Understanding proteomic diversity at the molecular level is essential for advances in biology and medicine, but sequencing proteins in their full, native form remains challenging. A study in Nature by Motone et al. introduces a nanopore sequencing method that can read single, full-length protein molecules, enabling the detection of single amino acid substitutions and post-translational modifications (PTMs).

The method involves a two-step process. First, a protein is loaded into a nanopore formed by the bacterial lipoprotein CsgC by electrophoretic force, facilitated by a negatively charged tail domain and the addition of a ‘stopper’ sequence. Then, ClpX unfoldase steadily pulls it back through in a trans-to-cis direction. Using synthetic proteins, the authors show their method can detect single amino acid substitutions and can map the activity of kinases, identifying site-specific PTMs such as phosphorylation.

在分子水平上了解蛋白质组学多样性对于生物学和医学的进步至关重要，但对蛋白质的完整天然形式进行测序仍然具有挑战性。Motone 等人在《自然》杂志上发表的一项研究介绍了一种纳米孔测序方法，该方法可以读取单个全长蛋白质分子，从而能够检测单个氨基酸取代和翻译后修饰 （PTM）。

该方法涉及两个步骤。首先，蛋白质通过电泳力加载到由细菌脂蛋白 CsgC 形成的纳米孔中，这得益于带负电荷的尾部结构域和添加“阻断”序列。然后，ClpX 去折叠酶将其稳定地沿反式到顺式方向拉回。使用合成蛋白质，作者表明他们的方法可以检测单个氨基酸替换，并且可以绘制激酶的活性，识别位点特异性 PTM，例如磷酸化。