9 research outputs found
An acid-compatible co-polymer for the solubilization of membranes and proteins into lipid bilayer-containing nanoparticles
The fundamental importance of membrane proteins in drug discovery has meant that membrane mimetic systems for studying membrane proteins are of increasing interest. One such system has been the amphipathic, negatively charged poly(styrene-co-maleic acid) (SMA) polymer to form âSMA Lipid Particlesâ (SMALPs) which have been widely adopted to solubilize membrane proteins directly from the cell membrane. However, SMALPs are only soluble under basic conditions and precipitate in the presence of divalent cations required for many downstream applications. Here, we show that the positively charged poly(styrene-co-maleimide) (SMI) forms similar nanoparticles with comparable efficiency to SMA, whilst remaining functional at acidic pH and compatible with high concentrations of divalent cations. We have performed a detailed characterization of the performance of SMI that enables a direct comparison with similar data published for SMA. We also demonstrate that SMI is capable of extracting proteins directly from the cell membrane and can solubilize functional human G-protein coupled receptors (GPCRs) expressed in cultured HEK 293T cells. âSMILPsâ thus provide an alternative membrane solubilization method that successfully overcomes some of the limitations of the SMALP method
Evidence of Lipid Exchange in Styrene Maleic Acid Lipid Particle (SMALP) Nanodisc Systems
This document is the Accepted Manuscript version of a Published Work that appeared in final form in Langmuir, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acs.langmuir.6b02927Styrene-alt-maleic Acid lipid particles (SMALPs) are self-assembled discoidal structures composed of a polymer belt and a segment of lipid bilayer, which are capable of encapsulating membrane proteins directly from the cell membrane. Here we present evidence of the exchange of lipids between such ânanodiscsâ and lipid monolayers adsorbed at either solid-liquid or air-liquid interfaces. This behavior has important implications for the potential uses of nanodiscs, including the potential to control lipid composition within nanodiscs containing membrane protein
The Interaction of Styrene Maleic Acid Copolymers with Phospholipids in Langmuir Monolayers, Vesicles and Nanodiscs; a Structural Study
Hypothesis Self-assembly of amphipathic styrene maleic acid copolymers with phospholipids in aqueous solution results in the formation of ânanodiscsâ containing a planar segment of phospholipid bilayer encapsulated by a polymer belt. Recently, studies have reported that lipids rapidly exchange between both nanodiscs in solution and external sources of lipids. Outstanding questions remain regarding details of polymer-lipid interactions, factors influencing lipid exchange and structural effects of such exchange processes. Here, the dynamic behaviour of nanodiscs is investigated, specifically the role of membrane charge and polymer chemistry. Experiments Two model systems are investigated: fluorescently labelled phospholipid vesicles, and Langmuir monolayers of phospholipids. Using fluorescence spectroscopy and time-resolved neutron reflectometry, the membrane potential, monolayer structure and composition are monitored with respect to time upon polymer and nanodisc interactions. Findings In the presence of external lipids, polymer chains embed throughout lipid membranes, the extent of which is governed by the net membrane charge. Nanodiscs stabilised by three different polymers will all exchange lipids and polymer with monolayers to differing extents, related to the properties of the stabilising polymer belt. These results demonstrate the dynamic nature of nanodiscs which interact with the local environment and are likely to deposit both lipids and polymer at all stages of use
Influence of Poly(styrene-<i>co</i>-maleic acid) Copolymer Structure on the Properties and Self-Assembly of SMALP Nanodiscs
Polymer
stabilized nanodiscs are self-assembled structures composed
of a polymer belt that wraps around a segment of lipid bilayer, and
as such are capable of encapsulating membrane proteins directly from
the cell membrane. To date, most studies on these nanodiscs have used
polyÂ(styrene-<i>co</i>-maleic acid) (SMA) with the term
SMA-lipid particles (SMALPs) coined to describe them. In this study,
we have determined the physical and thermodynamic properties of such
nanodiscs made with two different SMA copolymers. These include a
widely used and commercially available statistical polyÂ(styrene-<i>co</i>-maleic acid) copolymer (coSMA) and a reversible additionâfragmentation
chain transfer synthesized copolymer with narrow molecular weight
distribution and alternating styrene and maleic acid groups with a
polystyrene tail, (altSMA). We define phase diagrams for each polymer,
and show that, regardless of polymer topological structure, self-assembly
is driven by the free energy change associated with the polymers.
We also show that nanodisc size is polymer dependent, but can be modified
by varying polymer concentration. The thermal stability of each nanodisc
type is similar, and both can effectively solubilize proteins from
the <i>E. coli</i> membrane. These data show the potential
for the development of different SMA polymers with controllable properties
to produce nanodiscs that can be optimized for specific applications
and will enable more optimized and widespread use of the SMA-based
nanodiscs in membrane protein research