De novo design of a generalizable protein biosensor platform for point-of-care testing

用于即时测试的通用蛋白质生物传感器平台的从头设计

基本信息

批准号：
10836196
负责人：
Hsien Wei YEH
金额：
$ 24.9万
依托单位：
UNIVERSITY OF CALIFORNIA SANTA CRUZ
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10836196
关键词：
2019-nCoV Address Affinity Algorithms Antibodies Area Basic Science Binding Binding Proteins Biological Assay Biological Markers Bioluminescence Biosensor Body Fluids Botulinum Toxins Bypass COVID-19 COVID-19 pandemic Cardiac Caring Clinic Clinical Collection Color Colorimetry Consumption Coupled Coupling Creativeness Darkness Detection Development Development Plans Devices Diagnostic Diagnostic Reagent Diagnostic Services Directed Molecular Evolution Disease Drops ERBB2 gene Economics Energy Transfer Engineering Enzyme-Linked Immunosorbent Assay Equilibrium Event Fc domain Future Gene Order Goals Health Priorities Healthcare Hepatitis B Antibodies Hepatitis B Virus Home Hospitals Human Incubated Interleukin-6 Light Luciferases Measurable Measures Medical Mentors Mentorship Methods Mission Modality Molecular Molecular Conformation Monitor Monoclonal Antibodies Names Oligonucleotide Microarrays Patients Peptides Phase Protein Engineering Proteins Published Comment Reporter Reporting Research Research Personnel SARS-CoV-2 antibody Sampling Serology test Signal Transduction Specimen Surface System Technology Therapeutic Thermodynamics Time Treatment outcome Troponin I United States National Institutes of Health Work Yeasts antibody detection career career development clinically relevant complement system design disability emergency settings flu health care availability health care quality health care service improved instrument interest luminescence molecular diagnostics neutralizing antibody novel pandemic impact pathogen personalized medicine point of care point of care testing point-of-care diagnostics precision medicine protein biomarkers ratiometric receptor receptor binding seasonal influenza sensor synergism systemic inflammatory response tv watching

项目摘要

úú PROJECT SUMMARY/ABSTRACT Delivering diagnostic services at the point-of-care (POC) can improve the quality of healthcare in clinics, in emergency settings, or at home, which can potentially ease hospitals’ burden, for instance, during the COVID- 19 pandemic. Precision and personalized medicine revolution also require POC testing to provide readily available biomarker information to clinicians. The goal of this career development proposal is to create fast, inexpensive, sensitive, and reliable molecular diagnostics to address the 21st-century healthcare challenges. The central hypothesis is that we can efficiently utilize computational protein design to create modular allosteric protein switches, named LOCKR (Latching Orthogonal Cage–Key pRotein), that enable the rapid and reversible conformational changes upon interaction. As a proof of principle, we demonstrate that LOCKR- based biosensors can be configured to produce bioluminescence upon the addition of clinical targets (e.g., botulinum toxin, cardiac troponin I, HER2 receptor, Fc domain, anti-HBV mAb, anti-SARS-CoV2 antibodies, and SARS-CoV2 receptor-binding domain/spike protein, Fig 1 and 2) in homogeneous “all-in-solution” assays. Due to the modularity of LOCKR sensor platform and the advance in de novo binder design for arbitrary protein targets, we proposed the integration of both features as the universal strategy to develop tailored biosensors for user-defined targets. The main specific aims for the independent phase are to iteratively expand LOCKR-based diagnostics with the synergy of (1) de novo protein binder design to directly detect various disease protein biomarkers, and (2) indirectly detect the antibodies that compete with the designed interface, as POC devices; and (3) to repurpose the original luminescence signal with other compatible readouts by exchanging the reporter modules. For more specific proof-of-concept projects during the mentored phase, I describe in Aim 1 the use of state-of-the-art computational protein design methods to create an interleukin-6 binder and biosensor. In Aim 2, I propose a general way to develop antibody biosensors by demonstrating COVID-19 serological tests as an example. With my expertise in biosensor engineering, I attempt in Aim 3 to develop a ratiometric bioluminescence resonance energy transfer (BRET) biosensor to analyze the HBV antibody and a colorimetric biosensor to measure human cardiac troponin I level. Ultimately, I anticipate this new sensor platform is significant for the development of robust protein sensors that will be broadly applicable to arbitrary targets and enabling its POC compatible readouts for future diagnostics.

úú 项目概要/摘要在护理点 (POC) 提供诊断服务可以提高诊所的医疗保健质量紧急情况下或在家中，这可能会减轻医院的负担，例如在新冠疫情期间 19、精准化和个性化医疗革命也需要POC测试能够随时提供。向追随者提供可用的生物标志物信息该职业发展建议的目标是创造快速、廉价、灵敏且可靠的分子诊断技术可应对 21 世纪的医疗保健挑战。中心假设是我们可以有效地利用计算蛋白质设计来创建模块化变构蛋白开关，称为 LOCKR（闩锁正交笼 - 关键蛋白），能够快速作为原理证明，我们证明了 LOCKR- 相互作用时的可逆构象变化。基于生物传感器的配置可以在添加临床目标（例如，肉毒杆菌毒素、心肌肌钙蛋白 I、HER2 受体、Fc 结构域、抗 HBV mAb、抗 SARS-CoV2 抗体、和 SARS-CoV2 受体结合结构域/刺突蛋白，图 1 和 2）均质“全溶液” 由于 LOCKR 传感器平台的模块化和从头粘合剂设计的进步。任意蛋白质目标，我们提出将这两种特征整合作为开发的通用策略针对用户定义的目标定制的生物传感器独立阶段的主要具体目标是通过 (1) 从头蛋白结合剂设计的协同作用迭代扩展基于 LOCKR 的诊断，以直接检测各种疾病蛋白生物标志物，(2)间接检测与疾病蛋白竞争的抗体设计接口，作为POC设备；以及（3）将原始发光信号重新用于其他用途通过交换报告模块来实现更具体的概念验证项目。在指导阶段，我在目标 1 中描述了使用最先进的计算蛋白质设计方法来创建 IL-6 结合剂和生物传感器在目标 2 中，我提出了一种开发抗体的通用方法。以我在生物传感器方面的专业知识演示 COVID-19 血清学测试为例。工程方面，我尝试在目标 3 中开发比例生物发光共振能量转移 (BRET) 用于分析 HBV 抗体的生物传感器和用于测量人心肌肌钙蛋白 I 的比色生物传感器最终，我预计这个新的传感器平台对于稳健蛋白质的开发具有重要意义。传感器将广泛适用于任意目标，并支持未来的 POC 兼容读数诊断。