title: A high resolution electro-optical approach for investigating transition of soluble proteins to integral membrane proteins probed by colicin A.
abstract: The transition from water soluble state to an integral membrane protein state is a crucial step in the formation of the active form of many pore-forming or receptor proteins. Albeit this, high resolution techniques which allow assay of protein membrane binding and concomitant development of the final active form in the membrane await further development. Here, we describe a horizontal artificial bilayers setup allowing for simultaneous electrical and optical measurements at a single molecule level. We use the membrane binding and subsequent channel formation of colicin A (ColA) a water soluble bacteriocin secreted by some strains of Escherichia coli to demonstrate the potential of the combined electro-optical technique. Our results expand the knowledge on ColA molecular details which show that active ColA is monomeric; membrane binding is pH but not membrane-potential (Δϕ) dependent. ColA is at Δϕ=0 permeable for molecules ≥1 nm. Although ColA exhibits low ion conductance it facilitates permeation of large molecules. Our electro-optical recordings reveal ColA monomeric state and the chimeric character of its pore.
authors.affiliation: Universität Osnabrück, FB Biologie/Chemie, Barbara Str. 13, 49076 Osnabrück, Germany.
authors.forename: Alf
authors.initials: A
authors.lastname: Honigmann
authors.forename: Lakshmi Padmavathi
authors.initials: LP
authors.lastname: Pulagam
authors.forename: Michael
authors.initials: M
authors.lastname: Sippach
authors.forename: Philipp
authors.initials: P
authors.lastname: Bartsch
authors.forename: Heinz-Jürgen
authors.initials: HJ
authors.lastname: Steinhoff
authors.forename: Richard
authors.initials: R
authors.lastname: Wagner
mesh_terms: D003087:Colicins / Q000737:chemistry* / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D015640:Ion Channel Gating; D008051:Lipid Bilayers / Q000737:chemistry*; D008081:Liposomes / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D018613:Microscopy, Confocal / Q000379:methods*; D052899:Pore Forming Cytotoxic Proteins / Q000737:chemistry*; D011487:Protein Conformation; D055503:Protein Multimerization; D012995:Solubility
chemical_list: D003087:Colicins; D008051:Lipid Bilayers; D008081:Liposomes; D008565:Membrane Proteins; D052899:Pore Forming Cytotoxic Proteins