STTR Phase I: Single molecule sequencing of phosphorylated proteins for next-generation protein analyses and diagnostics

这个小型企业技术转让(STTR)第一阶段项目的更广泛的影响/商业潜力将是开发一种高度敏感的分析方法，该方法表征用于生命科学研究应用的蛋白质修饰。蛋白质是生命的纳米机器，是几乎所有药物和绝大多数诊断测试的靶标。蛋白质的一种修饰，如添加磷酸盐分子(称为磷酸化)，是改变蛋白质活性的关键触发因素。随后，这可能会从根本上改变细胞行为，例如影响胚胎生长或肿瘤的发展。研究蛋白质及其修饰的技术的市场范围从临床应用到基础研究，估计价值170亿美元，检测这些修饰是增长最快的应用，估计每年增长18%。建议的蛋白质测序分析可以比目前的技术高4-6个数量级的灵敏度来表征这些蛋白质修饰。这种敏感性使得只有少量样本(例如活体患者的活组织切片)或目标蛋白/修饰很少的新类别的实验得以实现，并在所有情况下都转化为大量的材料节省。蛋白质及其修饰的高灵敏度表征将为工业和学术实验室的科学家提供一种新型的有价值的定量数据。这个小型企业技术转移(STTR)第一阶段项目将开发单分子蛋白质测序分析(荧光测序)，供蛋白质组学科学家使用，以精确量化蛋白质分子上的多个磷酸化位点。当今最好的分析技术，质谱仪，在识别多个紧密排列的蛋白质修饰方面具有固有的局限性，如果在特定氨基酸上修饰的蛋白质少于10%，则无法产生准确的定量数据。蛋白质组学研究人员认为，更好地表征和量化磷酸化是一种需要。该项目将探索荧光测序的竞争能力，以区分和量化多个蛋白质中紧密间隔的修饰。该项目还将提供证据，证明该技术能够检测和量化100个总蛋白质中的单个磷酸化蛋白质。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The broader impact/commercial potential of this Small Business Technology Transfer (STTR) Phase I project will be to develop a highly sensitive assay that characterizes protein modifications for life science research applications. Proteins are life's nanomachines and serve as the targets for almost all drugs and the vast majority of diagnostic tests. One type of Modifications to proteins, such as addition of phosphate molecules (termed phosphorylation), are key triggers that alter protein activity. Subsequently, this can radically change cellular behaviour, such as affecting embryonic growth or development of tumors. The market for technologies studying proteins and their modifications range from clinical applications to fundamental research and is estimated at $17 B, and detecting these modifications is the fastest-growing application growing at an estimated 18% annually. The proposed protein-sequencing assay can characterize these protein modifications with 4-6 orders of magnitude greater sensitivity than current technologies. This sensitivity enables new classes of experiments in which only small samples are available (e.g. biopsies from living patients) or the target protein/modification is rare, and translates to substantial materials savings in all cases. The highly-sensitive characterization of proteins and their modifications will provide a new type of valuable quantitative data for scientists in industry and academic labs alike. This Small Business Technology Transfer (STTR) Phase I project will be to develop the single-molecule protein sequencing assay (fluorosequencing) for use by proteomics scientists to precisely quantify multiple phosphorylated sites on protein molecules. The best analytical technology today, mass-spectrometers, has an inherent limitation in identifying multiple (2) closely spaced protein modifications and cannot produce accurate quantitative data if fewer than 10% of the proteins are modified at the particular amino acid. Better characterization and quantification of phosphorylation is recognized as a need by proteomics researchers. The project will explore the competitive ability of fluorosequencing to distinguish and quantify closely spaced modifications in multiple proteins. The project will also provide evidence for the capability of the technology to detect and quantify a single phosphorylated protein amongst 100 total proteins.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.