title: Influence of calcium on direct incorporation of membrane proteins into in-plane lipid bilayer.
abstract: Reconstitution of transmembrane proteins by direct incorporation into supported lipid bilayers (SLBs) is a new method to provide suitable samples for high-resolution atomic force microscopy (AFM) analysis of membrane proteins. First experiments have reported successful incorporation of proteins into detergent-destabilized SLBs. Here, we analyzed by AFM the incorporation of membrane proteins in the presence of calcium, a divalent cation functionally important for several membrane proteins. Using lipid-phase-separated membranes, we first show that calcium strongly stabilizes the SLBs decreasing the insertion of low cmc detergents, dodecyl-beta-maltoside, dodecyl-beta-thiomaltoside, and N-hexadecylphosphocholine (Fos-Choline-16) and further insertion of proteins. However, high yield of protein insertion is recovered in the presence of calcium by increasing the detergent concentration in the solution. These data revealed the importance of the calcium in the structure of SLBs and provided new insights into the mechanism of protein insertion into these model membranes.
authors.affiliation: Institut Curie, UMR-CNRS 168 and LRC-CEA 34V, Paris Cedex 05, France.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Berquand
authors.forename: Daniel
authors.initials: D
authors.lastname: Lévy
authors.forename: Francesca
authors.initials: F
authors.lastname: Gubellini
authors.forename: Christian
authors.initials: C
authors.lastname: Le Grimellec
authors.forename: Pierre-Emmanuel
authors.initials: PE
authors.lastname: Milhiet
mesh_terms: D002118:Calcium / Q000494:pharmacology*; D003902:Detergents / Q000494:pharmacology; D008051:Lipid Bilayers / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry* / Q000648:ultrastructure; D018625:Microscopy, Atomic Force / Q000379:methods*
chemical_list: D003902:Detergents; D008051:Lipid Bilayers; D008565:Membrane Proteins; D002118:Calcium