BMRB: 25888

1H, 13C and 15N chemical shift assignments and solution structure for PARP-1 F1F2 domains in complex with a DNA single-strand break

Authors

Neuhaus, D., Eustermann, S., Yang, J., Wu, W.

Assembly

F1F2-DNA complex

Entity

1. F1F2 (polymer, Thiol state: all other bound), 214 monomers, 24062.18 Da Detail

MAESSDKLYR VEYAKSGRAS CKKCSESIPK DSLRMAIMVQ SPMFDGKVPH WYHFSCFWKV GHSIRHPDVE VDGFSELRWD DQQKVKKTAE AGGVTGKGQD GIGSKAEKTL GDFAAEYAKS NRSTCKGCME KIEKGQVRLS KKMVDPEKPQ LGMIDRWYHP GCFVKNREEL GFRPEYSASQ LKGFSLLATE DKEALKKQLP GVKSEGKRKG DEVD

2. DNA (45-MER) (polymer, Thiol state: not present), 45 monomers, 13934.83 Da Detail

GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCG

3. F1F2-DNA complex, entity ZN (non-polymer), 65.409 × 2 Da

Total weight

38127.824 Da

Max. entity weight

24062.18 Da

Entity Connection

na 8 Detail

---

ID	Type	Value order	Atom ID 1	Atom ID 2
1	na	sing	1:CYS21:SG	3:ZN1:ZN
2	na	sing	1:CYS24:SG	3:ZN1:ZN
3	na	sing	1:CYS56:SG	3:ZN1:ZN
4	na	sing	1:HIS53:ND1	3:ZN1:ZN
5	na	sing	1:CYS125:SG	3:ZN1:ZN
6	na	sing	1:CYS128:SG	3:ZN1:ZN
7	na	sing	1:CYS162:SG	3:ZN1:ZN
8	na	sing	1:HIS159:ND1	3:ZN1:ZN

Source organism

Homo sapiens

Exptl. method

solution NMR

Refine. method

simulated annealing

Data set

assigned_chemical_shifts

Chem. Shift Complete1

Sequence coverage: 82.2 %, Completeness: 34.5 %, Completeness (bb): 46.9 % Detail

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Polymer type: polypeptide(L) polydeoxyribonucleotide

	Total	1H	13C	15N
All	34.5 % (1020 of 2957)	23.9 % (423 of 1773)	41.0 % (396 of 965)	91.8 % (201 of 219)
Backbone	46.9 % (741 of 1581)	32.2 % (243 of 754)	47.7 % (297 of 622)	98.0 % (201 of 205)
Sidechain	20.3 % (318 of 1570)	17.7 % (180 of 1019)	25.7 % (138 of 537)	0.0 % (0 of 14)
Aromatic	8.4 % (24 of 286)	7.4 % (14 of 188)	10.6 % (10 of 94)	0.0 % (0 of 4)
Methyl	76.3 % (129 of 169)	71.9 % (64 of 89)	81.3 % (65 of 80)

1. F1F2

MAESSDKLYR VEYAKSGRAS CKKCSESIPK DSLRMAIMVQ SPMFDGKVPH WYHFSCFWKV GHSIRHPDVE VDGFSELRWD DQQKVKKTAE AGGVTGKGQD GIGSKAEKTL GDFAAEYAKS NRSTCKGCME KIEKGQVRLS KKMVDPEKPQ LGMIDRWYHP GCFVKNREEL GFRPEYSASQ LKGFSLLATE DKEALKKQLP GVKSEGKRKG DEVD

2. DNA (45-MER)

GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCG

Show sample condition

Sample #1

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
1	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
2	DNA (45-MER)	natural abundance		0.2 mM
3	TRIS	[U-2H]		50 mM
4	DTT	[U-2H]		1 mM
5	ZnSO4	natural abundance		0.1 mM
6	H2O	natural abundance	solvent	95 %
7	D2O	[U-2H]	solvent	5 %

Sample #2

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H (approx. 70%),15N,13C]. This sample was only used to confirm assignments by checking patterns of peaks; no numerical shift data from experiments with this sample contributed to the deposition (since isotope shifts could differ).

