Department of Botany and Plant Sciences, University of California, Riverside, Riverside, CA, USA.
Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA, USA.
Nat Biotechnol. 2022 Dec;40(12):1855-1861. doi: 10.1038/s41587-022-01364-5. Epub 2022 Jun 20.
A general method to generate biosensors for user-defined molecules could provide detection tools for a wide range of biological applications. Here, we describe an approach for the rapid engineering of biosensors using PYR1 (Pyrabactin Resistance 1), a plant abscisic acid (ABA) receptor with a malleable ligand-binding pocket and a requirement for ligand-induced heterodimerization, which facilitates the construction of sense-response functions. We applied this platform to evolve 21 sensors with nanomolar to micromolar sensitivities for a range of small molecules, including structurally diverse natural and synthetic cannabinoids and several organophosphates. X-ray crystallography analysis revealed the mechanistic basis for new ligand recognition by an evolved cannabinoid receptor. We demonstrate that PYR1-derived receptors are readily ported to various ligand-responsive outputs, including enzyme-linked immunosorbent assay (ELISA)-like assays, luminescence by protein-fragment complementation and transcriptional circuits, all with picomolar to nanomolar sensitivity. PYR1 provides a scaffold for rapidly evolving new biosensors for diverse sense-response applications.
一种生成针对用户定义分子的生物传感器的通用方法，可以为广泛的生物应用提供检测工具。在这里，我们描述了一种使用 PYR1（Pyrabactin Resistance 1）快速工程生物传感器的方法，PYR1 是一种植物脱落酸（ABA）受体，具有可塑的配体结合口袋和配体诱导异二聚化的要求，这有利于构建感应-响应功能。我们将该平台应用于进化出 21 种具有纳摩尔到微摩尔灵敏度的传感器，用于一系列小分子，包括结构多样的天然和合成大麻素以及几种有机磷。X 射线晶体学分析揭示了进化的大麻素受体对新配体识别的机制基础。我们证明，源自 PYR1 的受体可以很容易地应用于各种配体响应输出，包括酶联免疫吸附测定（ELISA）样测定、通过蛋白片段互补的发光和转录电路，所有这些都具有皮摩尔到纳摩尔的灵敏度。PYR1 为快速进化针对各种感应-响应应用的新型生物传感器提供了一个支架。
