Automated amino acid side-chain NMR assignment of proteins using 13C- and 15N-resolved 3D [1H,1H]-NOESY
Automated amino acid side-chain NMR assignment of proteins using 13C- and 15N-resolved 3D [1H,1H]-NOESY
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DOI：
10.1007/s10858-008-9259-x
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发表时间：
2008-09-01
影响因子：
2.7
通讯作者：
Wuethrich, Kurt
中科院分区：
文献类型：
--
作者：
Fiorito, Francesco;Herrmann, Torsten;Wuethrich, Kurt
ASCAN是一种用于蛋白质中氨基酸侧链的自动序列特异性核磁共振指定的新算法，它使用蛋白质的一级结构、先前多肽主干指定的(HN)-H-1、N-15、C-13(α)、C-13(β)和可能的H-1(α)的化学位移列表，以及一个或多个3D C-13-或N-15-分辨的[H-1，H-1]-NOESY谱。ASCAN还用于使用TOCSY类型的数据集作为补充输入。该程序根据从化学结构预期的核磁共振信号与实验观察到的NOESY峰型的比较来分配新的共振。该算法的核心部分是生成预期峰位置的程序，该程序基于在分配过程中针对不同氨基酸类型产生的分配和未分配共振的可变组合，以及基于所使用的核磁共振实验的对应的一组分配的接受标准。使用先前指定的共振峰的列表和从球状蛋白质的BMRB统计中获得的未指定信号的暂定化学移动值，生成涉及未指定共振的NOESY交叉峰的预期模式。将该方法用于101个氨基酸残基的蛋白质FimD(25-125)，结果表明84%的氢原子及其共价结合的重原子被指定为90%的正确率。使用这些侧链指定作为自动NOE指定和ATNOS/CANTID/DYANA程序套件的结构计算的输入，产生了质量相当的结构束，就原子坐标的精密度和准确度而言，与通过交互指定程序确定的参考结构的结构束类似。基于Ascan的结构测定高质量的基本原理是对所分配的侧链的分布进行分析，这表明在蛋白质的核心中几乎完成了所分配的残基，大多数不完全分配的残基位于蛋白质表面或附近。
ASCAN is a new algorithm for automatic sequence-specific NMR assignment of amino acid side-chains in proteins, which uses as input the primary structure of the protein, chemical shift lists of (HN)-H-1, N-15, C-13(alpha), C-13(beta) and possibly H-1(alpha) from the previous polypeptide backbone assignment, and one or several 3D C-13- or N-15-resolved [H-1,H-1]-NOESY spectra. ASCAN has also been laid out for the use of TOCSY-type data sets as supplementary input. The program assigns new resonances based on comparison of the NMR signals expected from the chemical structure with the experimentally observed NOESY peak patterns. The core parts of the algorithm are a procedure for generating expected peak positions, which is based on variable combinations of assigned and unassigned resonances that arise for the different amino acid types during the assignment procedure, and a corresponding set of acceptance criteria for assignments based on the NMR experiments used. Expected patterns of NOESY cross peaks involving unassigned resonances are generated using the list of previously assigned resonances, and tentative chemical shift values for the unassigned signals taken from the BMRB statistics for globular proteins. Use of this approach with the 101-amino acid residue protein FimD(25-125) resulted in 84% of the hydrogen atoms and their covalently bound heavy atoms being assigned with a correctness rate of 90%. Use of these side-chain assignments as input for automated NOE assignment and structure calculation with the ATNOS/CANDID/DYANA program suite yielded structure bundles of comparable quality, in terms of precision and accuracy of the atomic coordinates, as those of a reference structure determined with interactive assignment procedures. A rationale for the high quality of the ASCAN-based structure determination results from an analysis of the distribution of the assigned side chains, which revealed near-complete assignments in the core of the protein, with most of the incompletely assigned residues located at or near the protein surface.