Biomimetic Chemical Sensing by Fluorescence Signals Using a Virus-like Particle-Based Platform

The chemical receptors present in living organisms are promising tools for developing biomimetic chemical sensors. However, these receptors require lipid membranes for functioning under physiological conditions, which prevents their utilization in the production of cell-free in vitro chemical sensing systems. Here, we report the development of a cell-free biomimetic sensing platform using virus-like particles (VLPs) with intact ligand-gated Ca²⁺ channels and genetically encoded Ca²⁺ indicator (GECI). We observed that targeting GECI to the plasma membrane was essential for efficient loading GECI in the VLPs. Although the physiological Ca²⁺ concentration [Ca²⁺] maintained in the cells was low (∼10 nM), the concentration in the VLPs was high. This prevented the detection of the increase in [Ca²⁺] caused by binding of the ligand to the receptor. To address this problem, we employed Lyn-R-CEPIA1, which had low affinity for Ca²⁺, and a membrane targeting sequence. Thus, we succeeded in monitoring the activation of cyclic nucleotide-gated channels (CNG) on the VLPs by measuring the increase in fluorescence of Lyn-R-CEPIA1. Our VLP-based sensing system can act as a fundamental platform for all kinds of ligand-gated channels.