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

用于实时评估蛋白质足迹自由基产量的多波长荧光自由基剂量测定

基本信息

批准号：
10250755
负责人：
Scot Randy Weinberger
金额：
$ 25.14万
依托单位：
GENNEXT TECHNOLOGIES, INC.
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-05-01 至 2023-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10250755
关键词：
Address Adopted Adoption Amino Acids Antibodies Antibody Binding Sites Antineoplastic Agents Appearance Award Behavior Binding Binding Sites Biological Process Biological Products Biological Response Modifier Therapy Blood capillaries Chemistry Comparative Study Dangerousness Detection Drug Targeting Environment Epitopes Extracellular Protein Fluorescence Fluorine G-Protein-Coupled Receptors Generations Goals Growth Factor Higher Order Chromatin Structure Human Hydrophobicity Hydroxyl Radical Immune response Industry Label Laboratories Lasers Lead Letters Ligands Link Membrane Membrane Proteins Modification Molecular Conformation Monitor Monoclonal Antibodies National Institute of General Medical Sciences Nature Patients Performance Pharmaceutical Preparations Pharmacologic Substance Pharmacology Phase Phosphotransferases Play Process Production Protein Conformation Protein Footprinting Proteins Proteome Proteomics Reaction Reproducibility Research Role Safety Signal Transduction Small Business Innovation Research Grant Sodium Solvents Structure Surface Techniques Technology Time Toxin Transmembrane Domain Universities Washington Work adverse drug reaction analytical tool aqueous base detector dosimetry druggable target experimental study improved innovation interest new technology novel pathogen pathogenic virus professor programs research and development tool

项目摘要

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.

GenNext 第一阶段 SBIR 提交的题为“多波长荧光” 用于实时评估蛋白质足迹自由基产量的自由基剂量测定”是满足对新的和改进的更高阶工具的公认需求生物制药和膜蛋白靶点研究的结构分析（HOS）。一种新兴的 HOS 分析技术是羟基自由基足迹 (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通过随后的 MS 分析来鉴定蛋白质的外部部分。最近开发了一种新的、有价值的商业解决方案来执行HRPF。依赖于 OH 产生的三氟甲基 (TFM) 自由基的足迹技术三氟甲磺酸钠水溶液的自由基攻击已经开发出来，显示出巨大的前景与 HRPF 结合使用时，TFM 蛋白足迹 (TFMPF) 具有高度互补性。 HRPF，因为 TFMPF 有效地标记相对“沉默”的氨基酸残基当同时使用时，HRPF 和 TFMPF 可以提供大量的 OH 激进攻击。以及溶剂可及残留物的检测，因此代表了变革性的改进生物制药 HOS 评估。 TFMPF 的实践是由 Michael Gross 教授首创的。密苏里州圣路易斯华盛顿大学。同时显示出解决未满足问题的巨大希望药物研究中的挑战，TFMPF 的重现性受到以下挑战背景清除的可变性与 Gross 实验室合作，我们的工作。将把我们创新的 HRPF 自由基剂量测定技术扩展到 TFMPF。 Technologies 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 流程的联合使用从学术研究实验成为有价值的分析工具。转变为一种强大的技术，我们设想 HRPF 和 TFMPF 能够实现：单克隆抗体生物药物与互补位和表位的相互作用他们的膜靶点；阐明铅与正构或变构膜靶标；揭示次级信使信号级联 GPCR先导化合物；并检测正构/变构抗-的影响肿瘤针对激酶和生长因子等靶标。