Fast-tumbling bicelles constructed from native Escherichia coli lipids

Abstract

Solution-state NMR requires small membrane mimetic systems to allow for acquiring high-resolution data. At the same time these mimetics should faithfully mimic biological membranes. Here we characterized two novel fast-tumbling bicelle systems with lipids from two Escherichia coli strains. While strain 1 (AD93WT) contains a characteristic E. coli lipid composition, strain 2 (AD93-PE) is not capable of synthesizing the most abundant lipid in E. coli, phosphatidylethanolamine. The lipid and acyl chain compositions were characterized by (31)P and (13)C NMR. Depending on growth temperature and phase, the lipid composition varies substantially, which means that the bicelle composition can be tuned by using lipids from cells grown at different temperatures and growth phases. The hydrodynamic radii of the bicelles were determined from translational diffusion coefficients and NMR spin relaxation was measured to investigate lipid properties in the bicelles. We find that the lipid dynamics are unaffected by variations in lipid composition, suggesting that the bilayer is in a fluid phase under all conditions investigated here. Backbone glycerol carbons are the most rigid positions in all lipids, while head-group carbons and the first carbons of the acyl chain are somewhat more flexible. The flexibility increases down the acyl chain to almost unrestricted motion at its end. Carbons in double bonds and cyclopropane moieties are substantially restricted in their motional freedom. The bicelle systems characterized here are thus found to faithfully mimic E. coli inner membranes and are therefore useful for membrane interaction studies of proteins with E. coli inner membranes by solution-state NMR.

Phase Transitions in Small Isotropic Bicelles

“…In contrast, solutions with small particles which are not capable of spontaneous orientation are called isotropic bicelles (IsoBs). IsoBs have several features that make them an almost perfect membrane mimetic for structural studies of membrane proteins. − Unlike detergent micelles, bicelles contain lipids that form the plane patch of membrane and properly mimic some properties of the lipid bilayer. , The lipid composition of bicelles can be varied to simulate the membranes of different cells or their microdomains . Cholesterol, sphingolipids, glycerolipids, lipids with unsaturated fatty chains, or anionic headgroups can be added to the isotropic bicelles. − IsoBs can be formed using the very mild rim-forming surfactants, such as Facades, that do not cause the unfolding of soluble globular domains of large membrane proteins .…”

“…11,12 The lipid composition of bicelles can be varied to simulate the membranes of different cells or their microdomains. 13 Cholesterol, sphingolipids, glycerolipids, lipids with unsaturated fatty chains, or anionic headgroups can be added to the isotropic bicelles. 14−18 IsoBs can be formed using the very mild rim-forming surfactants, such as Facades, 19 that do not cause the unfolding of soluble globular domains of large membrane proteins.…”

Behavior of Most Widely Spread Lipids in Isotropic Bicelles

“…First, the bilayer thickness can influence both the structure and stability of the membrane proteins due to the hydrophobic mismatch or lipophobic effects. , This parameter may be mimicked by altering the length of the acyl chains of bicelle-forming lipids. Second, the electrostatic properties of the bilayer surface are important, especially for the juxtamembrane regions of the proteins. − This parameter may be reproduced by combining the lipids with various headgroups in bicelles, or preparing the bicelles from the native lipid extracts. , Last, some proteins can interact with specific and low-abundant components of the cell membrane, such as gangliosides, cholesterol, inositol phosphates, etc. , Such compounds can be simply added to the bicellar mixtures in necessary amounts. − …”

“…13−15 This parameter may be reproduced by combining the lipids with various headgroups in bicelles, or preparing the bicelles from the native lipid extracts. 16,17 Last, some proteins can interact with specific and low-abundant components of the cell membrane, such as gangliosides, cholesterol, inositol phosphates, etc. 18,19 Such compounds can be simply added to the bicellar mixtures in necessary amounts.…”

Structure and Dynamics of Membrane Proteins and Membrane Associated Proteins with Native Bicelles from Eukaryotic Tissues

“…7,35 Initial studies have shown that lipids extracted from prokaryotes can be reconstituted into bicelles or nanodics. 36,37 Our native bicelles are reconstituted with purified lipids from eukaryotic tissues and show native protein−lipid interactions with eukaryotic systems.…”