Photoelectron spectroscopy in a liquid microjet: unravelling the excited state dynamics of photoactive proteins

液体微射流中的光电子能谱：揭示光活性蛋白质的激发态动力学

• 批准号: EP/L005697/2
• 负责人: Graham Worth
• 金额: $ 12.92万
• 依托单位: University College London
• 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%2FL005697%2F2
• 关键词: Photoelectron; spectroscopy; liquid; microjet; unravelling

The extensive use of efficient light-induced processes in nature is inspiring efforts to exploit similar processes in functional synthetic systems. For example, fluorescent proteins have revolutionalised biological imaging. However, our understanding of the crucial role of the protein surrounding the chromophore in photoactive proteins is still far from complete. The aim of this research is to gain a molecular level understanding of the interactions between photoactive protein chromophores and their environment by a systematic investigation of the electronic structure and excited state dynamics of a series of chromophores in vacuo, in solution and in protein. Photoelectron spectroscopy is a particularly valuable tool for measuring the binding energies of electrons in molecules and femtosecond time-resolved photoelectron spectroscopy has emerged as a very powerful technique for probing the flow of energy in a molecule following photoexcitation. In femtosecond time-resolved photoelectron spectroscopy, a femtosecond laser pulse (pump) excites a molecule and after some delay a second femtosecond laser pulse (probe) ionises the molecule. The kinetic energy and angular distribution of the resulting photoelectrons provides information about the electronic and vibrational states of the molecule at the time of ionisation. Recording a series of photoelectron spectra at different pump-probe delays allows us to record a molecular "movie" of the flow of electronic and vibrational energy in the molecule. Time-resolved photoelectron spectroscopy has proved remarkably successful for investigating electronic structure and dynamics in the gas phase and in solids but aqueous solutions have presented more of a challenge. Recent technical advances in liquid microjet technology have enabled time-resolved photoelectron spectroscopy in solutions to become a real and exciting possibility. We will exploit these developments to create a photoelectron spectroscopy apparatus that is capable of investigating the femtosecond dynamics of chromophores in solution and in their protein environments. In a biological system, the molecular dynamics after photoexcitation are controlled by the molecular and electronic structure of the chromophore and by its interaction with the environment. For example, the environment of the GFP chromophore defines its optical properties: the chromophore is strongly fluorescent inside its barrel-shaped protein, while the fuorescence is lost when the protein is denatured but it returns upon renaturation; the isolated chromophore is non-fluorescent in aqueous solution and it is also non-fluorescent in the gas phase, yet the absorption spectrum of the isolated molecule in the gas phase is remarkably similar to that in the protein. In order to unravel the important role of the environment in defining the optical properties of fluorescent proteins, we will investigate how systematic changes to the electronic and structural properties of the chromophore and mutations to the protein influence binding energies and electronic relaxation following photoexcitation. High-level electronic structure and dynamics calculations will assist the interpretation of the experimental results. Organic chemistry and molecular biology methods will be employed to create the series of chromophores and proteins for systematic evaluation of the influence of electronic, structural, and environmental changes. The multidisciplinary team that has been assembled is ideally suited to tackle this important problem which will have an impact in many areas of science.

在自然界中广泛使用有效的光诱导过程是鼓舞人心的努力，利用功能合成系统中的类似过程。例如，荧光蛋白已经彻底改变了生物成像。然而，我们对光活性蛋白质中生色团周围蛋白质的关键作用的理解还远远没有完成。本研究的目的是获得一个分子水平的理解之间的相互作用的光活性蛋白质发色团和它们的环境通过一系列的发色团在真空中，在溶液中和蛋白质中的电子结构和激发态动力学的系统调查。光电子能谱是测量分子中电子结合能的一种特别有价值的工具，飞秒时间分辨光电子能谱已成为探测光激发后分子中能量流动的一种非常强大的技术。在飞秒时间分辨光电子能谱中，飞秒激光脉冲（泵浦）激发分子，并且在一些延迟之后，第二飞秒激光脉冲（探测）电离分子。所产生的光电子的动能和角分布提供了关于电离时分子的电子和振动状态的信息。在不同的泵浦-探测延迟下记录一系列光电子谱，使我们能够记录分子中电子和振动能量流动的分子“电影”。时间分辨光电子能谱已被证明是非常成功的研究电子结构和动力学在气相和固体，但水溶液中提出了更多的挑战。液体微射流技术的最新技术进展使溶液中的时间分辨光电子能谱成为一种真实的和令人兴奋的可能性。我们将利用这些发展，以创建一个光电子能谱仪，能够调查的飞秒动力学的发色团在溶液中，并在其蛋白质的环境。在生物系统中，光激发后的分子动力学受发色团的分子和电子结构及其与环境的相互作用控制。例如，GFP发色团的环境决定了它的光学性质：发色团在其桶形蛋白质内是强荧光的，而当蛋白质变性时荧光消失，但在复性时又恢复;分离的发色团在水溶液中不发荧光并且在气相中也不发荧光，然而，气相中分离分子的吸收光谱与蛋白质中的吸收光谱非常相似。为了解开的重要作用的环境中定义的荧光蛋白的光学性质，我们将研究如何系统的变化的电子和结构性质的发色团和突变的蛋白质影响结合能和电子弛豫光激发后。高层次的电子结构和动力学计算将有助于解释实验结果。有机化学和分子生物学的方法将被用来创建一系列的生色团和蛋白质的电子，结构和环境变化的影响进行系统的评估。已经组建的多学科团队非常适合解决这一重要问题，这将对许多科学领域产生影响。

项目成果

Photoelectron spectroscopy of isolated luciferin and infraluciferin anions in vacuo: competing photodetachment, photofragmentation and internal conversion.

• DOI: 10.1039/c7cp04815g
• 发表时间: 2017-08
• 期刊: Physical chemistry chemical physics : PCCP
• 作者: J. Woodhouse;Mariana Assmann;M. Parkes;H. Grounds;Steven J Pacman;James C. Anderson;G. Worth;H. Fielding

Shining light on the electronic structure and relaxation dynamics of the isolated oxyluciferin anion.

• DOI: 10.1039/d0cp03276j
• 发表时间: 2020-08
• 期刊: Physical chemistry chemical physics : PCCP
• 作者: A. Patel;Alice Henley;M. Parkes;Mariana Assmann;G. Worth;James C. Anderson;H. Fielding