title: Potential analytical applications of lysenin channels for detection of multivalent ions.
abstract: Transmembrane protein transporters possessing binding sites for ions, toxins, pharmaceutical drugs, and other molecules constitute excellent candidates for developing sensitive and selective biosensing devices. Their attractiveness for analytical purposes is enhanced by the intrinsic amplification capabilities shown when the binding event leads to major changes in the transportation of ions or molecules other than the analyte itself. The large-scale implementation of such transmembrane proteins in biosensing devices is limited by the difficulties encountered in inserting functional transporters into artificial bilayer lipid membranes and by the limitations in understanding and exploiting the changes induced by the interaction with the analyte for sensing purposes. Here, we show that lysenin, a pore-forming toxin extracted from earthworm Eisenia foetida, which inserts stable and large conductance channels into artificial bilayer lipid membranes, functions as a multivalent ion-sensing device. The analytical response consists of concentration and ionic-species-dependent macroscopic conductance inhibition most probably linked to a ligand-induced gating mechanism. Multivalent ion removal by chelation or precipitation restores, in most cases, the initial conductance and demonstrates reversibility. Changes in lipid bilayer membrane compositions leading to the absence of voltage-induced gating do not affect the analytical response to multivalent ions. Microscopic current analysis performed on individual lysenin channels in the presence of Cu(2+) revealed complex open-closed transitions characterized by unstable intermediate sub-conducting states. Lysenin channels provide an analytical tool with a built-in sensing mechanism for inorganic and organic multivalent ions, and the excellent stability in an artificial environment recommend lysenin as a potential candidate for single-molecule detection and analysis.
authors.affiliation: Department of Biological Sciences, University of Arkansas, Fayetteville, AR 72701, USA. DanielFologea@boisestate.edu
authors.forename: Daniel
authors.initials: D
authors.lastname: Fologea
authors.forename: Redwan
authors.initials: R
authors.lastname: Al Faori
authors.forename: Eric
authors.initials: E
authors.lastname: Krueger
authors.forename: Yuriy I
authors.initials: YI
authors.lastname: Mazur
authors.forename: Matt
authors.initials: M
authors.lastname: Kern
authors.forename: Matt
authors.initials: M
authors.lastname: Williams
authors.forename: Amir
authors.initials: A
authors.lastname: Mortazavi
authors.forename: Ralph
authors.initials: R
authors.lastname: Henry
authors.forename: Greg J
authors.initials: GJ
authors.lastname: Salamo
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D055664:Electrochemical Techniques / Q000379:methods; D007477:Ions / Q000032:analysis* / Q000378:metabolism; D008051:Lipid Bilayers / Q000378:metabolism; D009835:Oligochaeta / Q000378:metabolism*; D012680:Sensitivity and Specificity; D014118:Toxins, Biological / Q000378:metabolism*
chemical_list: D007477:Ions; D008051:Lipid Bilayers; D014118:Toxins, Biological; C107040:lysenin