To Hofmeister and beyond: an improved understanding of protein solubility and stability
致 Hofmeister 及其他人士:加深对蛋白质溶解度和稳定性的了解
基本信息
- 批准号:BB/P007066/1
- 负责人:
- 金额:$ 41.07万
- 依托单位:
- 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:
- 关键词:
项目摘要
Proteins are used increasingly widely, in pharmaceutical and diagnostic formulations, cosmetics, industrial processes, and detergents. A major problem is that proteins have limited solubility and/or stability in liquid formulations, leading over time to loss of quality, reduced benefit, wasted cost (especially as the protein is usually the most expensive part), and a lack of reproducibility and predictability. Similar problems abound in other spheres: for example many proteins give poor analytical spectra, cloudiness or haze in beverages, and cannot be crystallised, because of difficulties in keeping them in solution. There is thus a major need to improve stability without losing solubility or vice versa. The main approach used is to add high concentrations of ions and other molecules to the protein solution (known generally as excipients). Currently the selection of excipients is usually tackled on an ad hoc basis, using high-throughput testing of different solution conditions, and there is very little theoretical rationale that could be used as a basis for a more rational, cheaper and quicker route to improved performance. This proposal aims to provide a more rational basis for understanding and improving stability and solubility.We recently proposed a model that explains how salts added to a protein solution affect its solubility and stability, by competing with the protein for water molecules. Different salts compete better or worse than the protein, and therefore have different effects. The effects have been known for over 100 years and are generally called the Hofmeister effect, but there remains considerable disagreement over the explanation. This matters, because once we understand the physical basis for the Hofmeister effect, we can exploit it to develop better solution conditions. Our model was based largely on measurements using a spectroscopic technique known as NMR.The model differs in important ways from the current standard models. We will therefore start by using NMR to compare our model to the most popular of these, to eliminate the most significant likely objection to our model and clear the ground. Our model emphasises the role of water molecules in mediating changes in solubility and stability: in particular, it predicts that ions that stabilise proteins necessarily make them less soluble, and vice versa. If this is strictly true, then it places severe limits on what we can do to (for example) improve stability without compromising solubility. We shall therefore explore ways of getting round the problem. First, we shall investigate whether mixtures of ions behave simply in an additive way. To the extent that they do not, we might be able to exploit a window of opportunity. A particularly interesting pair of ions is a mixture of the two amino acids arginine and glutamate, which seem to work in a different way and are apparently 'special'. We shall investigate how they work, whether they are in fact different, and whether combining them with more typical Hofmeister ions helps. We will also investigate our suspicion that increasing the size of a protein makes the effect of added ions weaker: if true, this is useful information, since it guides the way we would go about stabilising different proteins.There are many organisms that can grow in high salt concentrations. Normally, high salt reduces either solubility or stability or both: so how do these organisms manage to grow successfully? The answer is that they produce high concentrations of specific small molecules to balance the high external ionic strength. We shall investigate how these molecules work, and whether they behave in the same way as 'normal' ions. If they work in different ways, then these can be exploited.Finally we will study whether external pressure can be used as an alternative way to stabilise proteins, and thus whether pressure would be a useful variable. The outcome will be a rational toolbox to guide solution conditions.
