Institute of Chemistry and Center for Nanoscience and Nanotechnology, The Hebrew University of Jerusalem, Safra Campus, Givat Ram, Jerusalem, 91904, Israel.
University of Bayreuth, Bioorganic Chemistry, Universitätsstraße 30, 95447, Bayreuth, Germany.
Biosens Bioelectron. 2021 Jan 15;172:112762. doi: 10.1016/j.bios.2020.112762. Epub 2020 Oct 24.
Sialylated glycans and glycoproteins are involved in cellular communication and are crucial for distinguishing between signal pathways. Sialylation levels and patterns modulate recognition events and are regulated by the enzymatic activity of sialyltransferases and neuraminidases. Abnormal activity of these enzymes is related to diseases such as cancer and viral infection. Monitoring these enzymatic activities offers valuable diagnostic tools. This work presents an impedimetric biosensing platform for following and detecting sialylation and desialylation processes. This platform is based on a native biantennary N-glycan substrate attached to a glassy carbon electrode. Changes in the molecular layer, as a result of enzymatic reactions, were detected by electrochemical impedance spectroscopy, displaying high sensitivity to the enzymatic surface reactions. Increase in the molecular layer roughness in response to the sialylation was visualized using atomic force microscopy. After enzymatic sialylation, the presence of sialic acid was confirmed using cyclic voltammetry by coupling of the redox active marker aminoferrocene. The sialylation showed selectivity toward the N-glycan compared to another glycan substrate. A time dependent sialylation was followed by electrochemical impedance spectroscopy, proving that the new system can be applied to evaluate the enzymatic kinetics. Our findings suggest that analyzing sialylation processes using this platform can become a useful tool for the detection of pathological states and pathogen invasion.
唾液酸化聚糖和糖蛋白参与细胞通讯，对于区分信号通路至关重要。唾液酸化水平和模式调节识别事件，并受唾液酸转移酶和神经氨酸酶的酶活性调节。这些酶的异常活性与癌症和病毒感染等疾病有关。监测这些酶活性提供了有价值的诊断工具。本工作提出了一种基于阻抗的生物传感平台，用于跟踪和检测唾液酸化和去唾液酸化过程。该平台基于附着在玻碳电极上的天然双天线 N-聚糖底物。通过电化学阻抗谱检测到酶反应导致的分子层变化，对酶表面反应具有高灵敏度。原子力显微镜显示，分子层粗糙度的增加对唾液酸化的响应。通过将氧化还原活性标记物氨基二茂铁偶联，在用循环伏安法进行酶唾液酸化后，确认了唾液酸的存在。与另一种糖基底物相比，唾液酸化对 N-聚糖表现出选择性。通过电化学阻抗谱跟踪时间依赖性唾液酸化，证明新系统可用于评估酶动力学。我们的研究结果表明，使用该平台分析唾液酸化过程可能成为检测病理状态和病原体入侵的有用工具。
