Integral membrane proteins (IMPs) play crucial roles in many aspects of biology by mediating the transfer of material and signals between cells and their environment. It is estimated that 20-30% of all open reading frames in the human genome encode membrane proteins, and that more than 50% of current pharmaceutical agents target IMPs1
. Our understanding of IMP structure and function, however, is hampered by difficulties associated with handling these proteins2
. Most IMPs are not soluble in aqueous buffer because they display large hydrophobic surfaces when properly folded; therefore, detergents are required to extract IMPs from the lipid bilayer and to maintain the native state in solution. Mild detergents are widely used for IMP manipulation, but many membrane proteins solubilized with these agents tend to denature and/or aggregate3
, making it difficult to conduct functional studies, spectroscopic analysis or crystallization trials.
Prior efforts to develop amphiphiles tailored for IMP applications have involved diverse strategies and achieved varying levels of success. Several peptide-based designs have been explored (peptitergents4
, lipopeptide detergents5
, short peptide surfactants6
), but so far have not gained broad acceptance. Amphiphilic polymers (“amphipols”7,8
) and discoidal lipid bilayers stabilized by an amphiphilic protein scaffold (“nanodiscs”9,10
) have proven to be versatile tools for studying IMPs in native-like states in aqueous solution. It is not clear, however, whether either of these approaches can yield high-quality crystals for diffraction analysis, a prominent objective of IMP studies. Furthermore, neither amphipols nor nanodiscs are designed to extract IMPs from biological membranes. Recently reported agents of low molecular weight, such as hemifluorinated surfactants (HFS)8,11
and cholic acid-based amphiphiles12
, have displayed promising properties, but the scope of their utility remains to be explored. Thus there has been a need for amphiphiles that can extract, stabilize, and promote crystallization of IMPs more effectively than do current detergents. Amphiphiles with this combination of capabilities would have to be easily prepared on a large scale, which would be extremely challenging for peptide- or protein-based agents.
Here we report a class of amphiphiles that display favorable behavior with a diverse set of membrane proteins. The design of these amphiphiles features a central quaternary carbon, which is intended to place subtle restraints on conformational flexibility13-15
. Since the quaternary carbon was derived from neopentyl glycol and since the hydrophilic groups in the examples discussed here are derived from maltose, we designate these compounds M
lycol (MNG) amphiphiles. The quaternary carbon distinguishes MNG architecture from conventional detergent structures and enables the incorporation of two hydrophilic and two lipophilic subunits. We hypothesized that the modulation of flexibility and distinctive orientations of hydrophilic and lipophilic surfaces would cause MNG amphiphiles to display properties distinct from those of analogous conventional detergents. These amphiphiles are also readily synthesized. We have evaluated their performance with multiple membrane proteins in diverse applications, including maintenance of native IMP folding, association and function, extraction from a native membrane, growth of high-quality crystals, and support of cell-free translation.