Experimental Research of Protein Translocation Using Solid-state Nanopore
Received date: 2017-06-16
Online published: 2017-09-04
Supported by
Project supported by the National Natural Science Foundation of China (Nos. 51375092, 51435003 and 51675101) and the Fundamental Research Funds for the Central Universities (No. 2242015R30002).
For proteins' diverse range of structural and functional features, they are important populations of biomolecules within organisms. Common methods to detect proteins are with the help of fluorescence or enzyme. Due to the advantages like lable-free and single-molecule detection, nanopore technology provides a novel platform for proteins' characterization. In this experiment, the patch clamp amplifier is used to apply the voltage and acquire the tiny current blockage. The Si3N4 membrane drilled with a nanopore separated the buffer solution into two sides:cis and trans. When the voltage applied into the buffer solution, charged proteins would been driven through the pore from one side to the other. Then, a series of current blockages could be obtained. By analysing these data, the size and conformation of the biomolecules could be acquired. In this paper, we using solid-state nanopore detected single protein and protein-protein complexes. The nanopore was characterized firstly. Then, both the applied voltage and the pH of the electrolyte solution were regulated. Under the low voltage, the sample proteins could be regarded as a rigid spheroid, and the dwell time is decreased with the voltage increasing. It was found that, the charges carried by proteins could be improved by higher pH of buffer solution, so that the dwell time would been shortened. Furthermore, based on the high specific between the antigen and antibody which are proteins, the translocation events before and after the addition of specific antigen into the solution with antibody were compared. Results showed that the relative current drop of the complex is larger than the pure antibody, implying that the antigen has been bound into the antibody. Due to the difference of excluded volume, the antibody and antigen-antibody complexes could be distinguished by the solid-state nanopore. In the future, the nanopore technology is promising to be applied into the recognition of multiply proteins and protein-protein interaction.
Sha Jingjie , Xu Bing , Chen Yunfei , Yang Yanjing . Experimental Research of Protein Translocation Using Solid-state Nanopore[J]. Acta Chimica Sinica, 2017 , 75(11) : 1121 -1125 . DOI: 10.6023/A17060271
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