All-Atoms Md Simulation of Protein Translocation through α-Hemolysin Nanopore: Implications for Protein Sequence/Structural Analyses

Abstract

The cell isolation from the external environment and the cellular compartmentalization is due to the presence of membranes composed by phospholipidic bilayers. Different proteins that play numerous biological functions are present in the membrane. These proteins are mainly involved in the communication between the cell and the external environment and between the different cellular compartments. Some of them form pores into the lipid bilayer that allow the translocation of molecules of different size and with different chemical-physical properties. The transport of these molecules across the membrane is crucial for the cellular survival. This feature of the membrane proteins has been extensively studied in order to use them as nanopores in biochemical and industrial fields.During the last decades nanopores have been exploited in the development of new techniques for nucleic acids sequencing. Much less effort has been dedicated to protein and polypeptide analysis using nanopores. Only in the last years pioneering studies appeared in the literature suggesting potentially revolutionary applications in the study of protein sequence and structure. Recent experimental data have demonstrated that, in order to be translocated across nanopores, proteins must be unfolded, with the unfolding process occurring through a multi-step mechanism. It has been observed, for instance, that the transport of the thioredoxin through the α-hemolysin nanopore occurs via a four steps process.In this work, all atom molecular dynamics simulations of the thioredoxin translocation across the α-hemolysin nanopore have been performed. The data obtained allowed the description of the molecular mechanisms at the basis of the protein transport at the atomic level. Our results allowed us to gain a deeper knowledge regarding the translocation of protein through nanopores, useful for the development of nanopore based applications for several crucial biochemical analyses.