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13C-detected NMR experiments for automatic resonance assignment of IDPs and multiple-fixing SMFT processing

机译:13C检测的NMR实验用于IDP的自动共振分配和多重固定SMFT处理

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摘要

Intrinsically disordered proteins (IDPs) have recently attracted much interest, due to their role in many biological processes, including signaling and regulation mechanisms. High-dimensional 13C direct-detected NMR experiments have proven exceptionally useful in case of IDPs, providing spectra with superior peak dispersion. Here, two such novel experiments recorded with non-uniform sampling are introduced, these are 5D HabCabCO(CA)NCO and 5D HNCO(CA)NCO. Together with the 4D (HACA)CON(CA)NCO, an extension of the previously published 3D experiments (Pantoja-Uceda and Santoro in J Biomol NMR 59:43–50, . doi:10.1007/s10858-014-9827-1), they form a set allowing for complete and reliable resonance assignment of difficult IDPs. The processing is performed with sparse multidimensional Fourier transform based on the concept of restricting (fixing) some of spectral dimensions to a priori known resonance frequencies. In our study, a multiple-fixing method was developed, that allows easy access to spectral data. The experiments were tested on a resolution-demanding alpha-synuclein sample. Due to superior peak dispersion in high-dimensional spectrum and availability of the sequential connectivities between four consecutive residues, the overwhelming majority of resonances could be assigned automatically using the TSAR program. Electronic supplementary materialThe online version of this article (doi:10.1007/s10858-015-9932-9) contains supplementary material, which is available to authorized users.
机译:由于其在许多生物过程(包括信号传导和调节机制)中的作用,内在无序蛋白(IDP)最近引起了人们的极大兴趣。高维 13 C直接检测NMR实验在IDP的情况下被证明格外有用,它提供了具有出色峰分散的光谱。在这里,介绍了两个用非均匀采样记录的此类新颖实验,它们是5D HabCabCO(CA)NCO和5D HNCO(CA)NCO。与4D(HACA)CON(CA)NCO一起,是先前已发表的3D实验的扩展(Pantoja-Uceda和Santoro的J Biomol NMR 59:43–50 ,. doi:10.1007 / s10858-014-9827-1) ,它们形成了一个集合,可以对困难的IDP进行完全可靠的谐振分配。基于将某些频谱尺寸限制(固定)到先验已知共振频率的概念,利用稀疏多维傅立叶变换执行该处理。在我们的研究中,开发了一种多重固定方法,可以轻松访问光谱数据。在要求分辨率的α-突触核蛋白样品上测试了实验。由于在高维光谱中具有出色的峰分散性,并且在四个连续残基之间具有连续的连通性,因此可以使用TSAR程序自动分配绝大多数共振。电子补充材料本文的在线版本(doi:10.1007 / s10858-015-9932-9)包含补充材料,授权用户可以使用。

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