蛋白质在制药和诊断配方、化妆品、工业过程和洗涤剂中的应用越来越广泛。一个主要问题是,蛋白质在液体制剂中的溶解度和/或稳定性有限,随着时间的推移,会导致质量下降、效益降低、成本浪费(特别是因为蛋白质通常是最昂贵的部分),以及缺乏重复性和可预测性。类似的问题在其他领域也比比皆是:例如,许多蛋白质的分析光谱很差,饮料中出现混浊或混浊,由于难以将其保持在溶液中,因此无法结晶。因此,主要需要在不损失溶解度的情况下提高稳定性,反之亦然。使用的主要方法是在蛋白质溶液中添加高浓度的离子和其他分子(通常称为赋形剂)。目前,辅料的选择通常是在特别的基础上进行的,使用不同溶液条件的高通量测试,几乎没有理论基础可以用作更合理、更便宜和更快地提高性能的基础。这一建议旨在为理解和改善稳定性和溶解性提供更合理的基础。我们最近提出了一个模型,解释了添加到蛋白质溶液中的盐是如何通过与蛋白质竞争水分子来影响其溶解性和稳定性的。不同的盐比蛋白质竞争更好或更差,因此有不同的影响。这种效应早在100多年前就已为人所知,通常被称为霍夫迈斯特效应,但对其解释仍存在相当大的分歧。这一点很重要,因为一旦我们了解了霍夫迈斯特效应的物理基础,我们就可以利用它来开发更好的解决条件。我们的模型在很大程度上是基于一种被称为NMR的光谱技术进行的测量。该模型与当前的标准模型有重要的不同。因此,我们将首先使用核磁共振将我们的模型与其中最流行的模型进行比较,以消除对我们的模型最重要的反对意见,并清除基础。我们的模型强调了水分子在调节溶解度和稳定性变化中的作用:特别是它预测,稳定蛋白质的离子必然会使蛋白质更难溶解,反之亦然。如果严格来说这是真的,那么它对我们可以做什么来(例如)提高稳定性而不损害溶解性施加了严格的限制。因此,我们将探索绕过这个问题的方法。首先,我们将研究离子混合物是否表现为简单的相加行为。如果他们不这么做,我们或许能够利用一扇机会之窗。一对特别有趣的离子是两种氨基酸精氨酸和谷氨酸的混合物,这两种氨基酸似乎以不同的方式工作,显然是“特殊的”。我们将研究它们是如何工作的,它们是否实际上是不同的,以及将它们与更典型的霍夫迈斯特离子结合是否有帮助。我们还将调查我们的怀疑,即增加蛋白质的大小会削弱添加离子的影响:如果是真的,这是有用的信息,因为它指导我们稳定不同的蛋白质。有许多生物可以在高盐浓度下生长。通常情况下,高盐会降低溶解度或稳定性,或者两者兼而有之:那么这些生物是如何成功生长的呢?答案是,它们会产生高浓度的特定小分子,以平衡高外部离子强度。我们将研究这些分子是如何工作的,以及它们的行为方式是否与“正常”离子相同。如果它们以不同的方式工作,那么这些都可以被利用。最后,我们将研究外部压力是否可以作为稳定蛋白质的替代方法,从而研究压力是否会是一个有用的变量。结果将是一个指导解决方案条件的Rational工具箱。
项目成果
期刊论文数量(3)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Allomorphy as a mechanism of post-translational control of enzyme activity.
- DOI:10.1038/s41467-020-19215-9
- 发表时间:2020-11-02
- 期刊:
- 影响因子:16.6
- 作者:Wood HP;Cruz-Navarrete FA;Baxter NJ;Trevitt CR;Robertson AJ;Dix SR;Hounslow AM;Cliff MJ;Waltho JP
- 通讯作者:Waltho JP
Enzymatic production of ß-glucose 1,6-bisphosphate through manipulation of catalytic magnesium coordination
通过操纵催化镁配位酶法生产β-葡萄糖1,6-二磷酸
- DOI:10.1039/d0gc03290e
- 发表时间:2021
- 期刊:
- 影响因子:9.8
- 作者:Wood H
- 通讯作者:Wood H
CORM-3 induces DNA damage through Ru(II) binding to DNA.
