CICA-Centro Interdisciplinar de Química e Bioloxía and Departamento de Química, Facultade de Ciencias, Universidade da Coruña. Campus de Elviña, 15071 A Coruña, Spain.
Centro Nacional de Biotecnología, CSIC, Darwin 3, 28049 Madrid, Spain.
ACS Sens. 2024 Oct 25;9(10):5052-5057. doi: 10.1021/acssensors.4c00986. Epub 2024 Sep 6.
The detection and monitoring of and its virulence factors, such as the LasB protease, are crucial for managing bacterial infections. Traditional fluorescent sensors for this protease face limitations in bacterial cultures due to interference from pigments like pyoverdine secreted by this opportunistic pathogen. We report here a Ln(III)-metallopeptide that combines a DO3A-Ln(III) complex and a sensitizing unit via a short peptide sequence as a simple, tunable, and selective probe for detecting 's LasB. The probe's luminescence switches off in the presence of 's secretome due to LasB cleavage but remains stable in other bacterial environments, such as non-LasB-secreting strains or cultures. It also resists degradation by other proteases, like human leukocyte elastase and trypsin, and remains stable in the presence of bioanalytes related to infections, such as glutathione, HO, and pyocyanin, and in complex media like FBS. Importantly, time-gated experiments completely remove the background fluorescence of pigments, thus demonstrating the potential of the developed Ln(III)-metallopeptide for real-time monitoring of LasB activity in bacterial cultures.
检测和监测及其毒力因子，如 LasB 蛋白酶，对于管理细菌感染至关重要。传统的荧光传感器用于该蛋白酶时，由于这种机会性病原体分泌的绿脓菌素等色素的干扰，在细菌培养物中受到限制。我们在这里报告了一种 Ln(III)-金属肽，它通过短肽序列将 DO3A-Ln(III) 络合物和敏化单元结合在一起，作为一种简单、可调谐和选择性的探针，用于检测 LasB。由于 LasB 切割，探针的发光在存在 '的分泌物时关闭，但在其他细菌环境中保持稳定，如不分泌 LasB 的菌株或 '的培养物。它还能抵抗其他蛋白酶，如人白细胞弹性蛋白酶和胰蛋白酶的降解，并在与 '感染相关的生物分析物（如谷胱甘肽、HO 和绿脓菌素）和复杂介质（如 FBS）存在的情况下保持稳定。重要的是，时间门控实验完全消除了 '色素的背景荧光，从而证明了所开发的 Ln(III)-金属肽在细菌培养物中实时监测 LasB 活性的潜力。
