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Structural Dynamics in LOV Domain Photosensor Proteins

LOV 结构域光传感器蛋白的结构动力学

基本信息

批准号：
EP/N033647/1
负责人：
Stephen Meech
金额：
$ 44.97万
依托单位：
University of East Anglia
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2016
资助国家：
英国
起止时间：
2016 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=EP%2FN033647%2F1
关键词：
Structural Dynamics LOV Domain Photosensor

项目摘要

The oxygen we breathe and the food we eat ultimately derive from photosynthesis, the conversion of the sun's rays into useful chemical energy by plants and bacteria. However, we can have too much sunshine. Just as humans can suffer from skin cancer due to harmful UV rays in the sun, so plants and bacteria can be damaged by too much sunlight. As a result of these conflicting demands it is essential for a wide range of living organisms to have some means of sensing light levels. That plants have such tools is obvious to anyone who has ever grown cress on a windowsill and seen it turn towards the light. What we are principally concerned with in this project is precisely how plants and bacteria sense light, and whether this process can be exploited in human applications. In this proposal we focus on one particularly useful family of photosensor proteins, the LOV (Light-Oxygen-Voltage) domains.Over the past twenty years many proteins have been discovered which detect light. The LOV domain proteins are part of a much larger group called the flavoproteins. 'Flavo-' means yellow indicating that these proteins are colored and thus have the ability to absorb light energy. In the photoactive flavoproteins, which includes the LOV domains, this energy is converted it into some useful structure change in the protein. This then stimulates further changes in associated proteins which ultimately gives rise to a specific biological response. This complex chain of events in known to be important in: determining when flowers open; making leaves turn towards the sun; causing bacteria to swim away from harmful sunlight; controlling circadian rhythms, etc. In a few cases the structures of these LOV domain proteins have been determined, and other experiments have shown what secondary proteins (or DNA) they are complexed with, which informs us about their function. However, very little is known about the mechanism of operation of photoactive flavoproteins, beyond the fact that the proteins binds a flavin molecule which absorbs blue light. The question at the heart of our research is how is the event of light absorption can be converted into a specific structure change which acts as a signal to initiate other processes in living cells.In this work we will use some of the most sophisticated methods of laser spectroscopy to record what happens to the proteins after they have absorbed light. It is through the application of such advanced physical methods to living systems that we can begin to understand (and even control) the chemistry of life. In this case we will stimulate the protein response with a short pulse of blue light (less than 100 million billionths of a second long) and use another short pulse of light to take ultrafast 'snapshots' of the structural changes as they happen. We will follow these structure changes right from the time of excitation all the way through to formation of the final signalling state. By thus observing protein function in real time we will obtain new insights into the mechanism of how plants 'see' light. We will then use some tricks of protein chemistry to test, probe and manipulate these structure changes. Our interest in these proteins is not simply curiosity as to how they work. Recently scientists have artificially incorporated light-activated proteins into various cells and then used light to trigger a particular response. The most famous example is the use of light to activate the firing of neurons in the brains of mice, but as other light-activated proteins (such as LOV domains) become better understood it will become possible to stimulate a variety of new phenomena. The ability to stimulate a specific process in a living cell with both time and space resolution will represent a powerful new tool for scientists trying to understand cellular functions, and will inform a variety of research in health sciences.

