1H, 13C and 15N chemical shift assignments and solution structure for PARP-1 F1F2 domains in complex with a DNA single-strand break
MAESSDKLYR VEYAKSGRAS CKKCSESIPK DSLRMAIMVQ SPMFDGKVPH WYHFSCFWKV GHSIRHPDVE VDGFSELRWD DQQKVKKTAE AGGVTGKGQD GIGSKAEKTL GDFAAEYAKS NRSTCKGCME KIEKGQVRLS KKMVDPEKPQ LGMIDRWYHP GCFVKNREEL GFRPEYSASQ LKGFSLLATE DKEALKKQLP GVKSEGKRKG DEVD
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 |
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 DEVD2. DNA (45-MER)
GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCGSolvent 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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 DEVD2. DNA (45-MER)
GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCGSolvent 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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 DEVD2. DNA (45-MER)
GCTGGCTTCG TAAGAAGCCA GCTCGCGGTC AGCTTGCTGA CCGCGSolvent 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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |
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 % |