DISRUPTIVE TECHNOLOGY FOR KINETIC STUDIES OF PROTEIN-SMALL MOLECULE BINDING

蛋白质-小分子可逆结合的动力学研究对于理解疾病的分子机制、分子诊断和药物先导物的开发至关重要。基于生物传感器的方法在此类研究中最为成熟，但它们需要将蛋白质或小分子固定在传感器上。固定化常常阻碍结合。在这里，我们提出的设备，将独特地使固定化自由蛋白质-小分子结合的动力学研究。 在3年的技术开发阶段，我们的2个实验室（37名HQP和约80万美元/年的运营资金）将使用拟议的仪器。在此阶段，我们将：（i）组装用于无固定动力学研究的独特仪器，（ii）开发用于快速和准确的信号处理和数据分析的算法，（iii）使用该仪器研究来自Yudin实验室和GSK产生的文库的小分子的结合动力学，（工业合作者），（iv）设计仪器和软件的商业版本，（v）启动技术商业化，以及（vi）设计并实施HQP使用该工具的培训方案。 在技术开发阶段之后，仪器将被放置在约克生物分析中心（YBioCore）。然后，来自约克10个实验室的经过培训的HQP将使用它进行相关研究。外部用户也可以访问。Krylov的实验室将每年为HQP举办为期3天的研讨会。讲习班将包括课堂和实践练习。仪器将由设施技术人员维护，但由经过培训的HQP操作。具有这些技能的HQP是制药和生物技术行业非常需要的。 在YBioCore中，该仪器将被研究疾病分子机制的生物学家和开发诊断探针和药物先导的化学家使用。该仪器的独特功能将有助于获得动力学数据，如果没有该仪器，这些数据将很难或不可能获得。利用该仪器获得的数据将有助于开发分子诊断和药物先导，这反过来将使加拿大人受益。 如果我们不能在2017年建立拟议的仪器，我们实验室的两个重要方向将几乎停滞。在Krylov的实验室，它是基于LSTDLPF的技术的开发，目前由3个PDF和2个格拉德生进行。在Yudin的实验室，这是大环蛋白结合研究，目前由4名格拉德生运行。因此，延迟获得仪器将大大延迟6名格拉德生完成学位要求和3名PDF的职业培训。延迟获得该仪器也将产生更广泛的影响，因为它也将延迟将我们的高性能技术转移到广泛的学术和工业最终用户社区，该技术正在为他们开发。

Kinetic studies of reversible protein-small molecule binding are pivotal to understanding molecular mechanisms of diseases and development of molecular diagnostics and drug leads. Biosensor-based methods are most established in such studies, but they require the immobilization of either protein or small molecule on the sensor. The immobilization often blocks the binding. Here we propose equipment that will uniquely enable immobilization-free kinetic studies of protein-small molecule binding. The proposed instrument will be used by our 2 labs (37 HQP and ~$0.8M/year operating funds) during a 3-year technology-development stage. During this stage, we will: (i) assemble a unique instrument for immobilization-free kinetic studies, (ii) develop algorithms for fast and accurate signal processing and data analysis, (iii) use the instrument for studying binding kinetics for small molecules from libraries generated by Yudin’s lab and GSK (industrial collaborator), (iv) design commercial versions of the instrument and software, (v) initiate commercialization of the technology, and (vi) design and implement a training protocol for use of the instrument by HQP. After the technology-development stage, the instrument will be placed in the York Bioanalytical Core Facility (YBioCore). It will then be used by trained HQP from 10 labs carrying out relevant research at York. It will also be accessible to external users. Krylov’s lab will hold annual 3-day workshops for HQP. The workshops will include classes as well as hands-on exercises. The instrument will be maintained by Facility technical staff but operated by trained HQP. HQP with such skills are highly desired by the pharma and biotech industries. When in YBioCore, the instrument will be used by biologists studying molecular mechanisms of diseases and chemists developing diagnostic probes and drug leads. The unique capabilities of the instrument will facilitate obtaining kinetic data that would be difficult or impossible without this instrument. The data obtained with the instrument will enable the development of molecular diagnostics and drug leads which, in turn, will benefit Canadians. If we are not able to build the proposed instrument in 2017, 2 important directions in our labs will be practically stalled. In Krylov’s lab it is the development of the LSTDLPF-based technology, which is currently carried out by 3 PDFs and 2 grad students. In Yudin’s lab, it is macrocycle-protein binding studies, which are currently run by 4 grad students. A delay in acquiring the instrument will, thus, significantly delay the completion of degree requirements by 6 grad students and the career training of 3 PDFs. A delay in obtaining the instrument will also have a much wider impact as it will also delay the transfer of our highly-enabling technology to a broad community of academic and industrial end-users for whom this technology is being developed.