#	Name	Isotope labeling	Type	Concentration
8	PARP-1 1-214	[U-15N; U-13C; U-70% 2H]		0.2 mM
9	DNA (45-MER)	natural abundance		0.2 mM
10	TRIS	[U-2H]		50 mM
11	DTT	[U-2H]		1 mM
12	ZnSO4	natural abundance		0.1 mM
13	H2O	natural abundance	solvent	95 %
14	D2O	[U-2H]	solvent	5 %

Sample #3

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N]

#	Name	Isotope labeling	Type	Concentration
15	PARP-1 1-214	[U-98% 2H; U-98% 15N]		0.2 mM
16	DNA (45-MER)	natural abundance		0.2 mM
17	TRIS	[U-2H]		50 mM
18	DTT	[U-2H]		1 mM
19	ZnSO4	natural abundance		0.1 mM
20	H2O	natural abundance	solvent	95 %
21	D2O	[U-2H]	solvent	5 %

Sample #4

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the delta-methyl groups of Ile and all methyl groups of Leu and Val residues, using sodium [4-13C, 3,3-2H2] alpha-ketobutyrate and sodium [3- 2H, 4,4'-13C2] alpha-ketoisovalerate as precursors to maximize protonation of methyl groups, for use in NOE experiments; sodium [3-2H, 4,4'-13C2] alpha-ketoisovalerate was prepared from sodium [4,4'-13C2] alpha-ketoisovalerate by exchange with 2H2O at pH 12.5 and 45 C for 3 hrs.

#	Name	Isotope labeling	Type	Concentration
22	PARP-1 1-214	see Sample details section		0.2 mM
23	DNA (45-MER)	natural abundance		0.2 mM
24	TRIS	[U-2H]		50 mM
25	DTT	[U-2H]		1 mM
26	ZnSO4	natural abundance		0.1 mM
27	H2O	natural abundance	solvent	95 %
28	D2O	[U-2H]	solvent	5 %

Sample #5

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the delta-methyl groups of Ile and all methyl groups of Leu and Val residues, using sodium [3,3-2H2,13C4] alpha-ketobutyrate and sodium [3- 2H,13C5] alpha-ketoisovalerate as precursors to produce linear chains of 13C in the sidechains of Val and Leu, for use in assignment experiments to link methyl signals to C-alpha signals.

#	Name	Isotope labeling	Type	Concentration
29	PARP-1 1-214	see Sample details section		0.2 mM
30	DNA (45-MER)	natural abundance		0.2 mM
31	TRIS	[U-2H]		50 mM
32	DTT	[U-2H]		1 mM
33	ZnSO4	natural abundance		0.1 mM
34	D2O	[U-2H]	solvent	100 %

Sample #6

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the methyl groups of Met residues in addition to Ile, Leu and Val methyl groups as in sample 4a.

#	Name	Isotope labeling	Type	Concentration
35	PARP-1 1-214	see Sample details section		0.2 mM
36	DNA (45-MER)	natural abundance		0.2 mM
37	TRIS	[U-2H]		50 mM
38	DTT	[U-2H]		1 mM
39	ZnSO4	natural abundance		0.1 mM
40	D2O	[U-2H]	solvent	100 %

Sample #7

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]; back-labeled with [1H,13C] in the methyl groups of Ile, Leu and Val methyl groups as in sample 4a and [1H,13C,15N] Phe residues.

