Enzyme activity is often a key measurement to make in a biological sample

酶活性通常是生物样品中进行的关键测量

• 批准号: 2279653
• 金额: --
• 依托单位: University of Glasgow
• 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-2279653
• 关键词: Enzyme; activity; often; key; measurement

Many disease states can be indicated by the relative activity of a particular enzyme and many drugs are targeted to up- or down-regulate enzyme activity. Enzymes are now also being used beyond biomedical science, in industrial processes, being synthetically modified to target the degradation and recycling of plastics and production of fuels. Therefore, we aim to develop simple, accurate enzyme activity assays that can ultimately be (1) multiplexed to detect several different enzymes at once and (2) miniaturised into high-throughput devices. These assays are based on the modification of the surface of luminescent nanomaterials such as "quantum dots" to create enzyme sensors. For example, a peptide bearing a terminal dye can be bound to the surface of a quantum dot nanoparticle. The dye fluorescence is excited by the luminescence from the nanoparticle (via resonant energy transfer). On cleavage of the peptide by a target enzyme (a hydrolase), the dye is dissociated from the nanoparticle and the dye emission is quenched - measuring the rate of this quenching gives an indication of enzyme activity. Such nanotechnologies have been demonstrated for the sensing of several different proteases (protein cleaving enzymes), and in our work we hope to demonstrate that the technique is extensible to luminescent nanomaterials beyond quantum dots, that display new and exciting optical properties (e.g. the very sharp emission of quantum plates for high degrees of multiplexing, or upconverting lanthanide nanoparticles for sensing in highly scattering media). We will generate novel surface chemistries of these nanoparticles to produce the best sensing results. The PhD student will synthesise enzyme reactive nanoparticles that operate on the mechanism described above, starting with quantum dots and moving on to explore quantum nanoplatelets and upconverting lanthanide nanoparticles as novel alternatives. The surface chemistry will be designed to measure the activity of enzyme Matrix Metalloprotease 1 (MMP1) or human pancreatic lipase - blood biomarkers of markers of liver fibrosis and pancreatitis respectively. The student will demonstrate proof-of-concept of sensing in blood, and begin working to incorporate the nanoparticles into practical assays. This will be undertaken with collaborators in biomedical engineering (microfluidics and nanofabrication) and medicine. During the project, the student will gain practical skills in nanomaterials synthesis, modification and characterisation; and will have opportunities to apply their modified nanoparticles to a range of related projects. For example, it will be of interest to investigate (with collaborators) if the developed nanotechnology can be made compatible with high throughput screening of modified and evolved enzymes for treating plastic waste, as an alternative application. The work fits within EPSRC themes of Healthcare Technologies, Physical Sciences and Manufacturing the Future (areas include: Chemical Biology, Analytical Science, Sensors and Instrumentation). We aim to provide tools that will contribute to the ongoing paradigm shift in medicine towards stratified/personalised treatment. By enabling the rapid screening of enzyme activity in a near-patient format, diagnosis, drug selection, and dosage monitoring can be improved. During the course of this work we will also advance our understanding of nanoparticle surfaces and their interaction with biological agents such as enzymes. This new knowledge will be disseminated widely, via work with collaborators, in the scientific literature and at relevant conferences.

许多疾病状态可以通过一种特定酶的相对活性来指示，许多药物的目标是上调或下调酶的活性。酶现在也被用于生物医学科学之外的工业过程中，被合成修饰以针对塑料的降解和回收以及燃料的生产。因此，我们的目标是开发简单，准确的酶活性测定，最终可以(1)多路复用，一次检测几种不同的酶，(2)小型化成高通量设备。这些检测是基于对发光纳米材料（如“量子点”）表面的修饰来制造酶传感器。例如，带有末端染料的肽可以结合到量子点纳米颗粒的表面。染料荧光被纳米粒子的发光激发（通过共振能量转移）。在肽被目标酶（水解酶）裂解时，染料与纳米颗粒分离，染料发射被淬灭——测量这种淬灭的速率可以指示酶的活性。这种纳米技术已经被证明可以感应几种不同的蛋白酶（蛋白质切割酶），在我们的工作中，我们希望证明该技术可以扩展到量子点以外的发光纳米材料，显示出新的和令人兴奋的光学特性（例如，用于高度复用的量子板的非常尖锐的发射，或用于在高散射介质中感应的上转换镧系纳米粒子）。我们将产生这些纳米颗粒的新表面化学反应，以产生最佳的传感结果。博士生将根据上述机制合成酶反应纳米粒子，从量子点开始，继续探索量子纳米血小板和上转化镧系纳米粒子作为新的替代品。表面化学将被设计用于测量酶基质金属蛋白酶1 （MMP1）或人胰腺脂肪酶的活性-分别是肝纤维化和胰腺炎标志物的血液生物标志物。学生将演示在血液中传感的概念验证，并开始将纳米颗粒纳入实际分析中。这将与生物医学工程（微流体和纳米制造）和医学的合作者一起进行。在项目期间，学生将获得纳米材料合成、改性和表征的实用技能；并且将有机会将他们改良的纳米颗粒应用到一系列相关的项目中。例如，研究（与合作者）开发的纳米技术是否可以与用于处理塑料废物的改性和进化酶的高通量筛选相兼容，作为一种替代应用，将是一件有趣的事情。这项工作符合EPSRC医疗技术、物理科学和未来制造业的主题（领域包括：化学生物学、分析科学、传感器和仪器仪表）。我们的目标是提供工具，将有助于医学正在进行的范式转变向分层/个性化治疗。通过使酶活性在近病人的格式快速筛选，可以改善诊断，药物选择和剂量监测。在这项工作的过程中，我们也将推进我们对纳米颗粒表面及其与酶等生物制剂相互作用的理解。这一新知识将通过与合作者的合作，在科学文献和有关会议上广泛传播。