我们呼吸的氧气和我们吃的食物最终来自光合作用，即植物和细菌将太阳光转化为有用的化学能。然而，我们可以有太多的阳光。正如人类会因阳光下有害的紫外线而患上皮肤癌一样，植物和细菌也会因过多的阳光而受损。由于这些相互冲突的需求，对范围广泛的生物来说，拥有一些感知光线水平的手段是必不可少的。对于任何在窗台上种植水芹并看到它转向阳光的人来说，植物都有这样的工具是显而易见的。我们在这个项目中主要关心的正是植物和细菌如何感知光线，以及这一过程是否可以用于人类应用。在这项研究中，我们关注一个特别有用的光传感器蛋白家族，LOV(光-氧-电压)结构域。在过去的二十年里，许多检测光的蛋白质已经被发现。LOV结构域蛋白是一个更大的称为黄素蛋白的组的一部分。‘Flavo-’的意思是黄色，表示这些蛋白质是有色的，因此具有吸收光能的能力。在包括LOV结构域的光活性黄素蛋白中，这种能量被转化为蛋白质中一些有用的结构变化。然后刺激相关蛋白质的进一步变化，最终引起特定的生物反应。已知这一复杂的事件链在以下方面很重要：确定花何时开放；使树叶转向太阳；使细菌游过有害的阳光；控制昼夜节律等。在少数情况下，这些LOV结构域蛋白质的结构已经确定，其他实验表明它们与哪些次级蛋白质(或DNA)结合，这告诉我们它们的功能。然而，人们对光活性黄素蛋白的作用机制知之甚少，除了这些蛋白与吸收蓝光的黄素分子结合这一事实之外。我们研究的核心问题是如何将光吸收事件转化为特定的结构变化，作为启动活细胞中其他过程的信号。在这项工作中，我们将使用一些最复杂的激光光谱方法来记录蛋白质吸收光后发生的情况。正是通过将这种先进的物理方法应用于生命系统，我们才能开始了解(甚至控制)生命的化学。在这种情况下，我们将用一个短的蓝光脉冲(不到亿分之一秒的长度)来刺激蛋白质的反应，并使用另一个短的光脉冲来拍摄结构变化发生时的超快快照。我们将跟踪这些结构的变化，从激励时间一直到最终信令状态的形成。通过实时观察蛋白质的功能，我们将对植物如何“看到”光的机制获得新的见解。然后，我们将使用蛋白质化学的一些技巧来测试、探测和操纵这些结构变化。我们对这些蛋白质的兴趣不仅仅是好奇它们是如何工作的。最近，科学家们人工将光激活蛋白整合到各种细胞中，然后利用光来触发特定的反应。最著名的例子是使用光来激活小鼠大脑中神经元的激活，但随着对其他光激活蛋白(如LOV结构域)的更好理解，它将有可能刺激各种新现象。对于试图了解细胞功能的科学家来说，以时间和空间分辨率刺激活细胞中特定过程的能力将是一个强大的新工具，并将为健康科学的各种研究提供信息。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Infrared spectroscopy reveals multi-step multi-timescale photoactivation in the photoconvertible protein archetype dronpa.

DOI：
10.1038/s41557-018-0073-0
发表时间：
2018-08
期刊：
Nature chemistry
影响因子：
21.8
作者：
Laptenok SP;Gil AA;Hall CR;Lukacs A;Iuliano JN;Jones GA;Greetham GM;Donaldson P;Miyawaki A;Tonge PJ;Meech SR
通讯作者：
Meech SR

Excited State Vibrations of Isotopically Labeled FMN Free and Bound to a Light-Oxygen-Voltage (LOV) Protein.

DOI：
10.1021/acs.jpcb.0c04943
发表时间：
2020-08-20
期刊：
The journal of physical chemistry. B
影响因子：
0
作者：
Iuliano JN;Hall CR;Green D;Jones GA;Lukacs A;Illarionov B;Bacher A;Fischer M;French JB;Tonge PJ;Meech SR
通讯作者：
Meech SR

Femtosecond stimulated Raman study of the photoactive flavoprotein AppABLUF

DOI：
10.1016/j.cplett.2017.03.030
发表时间：
2017-09-01
期刊：
CHEMICAL PHYSICS LETTERS
影响因子：
2.8
作者：
Hall, Christopher R.;Heisler, Ismael A.;Meech, Stephen R.
通讯作者：
Meech, Stephen R.

Femtosecond to Millisecond Dynamics of Light Induced Allostery in the Avena sativa LOV Domain.

DOI：
10.1021/acs.jpcb.7b00088
发表时间：
2017-02-09
期刊：
The journal of physical chemistry. B
影响因子：
0
作者：
Gil AA;Laptenok SP;French JB;Iuliano JN;Lukacs A;Hall CR;Sazanovich IV;Greetham GM;Bacher A;Illarionov B;Fischer M;Tonge PJ;Meech SR
通讯作者：
Meech SR

Functional dynamics of a single tryptophan residue in a BLUF protein revealed by fluorescence spectroscopy