Biological Nanopore Approach for Single‐Molecule Protein Sequencing

Abstract

Proteins are responsible for the occurrence and treatment of many diseases, and therefore protein sequencing will revolutionize proteomics and clinical diagnostics. Biological nanopore approach has proved successful for single‐molecule DNA sequencing, which resolves the identities of 4 natural deoxyribonucleotides based on the current blockages and duration times of their translocations across the nanopore confinement. However, open challenges still remain for biological nanopores to sequentially identify each amino acid (AA) of single proteins due to the inherent complexity of 20 proteinogenic AAs in charges, volumes, hydrophobicity and structures. Herein, we focus on recent exciting advances in biological nanopores for single‐molecule protein sequencing (SMPS) from native protein unfolding, control of peptide translocation, AA identification to applications in disease detection.

Forensic proteomics

“…Similar in many ways to the Oxford Nanopore third generation DNA sequencing platform, the potential of single protein molecule sequencing has obvious potential for forensics. In addition to increased sensitivity, this approach directly sequences protein and theoretically removes the need to match complex fragmentation spectra with sequences from reference protein databases [371]. There are still significant technical challenges [371].…”

“…In addition to increased sensitivity, this approach directly sequences protein and theoretically removes the need to match complex fragmentation spectra with sequences from reference protein databases [371]. There are still significant technical challenges [371]. Instead of 4 nucleotides there are 21 amino acids with many potential post-translational modifications, resulting in many different hydrodynamic volumes and current blockages [371,372].…”

“…There are still significant technical challenges [371]. Instead of 4 nucleotides there are 21 amino acids with many potential post-translational modifications, resulting in many different hydrodynamic volumes and current blockages [371,372]. Many of these resistivities are very similar [371].…”

Permeation eines 5.1‐kDa‐Peptides durch einen Proteinkanal: Molekulare Basis der Translokation von Protamin durch CymA aus Klebsiella Oxytoca**

“…[33] Mittlerweile ist dieses Gebiet weiter fortgeschritten und ermçglicht unter günstigen Bedingungen sogar die Unterscheidung einzelner Aminosäuren. [34][35][36][37][38] Andere großporige Kanäle wie CymA, die den Durchgang von Peptiden ermçglichen, sind fürdie Peptidsequenzierung von zentralem Interesse.…”

Sensitive Detection of Single-Nucleotide Polymorphisms by Solid Nanopores Integrated With DNA Probed Nanoparticles

“…As an emerging single-molecule detection technology, nanopores are promising sensors for DNA identification, proteomic detection, determination of epigenetic changes, and biomolecular mechanism exploration with the advantages of being label-free and high-throughput (Wang et al, 2017;Jeong et al, 2019;Spitzberg et al, 2019;Chen et al, 2021;Fragasso et al, 2021;Hu et al, 2021). The nanopore technique is inspired by the transmembrane protein channels embedded in lipid bilayers that allow ions and molecules inside and outside the living cell to exchange freely.…”