Tripod Amphiphiles for Membrane Protein Manipulation

Department of Chemistry, University of Wisconsin-Madison, 1101 University Avenue, Madison, WI 53706, USA. .
Molecular BioSystems (Impact Factor: 3.21). 12/2009; 6(1):89-94. DOI: 10.1039/b915162c
Source: PubMed


Integral membrane proteins (IMPs) are crucial biological components, mediating the transfer of material and information between cells and their environment. Many IMPs have proven to be difficult to isolate and study. High-resolution structural information on this class of proteins is limited, largely because of difficulties in generating soluble forms of such proteins that retain native folding and activity, and difficulties in generating high-quality crystals from such preparations. Isolated IMPs typically do not dissolve in aqueous solution, a property that arises from the large patches of hydrophobic surface necessary for favorable interactions with the core of a lipid bilayer. Detergents are generally required for IMP solubilization: hydrophobic segments of detergent molecules cluster around and shield from water the hydrophobic protein surfaces. The critical role played by detergents in membrane protein manipulation, and the fact that many IMPs are recalcitrant to solubilization and/or crystallization with currently available detergents, suggest that it should be valuable to explore new types of amphiphiles for these purposes. This review constitutes a progress report on our long-term effort to develop a new class of organic molecules, collectively designated "tripod amphiphiles," that are intended as alternatives to conventional detergents for membrane protein manipulation. One long-range goal of this research is to identify new types of amphiphiles that facilitate IMP crystallization. This review should help introduce an important biochemical need to organic chemists, and perhaps inspire new approaches to the problem.

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    • "This carbon in the lipophilic region limits the conformational freedom of this class of molecules, thereby making them rigid relative to conventional detergents. This rigidification likely enabled us to solve the crystal structures of several N-oxide TPAs themselves [37] and has the potential to facilitate the crystallization of a wide array of membrane proteins; for example, TPA-solubilized bR and potassium channel from Streptomyces lividans have been crystallized, although their structures have not yet been solved [34] [36]. Recent TPA advances had led to a series of molecules with bifurcated glucose headgroups with favorable solubilization and stabilization efficacy [35]. "
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    ABSTRACT: IntroductionPreparation of Expression ConstructsTransfer of Plasmid DNA to Rhodobacter via Conjugal MatingSmall-Scale Screening for Expression and Localization of Target Protein in RhodobacterLarge-Scale CultureDetergent Solubilization and Chromatographic Purification of Expressed Membrane ProteinsProtein Identification and Assessment of PurityPreparations of Specialized Rhodobacter MembranesAppendix: Media and Buffer FormulationsAbbreviationsReferences
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