有机磷水解酶和 pHlourin 的融合蛋白，用于有机磷农药测量的全细胞生物传感器。
Fusion proteins of organophosphorus hydrolase and pHluorin for a whole-cell biosensor for organophosphorus pesticide measurement.
机构信息
School of Bioscience and Biotechnology, Tokyo University of Technology, 1404-1 Katakura, Hachioji, Tokyo, 192-0982, Japan.
出版信息
Anal Sci. 2023 Sep;39(9):1515-1520. doi: 10.1007/s44211-023-00369-7. Epub 2023 Jun 1.
Fusion proteins composed of an organophosphorus hydrolase (OPH) and pHluorin, a pH-sensitive green fluorescent protein variant, were constructed as whole-cell biosensors to measure organophosphorus pesticides. pHluorin was used to detect the pH changes because of the hydrolase of paraoxon by OPH. To examine the order of fusion of OPH and pHluorin, pHluorin-OPH and OPH-pHluorin fusion proteins were constructed. In addition, a peptide linker consisting of 15 amino acid was inserted between pHluorin and OPH to reduce steric hindrance. OPH and pHluorin activities were evaluated in cells expressing the four fusion proteins. The both activities of pHluorin-OPH and pHluorin-linker-OPH were higher than that of OPH-pHluorin and OPH-linker-pHluorin. Effects of the peptide linker on the activities were slight. Therefore, pHluorin-OPH and pHluorin-linker-OPH were found to be suitable for organophosphorus pesticide measurements. Using cells expressing pHluorin-linker-OPH, 0.5 μg/mL of paraoxon could be measured.
融合蛋白由有机磷水解酶（OPH）和 pHluroin 组成，pHluroin 是一种对 pH 敏感的绿色荧光蛋白变体，可作为全细胞生物传感器来测量有机磷农药。pHluroin 用于检测由于 OPH 对氧乐果的水解而引起的 pH 变化。为了研究 OPH 和 pHluroin 的融合顺序，构建了 pHluroin-OPH 和 OPH-pHluroin 融合蛋白。此外，在 pHluroin 和 OPH 之间插入由 15 个氨基酸组成的肽接头，以减少空间位阻。在表达四种融合蛋白的细胞中评估了 OPH 和 pHluroin 的活性。pHluroin-OPH 和 pHluroin-linker-OPH 的两种活性均高于 OPH-pHluroin 和 OPH-linker-pHluroin。肽接头对活性的影响很小。因此，pHluroin-OPH 和 pHluroin-linker-OPH 适合用于有机磷农药的测量。使用表达 pHluroin-linker-OPH 的细胞，可以测量到 0.5μg/mL 的对氧乐果。
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