Measuring nanometre distance changes in biomolecules under pressure

测量压力下生物分子的纳米距离变化

• 批准号: 2276826
• 金额: --
• 依托单位: University of St Andrews
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2019
• 资助国家: 英国
• 起止时间: 2019 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2276826
• 关键词: Measuring; nanometre; distance; changes; biomolecules

Biomolecules, such as proteins, function by means of changes in their structural arrangement. Observing and understanding these changes can allow for detailed elucidation of biochemical mechanisms, which in turn can lead to advances in fields such as nanotechnology and drug discovery. Distances on a nanometre scale can be measured between pairs of specifically placed spin-label molecules using the pulsed EPR technique, double electron-electron resonance, DEER (PELDOR). Moving the locations of these labels within a protein can enable its structure to be mapped. As well as distances, the relative orientation of these labels can also be determined which can provide further information on the structure of the protein (G. Jeschke, Annu. Rev. Phys. Chem. (2012) 63, 419-446).While proteins typically exist in well-ordered native states under physiological conditions, their functions may cause them to move to excited states. Due to these states having higher energies, lower equilibrium populations and conformational flexibility, the excited states cannot be measured by traditional spectroscopic techniques. Interestingly, this can be overcome by subjecting the sample to high pressure, which can result in changes to the structural state of the protein and can populate the excited state (K. Akasaka, Biochemistry. (2003) 42, 10875-10885). Combining this with site-directed spin labelling and DEER spectroscopy can allow for these states to be accessed, if the sample can be rapidly frozen (M.T. Lerch et al, Proc. Natl. Acad. Sci. (2014) 111, E1201-E1210).In this work, we will apply this methodology to the messenger protein calmodulin which undergoes structural changes both with the calcium concentration as well as with the presence of binding proteins. We plan to measure at Q-band frequency and also at W-band using our home-built spectrometer, HiPER (P.A.S. Cruickshank et al, Rev. Sci. Instrum. (2009) 80, 103102). Agreed training requirements are to learn best research practice, scientific writing for publications and thesis, and to learn various research techniques. On top of these, further academic development is required in the form of QM-CDT training blocks, taught SUPA courses, attendance at the weekly colloquium, and transferrable skill courses. Further, involvement in public engagement and teaching is expected.

生物分子，如蛋白质，通过改变其结构排列来发挥作用。观察和理解这些变化可以详细阐明生物化学机制，这反过来又可以导致纳米技术和药物发现等领域的进步。纳米级的距离可以使用脉冲EPR技术，双电子-电子共振，DEER（PELDOR）测量特定放置的自旋标记分子对之间的距离。在蛋白质中移动这些标签的位置可以使其结构被映射。除了距离之外，还可以确定这些标签的相对方向，这可以提供有关蛋白质结构的进一步信息（G. Jeschke，Annu.（2012）63，419-446）。虽然蛋白质通常在生理条件下以良好有序的天然状态存在，但它们的功能可能导致它们移动到激发态。由于这些状态具有较高的能量，较低的平衡布居数和构象灵活性，激发态不能通过传统的光谱技术测量。有趣的是，这可以通过使样品经受高压来克服，这可以导致蛋白质的结构状态的变化，并且可以填充激发态（K。赤坂，生物化学。（2003）42，10875-10885）。将其与定点自旋标记和DEER光谱相结合可以允许访问这些状态，如果样品可以快速冷冻（M.T. Lerch等人，Proc.Natl. Acad. Sci.（2014）111，E1201-E1210）。在这项工作中，我们将这种方法应用于信使蛋白钙调蛋白，其随着钙浓度以及结合蛋白的存在而经历结构变化。我们计划使用我们自制的光谱仪HiPER（P.A.S.）在Q波段和W波段进行测量。Cruickshank等人，Rev. Sci.仪器。（2009）80，103102）。商定的培训要求是学习最佳研究实践，为出版物和论文撰写科学文章，并学习各种研究技术。除此之外，还需要进一步的学术发展，包括QM-CDT培训模块，教授SUPA课程，参加每周座谈会和可转移技能课程。此外，预计将参与公众参与和教学。