Multi-Wavelength Fluorescence Radical Dosimetry for Real-Time Assessment of Protein Footprinting Radical Yield

名为“多波长荧光”的GenNext第一阶段SBIR产品 用于实时评估蛋白质足迹自由基产率的自由基剂量学 满足对更高订单的新工具和改进工具的已知需求 生物药物的结构分析(HOS)和膜蛋白靶标研究。 一种新兴的HOS分析技术是羟基自由基足迹法(HRPF)。HRPF 涉及通过与羟基自由基反应对蛋白质外部进行不可逆标记 用随后的MS分析来鉴定蛋白质的外部部分。我们有 开发了执行HRPF的商业解决方案。最近一个新的和有价值的 依赖于OH生成的三氟甲基(TFM)自由基的足迹技术 氟化钠水溶液的自由基攻击已被开发出来，显示出巨大的前景 当与HRPF结合时。TFM蛋白足迹(TFMPF)具有很强的互补性 对于HRPF，因为TFMPF有效地标记了相对于 哦，激进的攻击。当HRPF和TFMPF一起使用时，可以提供大量的覆盖范围 以及溶剂可及残留物的检测，因此代表着一种变革性的 改进生物制药HOS评估。 TFMPF的实践是由Michael Gross教授开创的 华盛顿大学，密苏里州圣路易斯同时表现出解决未实现的问题的巨大承诺 制药研究中的挑战，TFMPF的重复性受到以下挑战 背景清除的可变性。与格罗斯实验室合作，我们的工作 将我们创新的HRPF自由基剂量学技术扩展到TFMPF。下一代 TECHICES是唯一一家将HRPF HOS分析产品商业化的公司。 我们的目标是将HRPF和TFMPF的组合使用从 将学术研究实验转化为有价值的分析工具。一旦简化， 转变为一种健壮的技术，我们设想将HRPF和 TFMPF使单抗生物药物与以下物质的相互作用成为可能 它们的膜靶标；阐明铅与正位或正构体结合的动力学 变构膜靶标；揭示次级信使信号级联 GPCR先导化合物；并检测正构/变构反 肿瘤对酶和生长因子等靶点的作用。

The GenNext Phase I SBIR submssion entitled “Multi-Wavelength Fluorescence Radical Dosimetry for Real-Time Assessment of Protein Footprinting Radical Yield” is responsive to the ackowledged need for new and improved tools for higher order structural analysis (HOS) of biopharmaceuticals and membrane protein target studies. An emerging HOS analysis technique is hydroxyl radical foot-printing (HRPF). HRPF involves the irreversible labeling of a protein’s exterior by reaction with hydroxyl radicals with subsequent MS analysis to identify the outer portions of the protein. We have developed commercial solutions to perform HRPF. Recently a new and valuable footprinting technique that relies upon trifluoromethyl (TFM) radicals created by OH radical attack of aqueous sodium triflinate has been developed that shows great promise when combined with HRPF. TFM protein footprinting (TFMPF) is highly complementary to HRPF, as TFMPF effectively labels amino acid residues that are relatively “silent” to OH radical attack. When used together, HRPF and TFMPF provide substantial coverage and detection of solvent accessible residues, and as such represent a transformative improvement to biopharmaceutical HOS assessment. The practice of TFMPF has been pioneered by Professor Michael Gross of Washington University, St. Louis, Mo. While showing great promise to address unmet challenges in pharmaceutical research, reproducibility for TFMPF is challenged by variability of background scavenging. Collaborating with the Gross laboratory, our work will extend our innovative HRPF radical dosimetry technology to TFMPF. GenNext Technologies is the only company commercializing products for HRPF HOS analysis. Our goal is to convert the combined use of HRPF and TFMPF process from an academic research experiment into a valuable analytical tool. Once simplified and transformed into a robust technique, we envision the facile combination of HRPF and TFMPF to enable: paratope and epitope the interaction of mAb biopharmaceuticals with their membrane targets; elucidate the dynamics of lead binding to orthosteric or allosteric membrane targets; to reveal secondary messenger signaling cascades of GPCR lead compounds; and to detect the impact of orthosteric / allosteric anti- neoplastics upon targets such as kinases and growth factors.