#	Name	Isotope labeling	Type	Concentration
41	PARP-1 1-214	see Sample details section		0.2 mM
42	DNA (45-MER)	natural abundance		0.2 mM
43	TRIS	[U-2H]		50 mM
44	DTT	[U-2H]		1 mM
45	ZnSO4	natural abundance		0.1 mM
46	D2O	[U-2H]	solvent	100 %

Sample #8

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): uniform [1H,12C,15N] Labelling for residues 105-214 (and GGG of insertion): [2H,15N,13C] back-labeled with [1H,13C] Ile, Leu Val and Met methyl groups labeled as in sample 5

#	Name	Isotope labeling	Type	Concentration
47	PARP-1 1-214	see Sample details section		0.2 mM
48	DNA (45-MER)	natural abundance		0.2 mM
49	TRIS	[U-2H]		50 mM
50	DTT	[U-2H]		1 mM
51	ZnSO4	natural abundance		0.1 mM
52	D2O	[U-2H]	solvent	100 %

Sample #9

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): uniform [1H,12C,15N] Labelling for residues 105-214 (and GGG of insertion): [2H,15N,13C] back labeled with [1H,13C] Met methyl groups as in sample 5 and [13C,15N,1H] Arg residues

#	Name	Isotope labeling	Type	Concentration
53	PARP-1 1-214	see Sample details section		0.2 mM
54	DNA (45-MER)	natural abundance		0.2 mM
55	TRIS	[U-2H]		50 mM
56	DTT	[U-2H]		1 mM
57	ZnSO4	natural abundance		0.1 mM
58	D2O	[U-2H]	solvent	100 %

Sample #10

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a and [1H,13C,15N] Arg residues Labelling for residues 105-214 (and GGG of insertion): uniform [1H,12C,15N]

#	Name	Isotope labeling	Type	Concentration
59	PARP-1 1-214	see Sample details section		0.2 mM
60	DNA (45-MER)	natural abundance		0.2 mM
61	TRIS	[U-2H]		50 mM
62	DTT	[U-2H]		1 mM
63	ZnSO4	natural abundance		0.1 mM
64	H2O	natural abundance	solvent	95 %
65	D2O	[U-2H]	solvent	5 %

Sample #11

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a and [1H,13C,15N] Phe residues Labelling for residues 105-214 (and GGG of insertion): uniform [1H,12C,15N]

#	Name	Isotope labeling	Type	Concentration
66	PARP-1 1-214	see Sample details section		0.2 mM
67	DNA (45-MER)	natural abundance		0.2 mM
68	TRIS	[U-2H]		50 mM
69	DTT	[U-2H]		1 mM
70	ZnSO4	natural abundance		0.1 mM
71	D2O	[U-2H]	solvent	100 %

Sample #12

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a Labelling for residues 105-214 (and GGG of insertion): uniform [2H,12C,15N] back-labeled with [1H,15N,13C] Pro

#	Name	Isotope labeling	Type	Concentration
72	PARP-1 1-214	see Sample details section		0.2 mM
73	DNA (45-MER)	natural abundance		0.2 mM
74	TRIS	[U-2H]		50 mM
75	DTT	[U-2H]		1 mM
76	ZnSO4	natural abundance		0.1 mM
77	D2O	[U-2H]	solvent	100 %

Sample #13

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
78	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
79	DNA (45-MER)	natural abundance		0.2 mM
80	TRIS	[U-2H]		50 mM
81	DTT	[U-2H]		1 mM
82	ZnSO4	natural abundance		0.1 mM
83	D2O	[U-2H]	solvent	100 %

Sample #14

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
84	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
85	DNA (45-MER)	natural abundance		0.2 mM
86	TRIS	[U-2H]		50 mM
87	DTT	[U-2H]		1 mM
88	ZnSO4	natural abundance		0.1 uM
89	sodium chloride	natural abundance		200 mM
90	H2O	natural abundance	solvent	95 %
91	D2O	[U-2H]	solvent	5 %

Hide sample codition

Chem. Shift Complete2

Sequence coverage: 75.3 %, Completeness: 26.9 %, Completeness (bb): 38.9 % Detail

Download

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Polymer type: polypeptide(L) polydeoxyribonucleotide

