CY Cergy Paris Université, CNRS, LAMBE, Cergy, 95000, France.
Department of Physics, University of South Florida, Tampa, Florida 33620, United States.
J Am Chem Soc. 2022 Feb 16;144(6):2716-2725. doi: 10.1021/jacs.1c11758. Epub 2022 Feb 4.
The implementation of a reliable, rapid, inexpensive, and simple method for whole-proteome identification would greatly benefit cell biology research and clinical medicine. Proteins are currently identified by cleaving them with proteases, detecting the polypeptide fragments with mass spectrometry, and mapping the latter to sequences in genomic/proteomic databases. Here, we demonstrate that the polypeptide fragments can instead be detected and classified at the single-molecule limit using a nanometer-scale pore formed by the protein aerolysin. Specifically, three different water-soluble proteins treated with the same protease, trypsin, produce different polypeptide fragments defined by the degree by which the latter reduce the nanopore's ionic current. The fragments identified with the aerolysin nanopore are consistent with the predicted fragments that trypsin could produce.
一种可靠、快速、廉价且简单的全蛋白质组鉴定方法的实现将极大地有益于细胞生物学研究和临床医学。目前，蛋白质是通过用蛋白酶切割它们，用质谱法检测多肽片段，并将后者映射到基因组/蛋白质组数据库中的序列来鉴定的。在这里，我们证明可以使用aerolysin 蛋白形成的纳米级孔在单分子极限下检测和分类多肽片段。具体来说，用相同的蛋白酶胰蛋白酶处理三种不同的水溶性蛋白质会产生不同的多肽片段，这些片段由后者降低纳米孔离子电流的程度来定义。用 aerolysin 纳米孔鉴定的片段与胰蛋白酶可能产生的预测片段一致。