- DOI:10.1042/bcj20220254
- 发表时间:2022-07-15
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
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Michael Williamson其他文献
Validation of a digit symbol substitution test for use in supervised and unsupervised assessment in mild Alzheimer’s disease
验证数字符号替换测试用于轻度阿尔茨海默病的监督和非监督评估
- DOI:
- 发表时间:
2022 - 期刊:
- 影响因子:2.2
- 作者:
Michael Williamson;P. Maruff;A. Schembri;Hannah Cummins;Laura J. Bird;E. Rosenich;Y. Lim - 通讯作者:
Y. Lim
Pubic symphysis diastasis sustained from a waterslide injury
- DOI:
10.1016/j.jcot.2018.01.002 - 发表时间:
2018-06-01 - 期刊:
- 影响因子:
- 作者:
Michael Williamson;Felice Vanacore;Caroline Hing - 通讯作者:
Caroline Hing
Intrapulmonary lymph nodes masquerading as renal cell carcinoma metastases
- DOI:
10.1016/s0090-4295(99)80202-7 - 发表时间:
1995-08-01 - 期刊:
- 影响因子:
- 作者:
Robert C. Kolosseus;Roy T. Temes;Richard M. Feddersen;Michael Williamson;Anthony Y. Smith - 通讯作者:
Anthony Y. Smith
Reduction in abdominal symptoms (CFAbd-Score), faecal M2-pyruvate-kinase and Calprotectin over one year of treatment with Elexacaftor-Tezacaftor-Ivacaftor in people with CF aged >=12 years - The RECOVER study
12 岁以上 CF 患者接受 Elexacaftor-Tezacaftor-Ivacaftor 治疗一年后,腹部症状 (CFAbd-Score)、粪便 M2-丙酮酸激酶和钙卫蛋白减少 - RECOVER 研究
- DOI:
10.1101/2023.07.10.23292435 - 发表时间:
2023 - 期刊:
- 影响因子:3.1
- 作者:
J. Mainz;Karen Lester;B. Elnazir;Michael Williamson;E. McKone;Des W Cox;Barry;Linnane;C. Zagoya;Franziska Duckstein;A. Barucha;Jane C Davies;Paul McNally - 通讯作者:
Paul McNally
Centrifuge modelling of the effect of base slab stiffness on long-term heave and swell pressure
底板刚度对长期隆起和膨胀压力影响的离心模型试验
- DOI:
10.1680/jgeot.22.00111 - 发表时间:
2022-08-17 - 期刊:
- 影响因子:5.200
- 作者:
Deryck Chan;Gopal Madabhushi;Giulia Viggiani;Michael Williamson;Yu Sheng Hsu - 通讯作者:
Yu Sheng Hsu
Michael Williamson的其他文献
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{{ truncateString('Michael Williamson', 18)}}的其他基金
A World-Leading National Network for NMR in the Physical and Life Science: Very-High Field Infrastructure at Sheffield
世界领先的物理和生命科学核磁共振国家网络:谢菲尔德的超高现场基础设施
- 批准号:
EP/S01358X/1 - 财政年份:2018
- 资助金额:
$ 41.07万 - 项目类别:
Research Grant
Validation of NMR protein structures using FIRST and RCI
使用 FIRST 和 RCI 验证 NMR 蛋白质结构
- 批准号:
BB/P020038/1 - 财政年份:2018
- 资助金额:
$ 41.07万 - 项目类别:
Research Grant
Upgrade to 600 MHz NMR spectrometer
升级至 600 MHz NMR 波谱仪
- 批准号:
BB/R000727/1 - 财政年份:2017
- 资助金额:
$ 41.07万 - 项目类别:
Research Grant
Internal dynamics in the enzyme barnase
芽孢杆菌RNA酶的内部动力学
- 批准号:
BB/J014966/1 - 财政年份:2012
- 资助金额:
$ 41.07万 - 项目类别:
Research Grant
Investigation of alternative states of barnase
芽孢杆菌RNA酶替代状态的研究
- 批准号:
BB/D015308/1 - 财政年份:2006
- 资助金额:
$ 41.07万 - 项目类别:
Research Grant
Remotely Operated Seafloor Drill with Extended Coring Depth Capability
具有扩展取芯深度功能的远程操作海底钻机
- 批准号:
9403812 - 财政年份:1995
- 资助金额:
$ 41.07万 - 项目类别:
Standard Grant
Feasibility Assesssment of a Deep Ocean Rock Coring Drill
深海岩石取芯钻机的可行性评估
- 批准号:
8361067 - 财政年份:1984
- 资助金额:
$ 41.07万 - 项目类别:
Standard Grant
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