	Total	1H	13C	15N
All	26.9 % (1593 of 5914)	19.5 % (690 of 3546)	26.2 % (506 of 1930)	90.6 % (397 of 438)
Backbone	38.9 % (1230 of 3162)	30.1 % (454 of 1508)	30.6 % (381 of 1244)	96.3 % (395 of 410)
Sidechain	13.6 % (427 of 3140)	11.6 % (236 of 2038)	17.6 % (189 of 1074)	7.1 % (2 of 28)
Aromatic	5.6 % (32 of 572)	4.8 % (18 of 376)	6.4 % (12 of 188)	25.0 % (2 of 8)
Methyl	45.6 % (154 of 338)	42.7 % (76 of 178)	48.8 % (78 of 160)

1. F1F2

MAESSDKLYR VEYAKSGRAS CKKCSESIPK DSLRMAIMVQ SPMFDGKVPH WYHFSCFWKV GHSIRHPDVE VDGFSELRWD DQQKVKKTAE AGGVTGKGQD GIGSKAEKTL GDFAAEYAKS NRSTCKGCME KIEKGQVRLS KKMVDPEKPQ LGMIDRWYHP GCFVKNREEL GFRPEYSASQ LKGFSLLATE DKEALKKQLP GVKSEGKRKG DEVD

2. DNA (45-MER)

GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCG

Show sample condition

Sample #1

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
1	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
2	DNA (45-MER)	natural abundance		0.2 mM
3	TRIS	[U-2H]		50 mM
4	DTT	[U-2H]		1 mM
5	ZnSO4	natural abundance		0.1 mM
6	H2O	natural abundance	solvent	95 %
7	D2O	[U-2H]	solvent	5 %

Sample #2

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H (approx. 70%),15N,13C]. This sample was only used to confirm assignments by checking patterns of peaks; no numerical shift data from experiments with this sample contributed to the deposition (since isotope shifts could differ).

#	Name	Isotope labeling	Type	Concentration
8	PARP-1 1-214	[U-15N; U-13C; U-70% 2H]		0.2 mM
9	DNA (45-MER)	natural abundance		0.2 mM
10	TRIS	[U-2H]		50 mM
11	DTT	[U-2H]		1 mM
12	ZnSO4	natural abundance		0.1 mM
13	H2O	natural abundance	solvent	95 %
14	D2O	[U-2H]	solvent	5 %

Sample #3

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N]

#	Name	Isotope labeling	Type	Concentration
15	PARP-1 1-214	[U-98% 2H; U-98% 15N]		0.2 mM
16	DNA (45-MER)	natural abundance		0.2 mM
17	TRIS	[U-2H]		50 mM
18	DTT	[U-2H]		1 mM
19	ZnSO4	natural abundance		0.1 mM
20	H2O	natural abundance	solvent	95 %
21	D2O	[U-2H]	solvent	5 %

Sample #4

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the delta-methyl groups of Ile and all methyl groups of Leu and Val residues, using sodium [4-13C, 3,3-2H2] alpha-ketobutyrate and sodium [3- 2H, 4,4'-13C2] alpha-ketoisovalerate as precursors to maximize protonation of methyl groups, for use in NOE experiments; sodium [3-2H, 4,4'-13C2] alpha-ketoisovalerate was prepared from sodium [4,4'-13C2] alpha-ketoisovalerate by exchange with 2H2O at pH 12.5 and 45 C for 3 hrs.

#	Name	Isotope labeling	Type	Concentration
22	PARP-1 1-214	see Sample details section		0.2 mM
23	DNA (45-MER)	natural abundance		0.2 mM
24	TRIS	[U-2H]		50 mM
25	DTT	[U-2H]		1 mM
26	ZnSO4	natural abundance		0.1 mM
27	H2O	natural abundance	solvent	95 %
28	D2O	[U-2H]	solvent	5 %

Sample #5

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the delta-methyl groups of Ile and all methyl groups of Leu and Val residues, using sodium [3,3-2H2,13C4] alpha-ketobutyrate and sodium [3- 2H,13C5] alpha-ketoisovalerate as precursors to produce linear chains of 13C in the sidechains of Val and Leu, for use in assignment experiments to link methyl signals to C-alpha signals.

