4 mM uniform 15N-labelled Arc repressor NL11/LN12 (Switch Arc)
90% H2O/10% D2O
2
7.5 mM 10% 13C-labelled Arc repressor NL11/LN12 (Switch Arc)
90% H2O, 10% D20
3
5 mM uniform 15N-labelled Arc repressor NL11/LN12 (Switch Arc)
100% D2O
試料状態
Conditions-ID
イオン強度
pH
圧 (kPa)
温度 (K)
1
20mMphosphate
4.8
ambient
303K
2
20mMphosphate
4.8
ambient
303K
3
20mMphosphate, 150mMKCl
4.67
ambient
303K
結晶化
*PLUS
手法: その他 / 詳細: NMR
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NMR測定
放射
プロトコル: SINGLE WAVELENGTH / 単色(M)・ラウエ(L): M
放射波長
相対比: 1
NMRスペクトロメーター
タイプ: Bruker DMX / 製造業者: Bruker / モデル: DMX / 磁場強度: 500 MHz
-
解析
NMR software
名称
バージョン
開発者
分類
XwinNMR
2.1
Bruker
collection
NMRPipe
1.1
Delaglio, Grzesiek, Vuister, Zhu, Pfeifer, Bax
解析
NMRDraw
2.1
Delaglio, Grzesiek, Vuister, Zhu, Pfeifer, Bax
解析
NMRView
3.1
Johnston, Blevins
データ解析
X-PLOR
3.1
Brunger
構造決定
X-PLOR
3.1
Brunger
精密化
精密化
手法: simulated annealing / ソフトェア番号: 1 詳細: Experimentally derived restraints included 810 nOe distance restraints per monomer, along with 3 hydrogen bond restraints, 31 phi-angle restraints and 3 chi1-angle restraints, for a total of ...詳細: Experimentally derived restraints included 810 nOe distance restraints per monomer, along with 3 hydrogen bond restraints, 31 phi-angle restraints and 3 chi1-angle restraints, for a total of 847 restraints per monomer spanning residues 5-53. Structure calculations were performed using a simulated annealing protocol designed for symmetric dimers (M. Nilges, Proteins Struct. Funct. Gen, 17, 297 (1993)). As starting points for the calculation, 28 structures were generated in which the conformation of the N-terminal region (residues 1-13) was random, and that of the remainder of the protein restrained to be similar to wild-type Arc. The use of completely random starting structures led to extremely poor convergence rates, but those calculations which did converge yielded structures similar to those obtained from calculations using non-random starting structures. Each semi-random starting structure was then annealed to a model structure using the distance, angle and hydrogen bond restraints mentioned above. Two "seed" restraints, Arg 40 HE-Phe 45 HD and Trp 14 HE3-Tyr 38 HE, were described as unambiguously intermolecular. In the wild-type Arc structure, the difference between the intra and intermolecular distances for these atoms is >10 A. Use of the seed restraints improved convergence but again did not alter the qualitative nature of the results. All other NOE distance restraints were described ambiguously using sum potentials. Hydrogen-bond restraints in alpha-helices were described as intramonomer. In the conformational search phase of the calculations, non-bonded interactions were computed only between C-alpha atoms with a van der Waals term of 0.1. 13 of 28 calculated structures were accepted with no angle violations >5 degrees and no more than two nOe violations >0.35 A. These comprise the ensemble submitted here.
代表構造
選択基準: closest to the average
NMRアンサンブル
コンフォーマー選択の基準: structures with the least restraint violations 計算したコンフォーマーの数: 28 / 登録したコンフォーマーの数: 13