A G-protein-coupled receptor-based sensor design platform for high throughput drug screening

提案摘要 G蛋白偶联受体（GPCR）是最大的跨膜受体家族，包括超过800个 人体内的受体类型。GPCR在各种生理过程中起着至关重要的作用。因此，委员会认为， GPCR是FDA批准的所有药物中34%的目标。尽管有如此多的人， 一半的GPCR仍然是FDA批准的药物的非靶向药物。这部分是由于缺乏一个具有成本效益， 用于监测GPCR后即时信号事件的高通量药物筛选平台 activation.为了进一步探索FDA药物靶向和非靶向GPCR的治疗潜力， 因此，有必要开发一种新的具有高信噪比的高通量GPCR药物筛选平台。 基于GPCR的荧光传感器对于高通量药物筛选是有利的，因为它们不 需要昂贵的试剂或底物，从而能够以低成本筛选数百种化合物。我的目标 设计基于GPCR的荧光传感器，可用于筛选G α i/o偶联和G α s-的药物。 偶联GPCR。为了设计和优化这个平台，我将设计κ阿片受体的传感器， 阿片受体、β-2-肾上腺素能受体和多巴胺D1受体。我会用一个合理的 蛋白质设计和定向进化来优化这些传感器。然后，我会给你做一个药物证明 与密歇根大学化学基因组学中心合作， 密歇根我预计这个平台比目前的GPCR配体药物筛选成本更低，更容易使用 平台该提案的完成将使工程的传感器广泛的高 高通量GPCR药物筛选这种新的荧光传感器设计平台将能够筛选新颖的 用于治疗目的的GPCR药物。

PROPOSAL SUMMARY G-protein-coupled receptors (GPCRs) are the largest family of transmembrane receptors, comprising over 800 receptor types in the human body. GPCRs play crucial roles in various physiological processes. Consequently, GPCRs are the target of 34% of all FDA approved pharmaceuticals. Despite this significant number, more than half of GPCRs remain untargeted by FDA approved drugs. This is partially due to the lack of a cost-effective, high-throughput drug screening platform for monitoring the immediate signaling events following GPCR activation. To further explore the therapeutic potential of both the FDA drug-targeted and untargeted GPCRs, it is necessary to develop a new high-throughput GPCR drug screening platform with a high signal-to-noise. GPCR-based fluorescent sensors are advantageous for high-throughput drug screening because they do not require expensive reagents or substrates, enabling the screening of hundreds of compounds at low costs. I aim to engineer GPCR-based fluorescent sensors that can be used to screen drugs for both Gαi/o-coupled and Gαs- coupled GPCRs. To design and optimize this platform, I will engineer sensors for the kappa opioid receptor, mu opioid receptor, beta-2-adrenergic receptor, and the dopamine D1 receptor. I will use a combination of rational protein design and directed evolution to optimize these sensors. Then, I will perform a proof of principle drug screening with these sensors in collaboration with the Center of Chemical Genomics at the University of Michigan. I expect this platform to cost less and be easier to use than the current GPCR ligand drug screening platforms. The completion of this proposal will allow the engineering of a wide array of sensors for high throughput GPCR drug screening. This new fluorescent sensor design platform will enable the screening of novel drugs for GPCRs that can be used for therapeutic purposes.