#	Name	Isotope labeling	Type	Concentration
29	PARP-1 1-214	see Sample details section		0.2 mM
30	DNA (45-MER)	natural abundance		0.2 mM
31	TRIS	[U-2H]		50 mM
32	DTT	[U-2H]		1 mM
33	ZnSO4	natural abundance		0.1 mM
34	D2O	[U-2H]	solvent	100 %

Sample #6

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the methyl groups of Met residues in addition to Ile, Leu and Val methyl groups as in sample 4a.

#	Name	Isotope labeling	Type	Concentration
35	PARP-1 1-214	see Sample details section		0.2 mM
36	DNA (45-MER)	natural abundance		0.2 mM
37	TRIS	[U-2H]		50 mM
38	DTT	[U-2H]		1 mM
39	ZnSO4	natural abundance		0.1 mM
40	D2O	[U-2H]	solvent	100 %

Sample #7

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]; back-labeled with [1H,13C] in the methyl groups of Ile, Leu and Val methyl groups as in sample 4a and [1H,13C,15N] Phe residues.

#	Name	Isotope labeling	Type	Concentration
41	PARP-1 1-214	see Sample details section		0.2 mM
42	DNA (45-MER)	natural abundance		0.2 mM
43	TRIS	[U-2H]		50 mM
44	DTT	[U-2H]		1 mM
45	ZnSO4	natural abundance		0.1 mM
46	D2O	[U-2H]	solvent	100 %

Sample #8

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): uniform [1H,12C,15N] Labelling for residues 105-214 (and GGG of insertion): [2H,15N,13C] back-labeled with [1H,13C] Ile, Leu Val and Met methyl groups labeled as in sample 5

#	Name	Isotope labeling	Type	Concentration
47	PARP-1 1-214	see Sample details section		0.2 mM
48	DNA (45-MER)	natural abundance		0.2 mM
49	TRIS	[U-2H]		50 mM
50	DTT	[U-2H]		1 mM
51	ZnSO4	natural abundance		0.1 mM
52	D2O	[U-2H]	solvent	100 %

Sample #9

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): uniform [1H,12C,15N] Labelling for residues 105-214 (and GGG of insertion): [2H,15N,13C] back labeled with [1H,13C] Met methyl groups as in sample 5 and [13C,15N,1H] Arg residues

#	Name	Isotope labeling	Type	Concentration
53	PARP-1 1-214	see Sample details section		0.2 mM
54	DNA (45-MER)	natural abundance		0.2 mM
55	TRIS	[U-2H]		50 mM
56	DTT	[U-2H]		1 mM
57	ZnSO4	natural abundance		0.1 mM
58	D2O	[U-2H]	solvent	100 %

Sample #10

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a and [1H,13C,15N] Arg residues Labelling for residues 105-214 (and GGG of insertion): uniform [1H,12C,15N]

#	Name	Isotope labeling	Type	Concentration
59	PARP-1 1-214	see Sample details section		0.2 mM
60	DNA (45-MER)	natural abundance		0.2 mM
61	TRIS	[U-2H]		50 mM
62	DTT	[U-2H]		1 mM
63	ZnSO4	natural abundance		0.1 mM
64	H2O	natural abundance	solvent	95 %
65	D2O	[U-2H]	solvent	5 %

Sample #11

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a and [1H,13C,15N] Phe residues Labelling for residues 105-214 (and GGG of insertion): uniform [1H,12C,15N]

#	Name	Isotope labeling	Type	Concentration
66	PARP-1 1-214	see Sample details section		0.2 mM
67	DNA (45-MER)	natural abundance		0.2 mM
68	TRIS	[U-2H]		50 mM
69	DTT	[U-2H]		1 mM
70	ZnSO4	natural abundance		0.1 mM
71	D2O	[U-2H]	solvent	100 %

Sample #12

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a Labelling for residues 105-214 (and GGG of insertion): uniform [2H,12C,15N] back-labeled with [1H,15N,13C] Pro

