Abstract：
The abnormal expression of protein tyrosine phosphatase 1B (PTP1B) is highly related to several serious human diseases. Therefore, an accurate PTP1B activity assay is beneficial to the diagnosis and treatment of these diseases. In this study, a dual-mode biosensing platform that enabled the sensitive and accurate assay of PTP1B activity was constructed based on the high-frequency (100 MHz) quartz crystal microbalance (QCM) and dual-signaling electrochemical (EC) ratiometric strategy. Covalent-organic framework@gold nanoparticles@ferrocene@single-strand DNA (COF@Au@Fc-S0) was introduced onto the QCM Au chip via the chelation between Zr4+ and phosphate groups (phosphate group of the phosphopeptide (P-peptide) on the QCM Au chip and the phosphate group of thiol-labeled single-stranded DNA (S0) on COF@Au@Fc-S0) and used as a signal reporter. When PTP1B was present, the dephosphorylation of the P-peptide led to the release of COF@Au@Fc-S0 from the QCM Au chip, resulting in an increase in the frequency of the QCM. Meanwhile, the released COF@Au@Fc-S0 hybridized with thiol/methylene blue (MB)-labeled hairpin DNA (S1-MB) on the Au NPs-modified indium-tin oxide (ITO) electrode. This caused MB to be far away from the electrode surface and Fc to be close to the electrode, leading to a decrease in the oxidation peak current of MB and an increase in the oxidation peak current of Fc. Thus, PTP1B-induced dephosphorylation of the P-peptide was monitored in real time by QCM, and PTP1B activity was detected sensitively and reliably using this innovative QCM-EC dual-mode sensing platform with an ultralow detection limit. This platform is anticipated to serve as a robust tool for the analysis of protein phosphatase activity and the discovery of drugs targeting protein phosphatase.
摘要：
蛋白酪氨酸磷酸酶1B(PTP1B)的异常表达与几种严重的人类疾病高度相关。因此,准确的PTP1B活性测定有利于这些疾病的诊断和治疗。在这项研究中,基于高频(100MHz)石英晶体微天平(QCM)和双信号电化学(EC)比率计量策略,构建了一个能够灵敏和准确测定PTP1B活性的双模式生物传感平台.共价有机框架@金纳米颗粒@二茂铁@单链DNA（COF@Au@Fc-S0）通过Zr4和磷酸基团（磷酸肽的磷酸基团（P肽）在QCMAu芯片上和COF@Au@Fc-S0上的硫醇标记的单链DNA（S0）的磷酸基团）之间的螯合被引入QCMAu芯片上，并用作报告信号。当PTP1B出现时，P-肽的去磷酸化导致COF@Au@Fc-S0从QCMAu芯片中释放,导致QCM的频率增加。同时,释放的COF@Au@Fc-S0在AuNP修饰的氧化铟锡（ITO）电极上与硫醇/亚甲基蓝（MB）标记的发夹DNA（S1-MB）杂交。这导致MB远离电极表面，Fc靠近电极，导致MB的氧化峰电流降低和Fc的氧化峰电流增加。因此，通过QCM实时监测PTP1B诱导的P肽去磷酸化，和PTP1B活性检测灵敏和可靠地使用这种创新的QCM-EC双模式传感平台具有超低的检测限。预计该平台将作为分析蛋白磷酸酶活性和发现靶向蛋白磷酸酶的药物的强大工具。
