Investigation of C-terminal domain of SARS nucleocapsid protein-Duplex DNA interaction using transistors and binding-site models

使用晶体管和结合位点模型研究 SARS 核衣壳蛋白 C 端结构域与双链 DNA 的相互作用

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作者:You-Ren Hsu, Yen-Wen Kang, Jung-Ying Fang, Geng-Yen Lee, Jen-Inn Chyi, Chung-Ke Chang, Chih-Cheng Huang, Chen-Pin Hsu, Tai-Huang Huang, Yu-Fen Huang, Yuh-Chang Sun, Chia-Hsien Hsu, Chih-Chen Chen, Sheng-Shian Li, J Andrew Yeh, Da-Jeng Yao, Fan Ren, Yu-Lin Wang

Abstract

AlGaN/GaN high electron mobility transistors (HEMTs) were used to sense the binding between double stranded DNA (dsDNA) and the severe acute respiratory syndrome coronavirus (SARS-CoV) nucleocapsid protein (N protein). The sensing signals were the drain current change of the HEMTs induced by the protein-dsDNA binding. Binding-site models using surface coverage ratios were utilized to analyze the signals from the HEMT-based sensors to extract the dissociation constants and predict the number of binding sites. Two dissociation constants, K D1 = 0.0955 nM, K D2 = 51.23 nM, were obtained by fitting the experimental results into the two-binding-site model. The result shows that this technique is more competitive than isotope-labeling electrophoretic mobility shift assay (EMSA). We demonstrated that AlGaN/GaN HEMTs were highly potential in constructing a semiconductor-based-sensor binding assay to extract the dissociation constants of nucleotide-protein interaction.

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