#	Name	Isotope labeling	Type	Concentration
72	PARP-1 1-214	see Sample details section		0.2 mM
73	DNA (45-MER)	natural abundance		0.2 mM
74	TRIS	[U-2H]		50 mM
75	DTT	[U-2H]		1 mM
76	ZnSO4	natural abundance		0.1 mM
77	D2O	[U-2H]	solvent	100 %

Sample #13

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
78	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
79	DNA (45-MER)	natural abundance		0.2 mM
80	TRIS	[U-2H]		50 mM
81	DTT	[U-2H]		1 mM
82	ZnSO4	natural abundance		0.1 mM
83	D2O	[U-2H]	solvent	100 %

Sample #14

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
84	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
85	DNA (45-MER)	natural abundance		0.2 mM
86	TRIS	[U-2H]		50 mM
87	DTT	[U-2H]		1 mM
88	ZnSO4	natural abundance		0.1 uM
89	sodium chloride	natural abundance		200 mM
90	H2O	natural abundance	solvent	95 %
91	D2O	[U-2H]	solvent	5 %

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Chem. Shift Complete3

Sequence coverage: 34.7 %, Completeness: 24.8 %, Completeness (bb): 34.4 % Detail

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Polymer type: polypeptide(L) polydeoxyribonucleotide

	Total	1H	13C	15N
All	24.8 % (2199 of 8871)	20.8 % (1107 of 5319)	22.5 % (651 of 2895)	67.1 % (441 of 657)
Backbone	34.4 % (1632 of 4743)	30.6 % (693 of 2262)	26.8 % (501 of 1866)	71.2 % (438 of 615)
Sidechain	14.2 % (670 of 4710)	13.5 % (414 of 3057)	15.7 % (253 of 1611)	7.1 % (3 of 42)
Aromatic	13.1 % (112 of 858)	17.4 % (98 of 564)	4.3 % (12 of 282)	16.7 % (2 of 12)
Methyl	33.1 % (168 of 507)	33.0 % (88 of 267)	33.3 % (80 of 240)

1. F1F2

MAESSDKLYR VEYAKSGRAS CKKCSESIPK DSLRMAIMVQ SPMFDGKVPH WYHFSCFWKV GHSIRHPDVE VDGFSELRWD DQQKVKKTAE AGGVTGKGQD GIGSKAEKTL GDFAAEYAKS NRSTCKGCME KIEKGQVRLS KKMVDPEKPQ LGMIDRWYHP GCFVKNREEL GFRPEYSASQ LKGFSLLATE DKEALKKQLP GVKSEGKRKG DEVD

2. DNA (45-MER)

GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCG

Show sample condition

Sample #1

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
1	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
2	DNA (45-MER)	natural abundance		0.2 mM
3	TRIS	[U-2H]		50 mM
4	DTT	[U-2H]		1 mM
5	ZnSO4	natural abundance		0.1 mM
6	H2O	natural abundance	solvent	95 %
7	D2O	[U-2H]	solvent	5 %

Sample #2

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H (approx. 70%),15N,13C]. This sample was only used to confirm assignments by checking patterns of peaks; no numerical shift data from experiments with this sample contributed to the deposition (since isotope shifts could differ).

#	Name	Isotope labeling	Type	Concentration
8	PARP-1 1-214	[U-15N; U-13C; U-70% 2H]		0.2 mM
9	DNA (45-MER)	natural abundance		0.2 mM
10	TRIS	[U-2H]		50 mM
11	DTT	[U-2H]		1 mM
12	ZnSO4	natural abundance		0.1 mM
13	H2O	natural abundance	solvent	95 %
14	D2O	[U-2H]	solvent	5 %

Sample #3

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N]

#	Name	Isotope labeling	Type	Concentration
15	PARP-1 1-214	[U-98% 2H; U-98% 15N]		0.2 mM
16	DNA (45-MER)	natural abundance		0.2 mM
17	TRIS	[U-2H]		50 mM
18	DTT	[U-2H]		1 mM
19	ZnSO4	natural abundance		0.1 mM
20	H2O	natural abundance	solvent	95 %
21	D2O	[U-2H]	solvent	5 %

