Department of Physics , Syracuse University , 201 Physics Building , Syracuse , New York 13244-1130 , United States.
Structural Biology, Biochemistry, and Biophysics Program , Syracuse University , 111 College Place , Syracuse , New York 13244-4100 , United States.
ACS Sens. 2019 Sep 27;4(9):2320-2326. doi: 10.1021/acssensors.9b00848. Epub 2019 Aug 21.
Protein detection in complex biological fluids has wide-ranging significance across proteomics and molecular medicine. Existing detectors cannot readily distinguish between specific and nonspecific interactions in a heterogeneous solution. Here, we show that this daunting shortcoming can be overcome by using a protein bait-containing biological nanopore in mammalian serum. The capture and release events of a protein analyte by the tethered protein bait occur outside the nanopore and are accompanied by uniform current openings. Conversely, nonspecific pore penetrations by nontarget components of serum, which take place inside the nanopore, are featured by irregular current blockades. As a result of this unique peculiarity of the readout between specific protein captures and nonspecific pore penetration events, our selective sensor can quantitatively sample proteins at single-molecule precision in a manner distinctive from those employed by prevailing methods. Because our sensor can be integrated into nanofluidic devices and coupled with high-throughput technologies, our approach will have a transformative impact in protein identification and quantification in clinical isolates for disease prognostics and diagnostics.
在蛋白质组学和分子医学中，复杂生物流体中的蛋白质检测具有广泛的意义。现有的检测器不能轻易区分异质溶液中的特异性和非特异性相互作用。在这里，我们表明，通过在哺乳动物血清中使用含有蛋白质诱饵的生物纳米孔可以克服这一令人畏惧的缺点。通过连接的蛋白质诱饵捕获和释放蛋白质分析物的事件发生在纳米孔之外，并伴有均匀的电流开口。相反，血清中非靶标成分的非特异性孔渗透发生在纳米孔内，其特征是不规则的电流阻断。由于这种独特的特点，在特异性蛋白质捕获和非特异性孔渗透事件之间的读出，我们的选择性传感器可以以不同于现有方法的方式以单分子精度定量采样蛋白质。由于我们的传感器可以集成到纳流控设备中，并与高通量技术相结合，因此我们的方法将在临床分离物中的蛋白质鉴定和定量方面产生变革性的影响，用于疾病预后和诊断。
