Polypeptide analysis for nanopore-based protein identification

作     者：Mazdak Afshar Bakshloo Safia Yahiaoui Fabien Piguet Manuela Pastoriza-Gallego Régis Daniel Jérôme Mathé John J.Kasianowicz Abdelghani Oukhaled Mazdak Afshar Bakshloo;Safia Yahiaoui;Fabien Piguet;Manuela Pastoriza-Gallego;Régis Daniel;Jér?me Mathé;John J.Kasianowicz;Abdelghani Oukhaled

作者机构：CY Cergy Paris UniversitéCNRSLAMBE95000CergyFrance UniversitéParis-SaclayUniv EvryCNRSLAMBE91025Evry-CourcouronnesFrance University of South FloridaDept.of PhysicsTampaFL 33620USA Freiburg Institute for Advanced StudiesUniversität FreiburgFreiburg 79104Germany 

出 版 物：《Nano Research》 (纳米研究（英文版）)

年 卷 期：2022年第15卷第11期

页      面：9831-9842页

核心收录：

学科分类：07[理学] 0805[工学-材料科学与工程（可授工学、理学学位）] 0703[理学-化学] 070202[理学-粒子物理与原子核物理] 0702[理学-物理学] 

基　　金：supported by the Agence Nationale de la Recherche ANR(ANR-17-CE09-0032-01 to A.O.,M.P.-G.,and F.P.) NIST Office of Law Enforcement Standards,and a Marie Skłodowska-Curie/Freiburg Institute for Advanced Studies Senior Fellowship(both to J.J.K.) We thank F.Gisou van der Goot(Ecole Polytechnique Federale de Lausanne,Switzerland)for providing the pET22b-proAL plasmid containing the proaerolysin sequence 

主　　题：protein identification nanopores trypsin enzymatic degradation 

摘      要：Presently,proteins are identified by cleaving them with proteases,measuring the mass to charge ratio of the fragments with a mass spectrometer,and matching the fragments to segments within known proteins in *** earlier demonstrated that a nanometer-scale pore formed by aerolysin(AeL)can discriminate between,and therefore identify,three similar size proteins from their trypsin-cleaved polypeptide *** this nanopore-protease method,the protein’s identity is instead determined from characteristic ionic current blockade patterns caused by the polypeptide fragments that enter the *** results also suggested that not all of the theoretically expected cleavage products partition into the *** better understand the mechanism by which polypeptide fragments are captured,and how different polypeptides reduce the pore’s ionic current,we studied the effects of 11 identical length polypeptides with different net charges and charge *** show that under certain experimental conditions,negative,positive,and neutral polypeptides are driven into the AeL pore by the same applied voltage *** capture rate and dwell time of polypeptides in the pore depend strongly on the ionic strength,the magnitude of the applied voltage,and the net charge and charge distribution of the *** dwell time distribution depends nonmonotonically on the applied voltage(regardless of the polymer’s net charge),and its maximum value depends on the polypeptide net charge and charge *** maximum dwell time for different polypeptides does not occur at the same applied voltage amplitude,which conceivably might complicate the detection and discrimination of some polypeptide *** additional experiments,computer simulations,and artificial intelligence research are needed to better understand how to optimize the partitioning of enzymatically cleaved fragments into the AeL nanopore,the method is still capable of accurately identifying