Sample #4

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the delta-methyl groups of Ile and all methyl groups of Leu and Val residues, using sodium [4-13C, 3,3-2H2] alpha-ketobutyrate and sodium [3- 2H, 4,4'-13C2] alpha-ketoisovalerate as precursors to maximize protonation of methyl groups, for use in NOE experiments; sodium [3-2H, 4,4'-13C2] alpha-ketoisovalerate was prepared from sodium [4,4'-13C2] alpha-ketoisovalerate by exchange with 2H2O at pH 12.5 and 45 C for 3 hrs.

#	Name	Isotope labeling	Type	Concentration
22	PARP-1 1-214	see Sample details section		0.2 mM
23	DNA (45-MER)	natural abundance		0.2 mM
24	TRIS	[U-2H]		50 mM
25	DTT	[U-2H]		1 mM
26	ZnSO4	natural abundance		0.1 mM
27	H2O	natural abundance	solvent	95 %
28	D2O	[U-2H]	solvent	5 %

Sample #5

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the delta-methyl groups of Ile and all methyl groups of Leu and Val residues, using sodium [3,3-2H2,13C4] alpha-ketobutyrate and sodium [3- 2H,13C5] alpha-ketoisovalerate as precursors to produce linear chains of 13C in the sidechains of Val and Leu, for use in assignment experiments to link methyl signals to C-alpha signals.

#	Name	Isotope labeling	Type	Concentration
29	PARP-1 1-214	see Sample details section		0.2 mM
30	DNA (45-MER)	natural abundance		0.2 mM
31	TRIS	[U-2H]		50 mM
32	DTT	[U-2H]		1 mM
33	ZnSO4	natural abundance		0.1 mM
34	D2O	[U-2H]	solvent	100 %

Sample #6

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C], back-labeled with [1H,13C] in the methyl groups of Met residues in addition to Ile, Leu and Val methyl groups as in sample 4a.

#	Name	Isotope labeling	Type	Concentration
35	PARP-1 1-214	see Sample details section		0.2 mM
36	DNA (45-MER)	natural abundance		0.2 mM
37	TRIS	[U-2H]		50 mM
38	DTT	[U-2H]		1 mM
39	ZnSO4	natural abundance		0.1 mM
40	D2O	[U-2H]	solvent	100 %

Sample #7

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]; back-labeled with [1H,13C] in the methyl groups of Ile, Leu and Val methyl groups as in sample 4a and [1H,13C,15N] Phe residues.

#	Name	Isotope labeling	Type	Concentration
41	PARP-1 1-214	see Sample details section		0.2 mM
42	DNA (45-MER)	natural abundance		0.2 mM
43	TRIS	[U-2H]		50 mM
44	DTT	[U-2H]		1 mM
45	ZnSO4	natural abundance		0.1 mM
46	D2O	[U-2H]	solvent	100 %

Sample #8

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): uniform [1H,12C,15N] Labelling for residues 105-214 (and GGG of insertion): [2H,15N,13C] back-labeled with [1H,13C] Ile, Leu Val and Met methyl groups labeled as in sample 5

#	Name	Isotope labeling	Type	Concentration
47	PARP-1 1-214	see Sample details section		0.2 mM
48	DNA (45-MER)	natural abundance		0.2 mM
49	TRIS	[U-2H]		50 mM
50	DTT	[U-2H]		1 mM
51	ZnSO4	natural abundance		0.1 mM
52	D2O	[U-2H]	solvent	100 %

Sample #9

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): uniform [1H,12C,15N] Labelling for residues 105-214 (and GGG of insertion): [2H,15N,13C] back labeled with [1H,13C] Met methyl groups as in sample 5 and [13C,15N,1H] Arg residues

