Coacervation of Lipid Bilayer in Natural Cell Membranes for Extraction, Fractionation, and Enrichment of Proteins in Proteomics Studies

This is the first
report where hexafluoroisopropanol (HFIP) was
used to induce the coacervation of lipid components in natural cell
membranes that would concomitantly result in solubilization, extraction,
and enrichment of hydrophobic proteins (e.g., integral membrane proteins,
IMP) into the coacervate phase, and extraction of hydrophilic proteins
in a separate aqueous phase. The incorporation of this innovative
approach in the proteomics workflow would allow the fractionation
of proteins in separate aqueous and coacervate phases and would also
eliminate the need for using surfactants. Subsequently, proteins can
be identified by the bottom-up proteomics approach where samples were
digested in solution after phase separation. Yeast cell wall proteins,
anchored membrane proteins, and proteins related to some regulatory
activities were mostly found in the aqueous-rich phase. On the other
hand, most integral membrane proteins, proteins involved in metabolic
processes, and proteins responsible for ions or drug binding were
identified in the coacervate phase. The detergent-free, facile, and
rapid process of natural lipid coacervation increased the number of
identified proteins by 8% (vs no-phase separation experiment). The
identification of all IMPs and organelle IMPs was improved by 13%
and 29%, respectively. In addition, 25% more low-abundance proteins
(less than 20 ppm) were identified.