通过通过溶解氧的旋转放松增强减少实验时间
Louis-Philippe Picard1, Dmitry Pichugin1, Shuya Kate Huang1
1Department of Chemistry, University of Toronto, Toronto, Canada.
Journal of biomolecular NMR
|February 17, 2025
概括
在蛋白质样本中溶解氧气显著加快了质子 (1H) 和-19 (19F) 旋转放松率. 这种方法将NMR实验时间缩短多达三倍,为蛋白质NMR研究提供了一种实用方法.
科学领域:
- 生物物理学的生物物理.
- 结构生物学 结构生物学
- 核磁共振 (NMR) 光谱学 核磁共振 (NMR) 光谱学
背景情况:
- 长质子 (1H) 旋转放松时间 (T1),在大型蛋白质和稀释旋转系统中常见,在NMR中增加实验持续时间.
- 二氧化物 (O2) 是一种超磁性物种,有效地提高了高玛核的自旋晶格放松率 (R1),包括1H和19F.
研究的目的:
- 为了研究溶解氧对各种蛋白质系统中1H和19F旋转放松的影响.
- 评估使用氧化的NMR实验节省时间的潜力.
- 评估标准NMR设置中氧化的稳定性和实际应用.
主要方法:
- 蛋白质样本的氧化 (酸脱酶,A2A受体,牛血清白蛋白) 用O2在9-10bar.
- 在氧化之前和之后测量1H和19F自旋格子放松率 (R1).
- 获取1H-15N HSQC和19F NMR光谱以评估节省时间.
- 在标准NMR管中分析T1放松和氧气保留.
主要成果:
- 在~9bar的氧化时,1H和19F R1速率增加到3-5 Hz,而没有氧气则是0.7-1.0 Hz.
- 对于1H-15N HSQC和19F NMR频谱,实验时间缩短了三倍.
- 旋转扩散效应为暴露于溶剂和埋藏的质子均的1H旋转放松作出了贡献.
- 溶解氧水平在标准NMR管中在≤20°C下在3-4天内保持稳定.
结论:
- 溶解氧是加速蛋白质NMR中自旋格子放松的实用和有效剂.
- 这种方法在不需要专门的设备的情况下为19F,1H-13C和1H-15NNNMR研究提供了显著的时间节省.
- 能够轻松引入或去除氧气的能力使其成为优化NMR实验的通用工具.
相关概念视频
01:28Atomic Nuclei: Types of Nuclear Relaxation
239
Nuclear relaxation restores the equilibrium population imbalance and can occur via spin–lattice or spin–spin mechanisms, which are first-order exponential decay processes.
In spin–lattice or longitudinal relaxation, the excited spins exchange energy with the surrounding lattice as they return to the lower energy level. Among several mechanisms that contribute to spin–lattice relaxation, magnetic dipolar interactions are significant. Here, the excited nucleus transfers...
In spin–lattice or longitudinal relaxation, the excited spins exchange energy with the surrounding lattice as they return to the lower energy level. Among several mechanisms that contribute to spin–lattice relaxation, magnetic dipolar interactions are significant. Here, the excited nucleus transfers...
239
01:12Double Resonance Techniques: Overview
183
Double resonance techniques in Nuclear Magnetic Resonance (NMR) spectroscopy involve the simultaneous application of two different frequencies or radiofrequency pulses to manipulate and observe two distinct nuclear spins. One important application of double resonance is spin decoupling, which selectively suppresses coupling with one type of nucleus while observing the NMR signal from another nucleus, simplifying the spectrum and enhancing resolution.
Spin decoupling is usually achieved by...
Spin decoupling is usually achieved by...
183
01:23Atomic Nuclei: Nuclear Relaxation Processes
602
In the absence of an external magnetic field, nuclear spin states are degenerate and randomly oriented. When a magnetic field is applied, the spins begin to precess and orient themselves along (lower energy) or against (higher energy) the direction of the field. At equilibrium, a slight excess population of spins exists in the lower energy state. Because the direction of the magnetic field is fixed as the z-axis, the precessing magnetic moments are randomly oriented around the z-axis.
602