Department of Structural Biology, St Jude Children's Research Hospital, Memphis, TN, USA.
Department of Structural Biology, St Jude Children's Research Hospital, Memphis, TN, USA.
Structure. 2022 Jan 6;30(1):69-79.e2. doi: 10.1016/j.str.2021.11.009. Epub 2021 Dec 15.
Methyl-TROSY spectroscopy has extended the reach of solution-state NMR to supra-molecular machineries over 100 kDa in size. Methyl groups are ideal probes for studying structure, dynamics, and protein-protein interactions in quasi-physiological conditions with atomic resolution. Successful implementation of the methodology requires accurate methyl chemical shift assignment, and the task still poses a significant challenge in the field. In this work, we outline the current state of technology for methyl labeling, data collection, data analysis, and nuclear Overhauser effect (NOE)-based automated methyl assignment approaches. We present MAGIC-Act and MAGIC-View, two Python extensions developed as part of the popular NMRFAM-Sparky package, and MAGIC-Net a standalone structure-based network analysis program. MAGIC-Act conducts statistically driven amino acid typing, Leu/Val pairing guided by 3D HMBC-HMQC, and NOESY cross-peak symmetry checking. MAGIC-Net provides model-based NOE statistics to aid in selection of a methyl labeling scheme. The programs provide a versatile, semi-automated framework for rapid methyl assignment.
Methyl-TROSY 光谱学将溶液态 NMR 的应用范围扩展到了超过 100 kDa 的超分子机器，甲基基团是在接近生理条件下研究结构、动态和蛋白质-蛋白质相互作用的理想探针，具有原子分辨率。该方法的成功实施需要准确的甲基化学位移赋值，而这在该领域仍然是一个重大挑战。在这项工作中，我们概述了目前用于甲基标记、数据收集、数据分析和基于核 Overhauser 效应 (NOE) 的自动甲基赋值方法的技术现状。我们介绍了 MAGIC-Act 和 MAGIC-View，这两个 Python 扩展是作为流行的 NMRFAM-Sparky 包的一部分开发的，以及 MAGIC-Net 一个独立的基于结构的网络分析程序。MAGIC-Act 进行基于统计的氨基酸类型推断，3D HMBC-HMQC 引导的 Leu/Val 配对，以及 NOESY 交叉峰对称性检查。MAGIC-Net 提供基于模型的 NOE 统计数据，以帮助选择甲基标记方案。这些程序提供了一个通用的、半自动的快速甲基赋值框架。
