIFFRACTION	Local ncs	0.08847	0.05009
15	30	HHH	X-RAY DIFFRACTION	Local ncs	0.08847	0.05009
16	31	BBB	X-RAY DIFFRACTION	Local ncs	0.08297	0.0501
16	32	III	X-RAY DIFFRACTION	Local ncs	0.08297	0.0501
17	33	BBB	X-RAY DIFFRACTION	Local ncs	0.0771	0.0501
17	34	JJJ	X-RAY DIFFRACTION	Local ncs	0.0771	0.0501
18	35	CCC	X-RAY DIFFRACTION	Local ncs	0.08973	0.05009
18	36	DDD	X-RAY DIFFRACTION	Local ncs	0.08973	0.05009
19	37	CCC	X-RAY DIFFRACTION	Local ncs	0.0858	0.05009
19	38	EEE	X-RAY DIFFRACTION	Local ncs	0.0858	0.05009
20	39	CCC	X-RAY DIFFRACTION	Local ncs	0.08647	0.05009
20	40	FFF	X-RAY DIFFRACTION	Local ncs	0.08647	0.05009
21	41	CCC	X-RAY DIFFRACTION	Local ncs	0.08417	0.0501
21	42	GGG	X-RAY DIFFRACTION	Local ncs	0.08417	0.0501
22	43	CCC	X-RAY DIFFRACTION	Local ncs	0.09276	0.05009
22	44	HHH	X-RAY DIFFRACTION	Local ncs	0.09276	0.05009
23	45	CCC	X-RAY DIFFRACTION	Local ncs	0.09349	0.05009
23	46	III	X-RAY DIFFRACTION	Local ncs	0.09349	0.05009
24	47	CCC	X-RAY DIFFRACTION	Local ncs	0.08662	0.05009
24	48	JJJ	X-RAY DIFFRACTION	Local ncs	0.08662	0.05009
25	49	DDD	X-RAY DIFFRACTION	Local ncs	0.07205	0.0501
25	50	EEE	X-RAY DIFFRACTION	Local ncs	0.07205	0.0501
26	51	DDD	X-RAY DIFFRACTION	Local ncs	0.07939	0.0501
26	52	FFF	X-RAY DIFFRACTION	Local ncs	0.07939	0.0501
27	53	DDD	X-RAY DIFFRACTION	Local ncs	0.08191	0.05009
27	54	GGG	X-RAY DIFFRACTION	Local ncs	0.08191	0.05009
28	55	DDD	X-RAY DIFFRACTION	Local ncs	0.08082	0.05009
28	56	HHH	X-RAY DIFFRACTION	Local ncs	0.08082	0.05009
29	57	DDD	X-RAY DIFFRACTION	Local ncs	0.08312	0.05009
29	58	III	X-RAY DIFFRACTION	Local ncs	0.08312	0.05009
30	59	DDD	X-RAY DIFFRACTION	Local ncs	0.07647	0.0501
30	60	JJJ	X-RAY DIFFRACTION	Local ncs	0.07647	0.0501
31	61	EEE	X-RAY DIFFRACTION	Local ncs	0.07878	0.0501
31	62	FFF	X-RAY DIFFRACTION	Local ncs	0.07878	0.0501
32	63	EEE	X-RAY DIFFRACTION	Local ncs	0.08631	0.0501
32	64	GGG	X-RAY DIFFRACTION	Local ncs	0.08631	0.0501
33	65	EEE	X-RAY DIFFRACTION	Local ncs	0.07963	0.0501
33	66	HHH	X-RAY DIFFRACTION	Local ncs	0.07963	0.0501
34	67	EEE	X-RAY DIFFRACTION	Local ncs	0.07603	0.0501
34	68	III	X-RAY DIFFRACTION	Local ncs	0.07603	0.0501
35	69	EEE	X-RAY DIFFRACTION	Local ncs	0.07306	0.0501
35	70	JJJ	X-RAY DIFFRACTION	Local ncs	0.07306	0.0501
36	71	FFF	X-RAY DIFFRACTION	Local ncs	0.08101	0.05009
36	72	GGG	X-RAY DIFFRACTION	Local ncs	0.08101	0.05009
