TROSY triple-resonance four-dimensional NMR spectroscopy of a 46 ns tumbling protein
TROSY triple-resonance four-dimensional NMR spectroscopy of a 46 ns tumbling protein
复制标题
DOI：
10.1021/ja984056t
复制
发表时间：
1999-03-24
影响因子：
15
通讯作者：
Kay, LE
中科院分区：
文献类型：
--
作者：
Yang, DW;Kay, LE
两个四维TROSY三重共振为基础的实验提出了高分子量蛋白质和蛋白质复合物的骨架分配。实验4D-HNCACO和4D-HNCOCA分别建立了形式（C-13（（i，i-1）alpha），C-13 '（（i，i-1）），N-15（（i）），HN（i））和（C-13（（i-1）alpha），C-13'（（i-1）），N-15（（i）），HN（i））的相关性。这两个序列都使用TROSY的实现，其相对于先前的序列提供了改进的灵敏度，这对于应用于具有40-50 ns量级的相关时间的系统至关重要。实验的效用被证明通过应用到46 ns翻滚复合物的370个残基的麦芽糖结合蛋白和β-环糊精。在HNCACO和HNCOCA中分别观察到约95%的预期残基内和残基间相关性，平均信噪比值约为35/1。该方法有望成为特别强大的应用程序，以高分子量的复合物组成的标记片段和未标记的组件或蛋白质与片段标记。
Two four-dimensional TROSY triple resonance based experiments are presented for backbone assignment of high molecular weight proteins and protein complexes. The experiments, 4D-HNCACO and 4D-HNCOCA, establish correlations Of the form (C-13((i,i-1)alpha),C-13'((i,i-1)),N-15((i)),HN(i)) and (C-13((i-1)alpha),C-13'((i-1)),N-15((i)),HN(i)), respectively. Both sequences use an implementation of TROSY that offers improved sensitivity relative to previous sequences, critical for application to systems with correlation times on the order of 40-50 ns. The utility of the experiments is demonstrated by an application to a 46 ns tumbling complex of the 370 residue maltose binding protein and beta-cyclodextrin. Approximately 95% of the expected intra- and interresidue correlations are observed in the HNCACO and HNCOCA, respectively, with average signal-to-noise values of approximately 35/1. The methodology promises to be particularly powerful for applications to high molecular weight complexes comprised of a labeled fragment and unlabeled components or proteins with segmental labeling.