Utility of methyl side chain probes for solution NMR studies of large proteins

Andrew C. McShan
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引用次数: 2

Abstract

Selective isotopic labeling of methyl side chain groups in proteins and other biomolecules, combined with advances in perdeuteration, new pulse sequences, and high field spectrometers with cryogenic probes, has revolutionized the field of solution nuclear magnetic resonance (NMR) spectroscopy by enabling characterization of macromolecular systems with molecular weights above 1 MDa in their native aqueous environment. This tutorial provides a basic overview for how modern NMR spectroscopists can utilize methyl side chain probes to study their system of interest. The advantages and limitations of methyl side chain probes are discussed. In addition, the preparation of selectively 13C-methyl labeled recombinant protein samples, strategies for manual and automated methyl NMR resonance assignment, and the application of methyl probes for characterization of dynamics and conformational exchange are discussed. A sneak preview for ways in which methyl probes are expected to continue to advance the field of biomolecular NMR towards new horizons in solution studies of large supramolecular complexes is also presented.

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甲基侧链探针在大蛋白质溶液核磁共振研究中的应用
蛋白质和其他生物分子中甲基侧链基团的选择性同位素标记,结合渗透氘化、新脉冲序列和带低温探针的高场光谱仪的进展,通过在其原生水环境中表征分子量大于1mda的大分子体系,彻底改变了溶液核磁共振(NMR)光谱领域。本教程提供了现代核磁共振波谱学家如何利用甲基侧链探针来研究他们感兴趣的系统的基本概述。讨论了甲基侧链探针的优点和局限性。此外,还讨论了选择性13c -甲基标记重组蛋白样品的制备、手工和自动甲基核磁共振分配策略以及甲基探针在表征动力学和构象交换中的应用。还介绍了甲基探针有望继续推进生物分子核磁共振领域在大型超分子复合物溶液研究中的新领域的方法。
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