37	73	FFF	X-RAY DIFFRACTION	Local ncs	0.08059	0.05009
37	74	HHH	X-RAY DIFFRACTION	Local ncs	0.08059	0.05009
38	75	FFF	X-RAY DIFFRACTION	Local ncs	0.07671	0.0501
38	76	III	X-RAY DIFFRACTION	Local ncs	0.07671	0.0501
39	77	FFF	X-RAY DIFFRACTION	Local ncs	0.06876	0.0501
39	78	JJJ	X-RAY DIFFRACTION	Local ncs	0.06876	0.0501
40	79	GGG	X-RAY DIFFRACTION	Local ncs	0.08382	0.05009
40	80	HHH	X-RAY DIFFRACTION	Local ncs	0.08382	0.05009
41	81	GGG	X-RAY DIFFRACTION	Local ncs	0.09123	0.05009
41	82	III	X-RAY DIFFRACTION	Local ncs	0.09123	0.05009
42	83	GGG	X-RAY DIFFRACTION	Local ncs	0.07805	0.05009
42	84	JJJ	X-RAY DIFFRACTION	Local ncs	0.07805	0.05009
43	85	HHH	X-RAY DIFFRACTION	Local ncs	0.08473	0.05009
43	86	III	X-RAY DIFFRACTION	Local ncs	0.08473	0.05009
44	87	HHH	X-RAY DIFFRACTION	Local ncs	0.07551	0.05009
44	88	JJJ	X-RAY DIFFRACTION	Local ncs	0.07551	0.05009
45	89	III	X-RAY DIFFRACTION	Local ncs	0.07359	0.0501
45	90	JJJ	X-RAY DIFFRACTION	Local ncs	0.07359	0.0501

LS refinement shell

Resolution (Å)	Rfactor Rfree	Num. reflection Rfree	Rfactor Rwork	Num. reflection Rwork	Refine-ID	% reflection obs (%)
2.8-2.873	0.412	643	0.386	11472	X-RAY DIFFRACTION	98.8737
2.873-2.951	0.379	591	0.387	11230	X-RAY DIFFRACTION	99.0033
2.951-3.037	0.402	567	0.356	10905	X-RAY DIFFRACTION	98.9221
3.037-3.13	0.372	529	0.316	10646	X-RAY DIFFRACTION	98.8851
3.13-3.233	0.346	520	0.296	10295	X-RAY DIFFRACTION	98.8122
3.233-3.347	0.283	518	0.255	9944	X-RAY DIFFRACTION	98.6888
3.347-3.473	0.294	530	0.245	9570	X-RAY DIFFRACTION	98.6039
3.473-3.615	0.275	487	0.218	9245	X-RAY DIFFRACTION	98.5419
3.615-3.775	0.26	473	0.207	8841	X-RAY DIFFRACTION	98.3215
3.775-3.959	0.25	462	0.199	8469	X-RAY DIFFRACTION	98.2508
3.959-4.174	0.22	395	0.182	8087	X-RAY DIFFRACTION	97.8655
4.174-4.427	0.219	430	0.173	7606	X-RAY DIFFRACTION	98.0239
4.427-4.732	0.242	394	0.175	7167	X-RAY DIFFRACTION	97.8517
4.732-5.111	0.22	380	0.171	6667	X-RAY DIFFRACTION	97.4958
5.111-5.599	0.256	326	0.186	6159	X-RAY DIFFRACTION	97.3578
5.599-6.259	0.26	300	0.179	5596	X-RAY DIFFRACTION	97.1495
6.259-7.226	0.233	262	0.191	4940	X-RAY DIFFRACTION	96.4584
7.226-8.848	0.206	234	0.167	4196	X-RAY DIFFRACTION	96.3043
8.848-12.502	0.18	189	0.168	3315	X-RAY DIFFRACTION	96.0526
12.502-125.735	0.294	101	0.278	1948	X-RAY DIFFRACTION	94.7734