Aqueous polymer two‐phase systems: Effective tools for plasma membrane proteomics

Abstract

Plasma membranes (PMs) are of particular importance for all living cells. They form a selectively permeable barrier to the environment. Many essential tasks of PMs are carried out by their proteinaceous components, including molecular transport, cell-cell interactions, and signal transduction. Due to the key role of these proteins for cellular function, they take center-stage in basic and applied research. A major problem towards in-depth identification and characterization of PM proteins by modern proteomic approaches is their low abundance and immense heterogeneity in different cells. Highly selective and efficient purification protocols are hence essential to any PM proteome analysis. An effective tool for preparative isolation of PMs is partitioning in aqueous polymer two-phase systems. In two-phase systems, membranes are separated according to differences in surface properties rather than size and density. Despite their rare application to the fractionation of animal tissues and cells, they represent an attractive alternative to conventional fractionation protocols. Here, we review the principles of partitioning using aqueous polymer two-phase systems and compare aqueous polymer two-phase systems with other methods currently used for the isolation of PMs.

Dissecting the Subcellular Compartmentation of Proteins and Metabolites in Arabidopsis Leaves Using Non-aqueous Fractionation

“…This could be done by applying orthogonal separation methods. For example, phase-partitioning-based separation has been successfully used in parallel with density-based separation to separate mitochondria from thylakoids (65), and two-phase partitioning is a common method for membrane separations (66). Otherwise, in order to further refine the separation based on density, one possibility would be to alter the ratio of the two non-polar solvents tetrachloroethylene and heptane, and therefore the range of the density along the gradient.…”

Simulation of polyethylene glycol and calcium-mediated membrane fusion

“…To do so, we simulated ionic solutions of PEG chains. For an electrolyte solution dissolved in a binary solvent, ions induce a phase separation when the ion solvation energy for a specific solvent overcomes the entropy of mixing 21 (low in polymer solutions because of chain connectivity). An initial homogeneous water/polymer mixture thus separates into coexisting fluid phases (so-called salting-out effect).…”

Platelet membrane proteomics: a novel repository for functional research

“…A common system is the polyethyleneglycol (PEG)/ dextran system, where plasma membranes show the highest affinity for the more hydrophobic upper PEG phase in comparison with other membrane vesicles. 8 We already successfully used this technique for analysis of plasma membrane N-glycosylation sites on human platelets. 9,10 In contrast, the current study intends to provide a general overview of membrane proteins within these preparations to allow for a more global assessment of the membrane protein composition.…”