Pollution of heavy metals in agricultural environments is a growing problem to the health of the world’s human population. Green, low-cost, and efficient detection methods can help control such pollution. In this study, a protein biosensor, mApple-D6A3, was built from rice-derived Cd²⁺-binding protein D6A3 fused with the red fluorescent protein mApple at the N-terminus to detect the contents of heavy metals. Fluorescence intensity of mApple fused with D6A3 indicated the biosensor’s sensitivity to metal ions and its intensity was more stable under alkaline conditions. mApple-D6A3 was most sensitive to Cu²⁺, then Ni²⁺, then Cd²⁺. Isothermal titration calorimetry experiments demonstrated that mApple-D6A3 successfully bound to each of these three metal ions, and its ability to bind the ions was, from strongest to weakest, Cu²⁺ > Cd²⁺ > Ni²⁺. There were strong linear relationships between the fluorescence intensity of mApple-D6A3 and concentrations of Cd²⁺ (0–100 μM), Cu²⁺ (0–60 μM) and Ni²⁺ (0–120 μM), and their respective R² values were 0.994, 0.973 and 0.973. When mApple-D6A3 was applied to detect concentrations of heavy metal ions in water (0–0.1 mM) or culture medium (0–1 mM), its accuracy for detection attained more than 80%. This study demonstrates the potential of this biosensor as a tool for detection of heavy metal ions.

农业环境中的重金属污染是世界人口健康日益严重的问题。绿色，低成本和高效的检测方法可以帮助控制此类污染。在这项研究中，蛋白质生物传感器mApple-D6A3是由水稻衍生的Cd ²⁺结合蛋白D6A3与红色荧光蛋白mApple在N端融合而成，以检测重金属的含量。mApple与D6A3融合的荧光强度表明该生物传感器对金属离子的敏感性，并且在碱性条件下其强度更加稳定。mApple-D6A3对Cu ²⁺，然后Ni ²⁺，然后Cd ²⁺最敏感。等温滴定量热法实验表明，mApple-D6A3成功结合了这三种金属离子，其结合离子的能力从最强到最弱，从Cu ²⁺ > Cd ²⁺ > Ni ^2+开始。mApple-D6A3的荧光强度与Cd ²⁺（0–100μM ），Cu ²⁺（0–60μM）和Ni ²⁺（0–120μM）的浓度及其各自的R之间存在强线性关系²值分别为0.994、0.973和0.973。当mApple-D6A3用于检测水（0–0.1 mM）或培养基（0–1 mM）中重金属离子的浓度时，其检测精度达到80％以上。这项研究证明了这种生物传感器作为检测重金属离子的工具的潜力。