A low velocity molecular collider for computer-controlled biochemical reactions.

用于计算机控制生化反应的低速分子对撞机。

• 批准号: 2547065
• 金额: --
• 依托单位: University College London
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2021
• 资助国家: 英国
• 起止时间: 2021 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2547065
• 关键词: low; velocity; molecular; collider; computer

There are diverse techniques for studying biomolecular interactions e.g. proteins-ligand, proteins-nucleic acidand protein-protein interactions. On-rates, off-rates and equilibrium constants are of wide interest in industrialand fundamental research. Drug development needs to quickly isolate, identify and characterise "strongbinder" small molecules or biologics, especially in screening. Cryo-EM has energised the study of multicomponentprotein machines and the disciplines of mechanistic biochemistry are being revived. However,proteins are expensive to produce and purify but for biological realism must be used at high concentrationbut most analytical approaches create large dilutions. This creates demand for miniaturised reaction systemsthat concentrate samples, allow precise manipulation of reactions conditions and give molecularcharacterisation. Beyond determination of physical constants there is also the need to establish if one proteinmodifies another during interaction e.g. some components of complexes are enzymes and assembled (orpartly assembled) complexes may be enzymes e.g. signalling scaffold assemblies, ribosomes, coatomers,GroEL/ES chaperones etc.We will use radically new electrophoretic methods to create a platform for computer manipulation ofbiochemical interactions. With their prototype QbQ ("cubic") system GMD, the industry partner, has built aplatform that allows the generation and manipulation of the motions of ultra-sharp molecular bands in amicrofluidic channel (See Figure 1 below). By developing QbQ for protein work we will discover how to controlprotein concentrations, band composition, band width and direction of motion of bands inside a fluid channel.We will then "collide" bands with each other in the sense of crossing motion paths while characteristics ofspeed, concentration, encounter time/duration and temperature are controlled by software.QbQ has potential to complement and then surpass the most well-established analytical approaches. Itavoids steric hindrance from absorption of one binding partner to a surface (SPR) and negates mass transportlimitations (SPR and NMR). QbQ preserves the advantages of microscale thermophoresis (MST) overdynamic light scattering (DLS) and similar techniques e.g. mm long light paths and mg/ml concentrations yetsurpasses MST because dilute proteins are concentrated on the chip and the motions and interactions ofproteins can be controlled simultaneously.Ultimately this creates the possibility for logical operations and loops to control the progression of chemicalprocesses. For example, "if (sample A x sample B == Characteristic C) then react C with D Else react C withE". Having software controlled chemical reactions can create possibilities of mimicking complex, sequentialbiochemical processes in a small chip. The system can lend itself to quantitative (screening) and qualitative(analytical) investigations. We will specifically investigate protein-protein interactions in signal transductionbut also create "value branch points" at which the research progress can be captured as design principles fornew devices useful in other applications (drug screening).This project fits the BBSRC definitions used in the DTP3 application for allocation of LIDo researchstudentships, specifically "Technology Development". Signalling is a general BBSRC interest and specificallyencompassed in (i) Regenerative Biology (ii) Immunology and (iii) Stem cells aims.

研究生物分子相互作用的技术有多种，如蛋白质-配体、蛋白质-核酸和蛋白质-蛋白质相互作用。结合速率、解离速率和平衡常数在工业和基础研究中具有广泛的意义。药物开发需要快速分离、识别和鉴定“强结合剂”小分子或生物制剂，特别是在筛选中。Cryo-EM为多组分蛋白质机器的研究注入了活力，机械生物化学学科正在复兴。然而，蛋白质的生产和纯化是昂贵的，但为了生物现实主义，必须以高浓度使用，但大多数分析方法产生大的稀释。这就产生了对浓缩样品、精确控制反应条件并进行分子表征的反应系统的需求。除了确定物理常数外，还需要确定一种蛋白质是否在相互作用过程中改变另一种蛋白质，例如复合物的某些组分是酶，组装（或部分组装）的复合物可能是酶，例如信号支架组装体、核糖体、外被体、GroEL/ES分子伴侣等。我们将使用全新的电泳方法来创建一个生物化学相互作用的计算机操作平台。凭借其原型QbQ（“立方体”）系统，行业合作伙伴GMD已经建立了一个平台，可以在微流体通道中生成和操纵超尖锐分子带的运动（见下图1）。通过为蛋白质工作开发QbQ，我们将发现如何控制蛋白质浓度、条带组成、带宽和流体通道内条带的运动方向。然后，我们将在交叉运动路径的意义上使条带相互“碰撞”，而速度、浓度、遭遇时间/持续时间和温度由软件控制。QbQ有潜力补充，然后超越最好的-建立分析方法。它消除了一个结合伴侣吸附到表面（SPR）的空间位阻，并消除了质量传递限制（SPR和NMR）。QbQ保留了微尺度热泳（MST）优于动态光散射（DLS）和类似技术的优点，如mm长的光路和mg/ml浓度，但超过了MST，因为稀释的蛋白质集中在芯片上，并且蛋白质的运动和相互作用可以同时控制，这最终为逻辑操作和循环控制化学过程的进展创造了可能性。例如，“如果（样品A x样品B ==特征C），则C与D反应，否则C与E反应”。通过软件控制化学反应，可以在一个小芯片上模拟复杂的、连续的生化过程。该系统可用于定量（筛选）和定性（分析）调查。我们将专门研究信号转导中的蛋白质-蛋白质相互作用，但也创造“价值分支点”，在该点上，研究进展可以被捕获为在其他应用（药物筛选）中有用的新设备的设计原则。信号转导是BBSRC的一个普遍兴趣，特别是在（i）再生生物学（ii）免疫学和（iii）干细胞目标中。