#	Name	Isotope labeling	Type	Concentration
53	PARP-1 1-214	see Sample details section		0.2 mM
54	DNA (45-MER)	natural abundance		0.2 mM
55	TRIS	[U-2H]		50 mM
56	DTT	[U-2H]		1 mM
57	ZnSO4	natural abundance		0.1 mM
58	D2O	[U-2H]	solvent	100 %

Sample #10

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a and [1H,13C,15N] Arg residues Labelling for residues 105-214 (and GGG of insertion): uniform [1H,12C,15N]

#	Name	Isotope labeling	Type	Concentration
59	PARP-1 1-214	see Sample details section		0.2 mM
60	DNA (45-MER)	natural abundance		0.2 mM
61	TRIS	[U-2H]		50 mM
62	DTT	[U-2H]		1 mM
63	ZnSO4	natural abundance		0.1 mM
64	H2O	natural abundance	solvent	95 %
65	D2O	[U-2H]	solvent	5 %

Sample #11

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a and [1H,13C,15N] Phe residues Labelling for residues 105-214 (and GGG of insertion): uniform [1H,12C,15N]

#	Name	Isotope labeling	Type	Concentration
66	PARP-1 1-214	see Sample details section		0.2 mM
67	DNA (45-MER)	natural abundance		0.2 mM
68	TRIS	[U-2H]		50 mM
69	DTT	[U-2H]		1 mM
70	ZnSO4	natural abundance		0.1 mM
71	D2O	[U-2H]	solvent	100 %

Sample #12

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Sortase ligated, block-labelled sample. (N.B. residues 103 and 104 of WT sequence deleted, additional residues LPETGGG inserted between residues 102 and 105; this sample was not used for making any assignments of residues in this region, which is in the flexible linker between domains). Labelling for residues 1-102 (and LPET of insertion): [2H,15N,13C] back-labeled with Ile, Leu and Val methyl groups labeled as in sample 4a Labelling for residues 105-214 (and GGG of insertion): uniform [2H,12C,15N] back-labeled with [1H,15N,13C] Pro

#	Name	Isotope labeling	Type	Concentration
72	PARP-1 1-214	see Sample details section		0.2 mM
73	DNA (45-MER)	natural abundance		0.2 mM
74	TRIS	[U-2H]		50 mM
75	DTT	[U-2H]		1 mM
76	ZnSO4	natural abundance		0.1 mM
77	D2O	[U-2H]	solvent	100 %

Sample #13

Solvent system 100% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
78	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
79	DNA (45-MER)	natural abundance		0.2 mM
80	TRIS	[U-2H]		50 mM
81	DTT	[U-2H]		1 mM
82	ZnSO4	natural abundance		0.1 mM
83	D2O	[U-2H]	solvent	100 %

Sample #14

Solvent system 95% H2O/5% D2O, Pressure 1 atm, Temperature 273 K, pH 7.2, Details Uniform [2H,15N,13C]

#	Name	Isotope labeling	Type	Concentration
84	PARP-1 1-214	[U-15N; U-13C; U-2H]		0.2 mM
85	DNA (45-MER)	natural abundance		0.2 mM
86	TRIS	[U-2H]		50 mM
87	DTT	[U-2H]		1 mM
88	ZnSO4	natural abundance		0.1 uM
89	sodium chloride	natural abundance		200 mM
90	H2O	natural abundance	solvent	95 %
91	D2O	[U-2H]	solvent	5 %

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Protein Blocks Logo

Calculated from 78 models in PDB: 2N8A, Strand ID: A Detail

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Release date

2015-11-24

Citation

Structural Basis of Detection and Signaling of DNA Single-Strand Breaks by Human PARP-1
Eustermann, S., Wu, W., Langelier, M., Yang, J., Easton, L.E., Riccio, A., Pascal, J.M., Neuhaus, D.
Mol. Cell (2015), 60, 742-754, PubMed 26626479 , DOI 10.1016/j.molcel.2015.10.032 , Abstract

Related entities 1. F1F2, : 1 : 2 : 23 entities Detail

Experiments performed 26 experiments Detail

NMR combined restraints 4 contents Detail