School of Biomedical Sciences, University of Leeds, Leeds, UK.
Defence Science and Technology Laboratory, Salisbury, UK.
Adv Biochem Eng Biotechnol. 2024;187:1-35. doi: 10.1007/10_2023_241.
Rapid detection of protein and small-molecule analytes is a valuable technique across multiple disciplines, but most in vitro testing of biological or environmental samples requires long, laborious processes and trained personnel in laboratory settings, leading to long wait times for results and high expenses. Fusion of recognition with reporter elements has been introduced to detection methods such as enzyme-linked immunoassays (ELISA), with enzyme-conjugated secondary antibodies removing one of the many incubation and wash steps. Chimeric protein switch biosensors go further and provide a platform for homogenous mix-and-read assays where long wash and incubation steps are eradicated from the process. Chimeric protein switch biosensors consist of an enzyme switch (the reporter) coupled to a recognition element, where binding of the analyte results in switching the activity of the reporter enzyme on or off. Several chimeric protein switch biosensors have successfully been developed for analytes ranging from small molecule drugs to large protein biomarkers. There are two main formats of chimeric protein switch biosensor developed, one-component and multi-component, and these formats exhibit unique advantages and disadvantages. Genetically fusing a recognition protein to the enzyme switch has many advantages in the production and performance of the biosensor. A range of immune and synthetic binding proteins have been developed as alternatives to antibodies, including antibody mimetics or antibody fragments. These are mainly small, easily manipulated proteins and can be genetically fused to a reporter for recombinant expression or manipulated to allow chemical fusion. Here, aspects of chimeric protein switch biosensors will be reviewed with a comparison of different classes of recognition elements and switching mechanisms.
快速检测蛋白质和小分子分析物是多个学科都非常有价值的技术，但大多数生物或环境样本的体外检测都需要在实验室环境中经过漫长而繁琐的过程和经过培训的人员，导致结果等待时间长且费用高。识别与报告元件的融合已被引入到酶联免疫吸附测定（ELISA）等检测方法中，其中酶结合的二级抗体去除了许多孵育和洗涤步骤之一。嵌合蛋白开关生物传感器更进一步，为均相混合和读取测定提供了一个平台，其中从过程中消除了长洗涤和孵育步骤。嵌合蛋白开关生物传感器由酶开关（报告器）与识别元件偶联而成，其中分析物的结合导致报告酶的活性开启或关闭。已经成功开发了几种用于从小分子药物到大蛋白生物标志物的分析物的嵌合蛋白开关生物传感器。已经开发出两种主要格式的嵌合蛋白开关生物传感器，即单组分和多组分，这些格式表现出独特的优缺点。在生物传感器的生产和性能方面，将识别蛋白与酶开关基因融合具有许多优势。已经开发了一系列免疫和合成结合蛋白作为抗体的替代品，包括抗体模拟物或抗体片段。这些主要是小的、易于操作的蛋白，可以与报告蛋白进行基因融合以进行重组表达，或进行操作以允许化学融合。在这里，将对嵌合蛋白开关生物传感器的各个方面进行综述，并比较不同类别的识别元件和开关机制。
