Upgrade to 600 MHz NMR spectrometer

升级至 600 MHz NMR 波谱仪

• 批准号: BB/R000727/1
• 负责人: Michael Williamson
• 金额: $ 57.38万
• 依托单位: University of Sheffield
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FR000727%2F1
• 关键词: Upgrade; 600; MHz; NMR; spectrometer

Nuclear Magnetic Resonance (NMR) is a spectroscopic technique closely related to Magnetic Resonance Imaging (MRI). It is a very versatile technique. Its biggest application lies in its ability to provide detailed information on interactions between molecules. Proteins up to about 30 kDa (250 amino acid residues) can have backbone signals assigned rapidly and almost automatically. This means that we can rapidly and simply work out which signal comes from which amino acid, thereby providing a fingerprint of the protein. Addition of a ligand (for example a small molecule drug, or another protein) produces changes in signals, which are straightforwardly analysed to determine where the ligand is binding, and often to produce a binding affinity. With larger proteins, similar information can be obtained in slightly different ways although with rather more experimental effort. Very large proteins (including membrane proteins and receptors in intact cells) are still amenable to investigation, and there are straightforward techniques for identifying which ligands bind to such targets, and which parts of the ligands are most closely in contact. NMR has many other uses. It can be used to calculate the structures of proteins in solution, to analyse local and global mobility, to look at structural change on alteration of solution conditions or addition of ligands, to measure local pKa values of charged sidechains, to analyse the metabolites present in complex mixtures such as body fluids, to follow metabolic and enzymatic processes, and much more. Extracting such information is not always straightforward and often requires expert advice and intervention.Currently we carry out many such analyses on a range of targets, often as a result of problems being investigated by external users that we are asked to advise on. Selection of projects is made on the basis of scientific importance rather than funding, although funding is clearly of considerable importance. Our current equipment is old and becoming obsolete, and is therefore likely to fail catastrophically at some point over the next few years. We are looking to replace it with new equipment, which has very similar capabilities, although the intervening years have led to improvements in many systems, for example more stable electronics and more rapid switching. The upgraded instrument will therefore be able to do the same things as our current instrument, but better and faster, with fewer instrumental artifacts and limitations. It will have some new capabilities, which will enable us to carry out a wider range of investigations, and will bring it up to the highest international standards. The current instrument has lasted 18 years: we expect the upgraded instrument to last at least another 10 and hopefully considerably longer. The funding required is thus a very cost-effective solution to the provision of world-class NMR facilities. The upgraded instrument will be used for a wide range of projects, with users mainly from Sheffield but also further afield. These include investigation of amyloid proteins (the cause of Alzheimer's disease); fundamental research on protein stability and solubility in solution; work on the mechanism of kinases and phosphoryl transferases (key regulatory enzymes within metabolic pathways; investigations on bacterial proteins that recognise specific parts of bacterial cell walls and could therefore be used as a basis for both antibiotics and novel diagnostic tools; investigations of the function of specific proteins; and the development of high-pressure NMR as a tool for investigating the relatively low-population active forms of proteins. The instrument therefore provides an important resource for many problems in biomolecular research.

核磁共振（NMR）是一种与磁共振成像（MRI）密切相关的光谱技术。这是一种非常通用的技术。它最大的应用在于能够提供分子之间相互作用的详细信息。高达约 30 kDa（250 个氨基酸残基）的蛋白质可以快速且几乎自动地分配主链信号。这意味着我们可以快速、简单地找出哪个信号来自哪个氨基酸，从而提供蛋白质的指纹。添加配体（例如小分子药物或另一种蛋白质）会产生信号变化，可以直接分析信号以确定配体的结合位置，并且通常会产生结合亲和力。对于较大的蛋白质，尽管需要更多的实验工作，但可以通过略有不同的方式获得类似的信息。非常大的蛋白质（包括完整细胞中的膜蛋白和受体）仍然适合研究，并且有直接的技术来识别哪些配体与这些靶标结合，以及配体的哪些部分接触最紧密。核磁共振还有许多其他用途。它可用于计算溶液中蛋白质的结构、分析局部和全局迁移率、观察溶液条件改变或添加配体时的结构变化、测量带电侧链的局部 pKa 值、分析溶液中存在的代谢物。复杂的混合物，如体液，遵循代谢和酶促过程等等。提取此类信息并不总是那么简单，通常需要专家的建议和干预。目前，我们对一系列目标进行了许多此类分析，通常是外部用户调查问题的结果，我们被要求提供建议。项目的选择是根据科学重要性而不是资金来进行的，尽管资金显然相当重要。我们现有的设备已经过时，因此很可能在未来几年的某个时候出现灾难性故障。我们正在寻求用新设备取代它，新设备具有非常相似的功能，尽管这些年来许多系统都得到了改进，例如更稳定的电子设备和更快速的切换。因此，升级后的仪器将能够执行与我们当前仪器相同的操作，但更好更快，仪器伪影和限制更少。它将拥有一些新的功能，使我们能够进行更广泛的调查，并使其达到最高的国际标准。目前的仪器已使用 18 年：我们预计升级后的仪器将至少再使用 10 年，甚至更长时间。因此，所需资金是提供世界一流核磁共振设施的非常具有成本效益的解决方案。升级后的仪器将用于广泛的项目，用户主要来自谢菲尔德，但也有更远的地方。其中包括淀粉样蛋白（阿尔茨海默病的病因）的研究；蛋白质在溶液中稳定性和溶解度的基础研究；研究激酶和磷酸转移酶的机制（代谢途径中的关键调节酶；研究识别细菌细胞壁特定部分的细菌蛋白，因此可用作抗生素和新型诊断工具的基础；研究因此，该仪器为生物分子研究中的许多问题提供了重要的资源。

项目成果

Improved methodology for protein NMR structure calculation using hydrogen bond restraints and ANSURR validation: the SH2 domain of SH2B1

使用氢键约束和 ANSURR 验证改进蛋白质 NMR 结构计算方法：SH2B1 的 SH2 结构域

• DOI: http://dx.10.1101/2022.08.29.505644
• 发表时间: 2022
• 作者: Fowler N

Two-site recognition of Staphylococcus aureus peptidoglycan by lysostaphin SH3b

溶葡萄球菌素 SH3b 对金黄色葡萄球菌肽聚糖的双位点识别

• DOI: http://dx.10.1038/s41589-019-0393-4
• 发表时间: 2019
• 期刊: Nature Chemical Biology
• 影响因子: 14.8
• 作者: Gonzalez

The accuracy of NMR protein structures in the Protein Data Bank.

蛋白质数据库中 NMR 蛋白质结构的准确性。

• DOI: http://dx.10.1016/j.str.2021.07.001
• 发表时间: 2021
• 期刊: 1993)
• 作者: Fowler NJ

Clostridioides difficile canonical L,D-transpeptidases catalyze a novel type of peptidoglycan cross-links and are not required for beta-lactam resistance.

艰难梭菌经典 L,D-转肽酶可催化新型肽聚糖交联，并且不是 β-内酰胺耐药性所必需的。

• DOI: http://dx.10.1016/j.jbc.2023.105529
• 发表时间: 2024
• 期刊: The Journal of biological chemistry
• 作者: Galley NF

A method for validating the accuracy of NMR protein structures.

一种验证 NMR 蛋白质结构准确性的方法。

• DOI: http://dx.10.1038/s41467-020-20177-1
• 发表时间: 2020
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Fowler NJ