108 results found
Pyle E, Guo C, Hofmann T, et al., 2019, Protein-Lipid Interactions Stabilize the Oligomeric State of BOR1p from Saccharomyces cerevisiae., Anal Chem
The BOR proteins are integral membrane transporters which mediate efflux of boron. Structures of two BOR family members from Arabidopsis thaliana and Saccharomyces mikitiae indicate that the proteins exist as dimers. However, it remains unclear whether dimer formation is dependent on protein-lipid interactions or whether the dimer is the functional form of the protein. Here, we used the BOR1p protein from Saccharomyces cerevisiae (ScBOR1p), recombinantly expressed in its native host, to explore these aspects of BOR transporter structure and function. Native mass spectrometry (MS) revealed that ScBOR1p isolates as a monomer in a range of detergents. Lipidomics analysis showed that ScBOR1p co-isolates with phosphatidylserine (PS), phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). Delipidation of ScBOR1p followed by addition of PS or PE causes formation of ScBOR1p dimers. Using a homology model of ScBOR1p, we identified a possible lipid binding site at the dimer interface comprising residues Arg265, Arg267, Arg480, and Arg481. A quadruple 4R/A mutant was expressed and isolated and also shown to be monomeric by native MS, and addition of PS or PE to this mutant did not reform the dimer. Functional complementation analysis revealed that the 4R/A mutant had boron efflux activity, suggesting that the ScBOR1p monomer is responsible for transport function. Taken together, these data strongly indicate that the physiological form of the ScBOR1p is the dimer and that dimer formation is dependent on association with membrane lipids.
Sadaf A, Ramos M, Mortensen JS, et al., 2019, Conformationally restricted monosaccharide-cored glycoside amphiphiles: the effect of detergent headgroup variation on membrane protein stability., ACS Chemical Biology, Vol: 14, Pages: 1717-1726, ISSN: 1554-8929
Detergents are widely used to isolate membrane proteins from lipid bilayers, but many proteins solubilized in conventional detergents are structurally unstable. Thus, there is major interest in the development of novel amphiphiles to facilitate membrane protein research. In this study, we have designed and synthesized novel amphiphiles with a rigid scyllo-inositol core, designated scyllo-inositol glycosides (SIGs). Varying the headgroup structure allowed the preparation of three sets of SIGs that were evaluated for their effects on membrane protein stability. When tested with a few model membrane proteins, representative SIGs conferred enhanced stability to the membrane proteins compared to a gold standard conventional detergent (DDM). Of the novel amphiphiles, a SIG designated STM-12 was most effective at preserving the stability of the multiple membrane proteins tested here. In addition, a comparative study of the three sets suggests that several factors, including micelle size and alkyl chain length, need to be considered in the development of novel detergents for membrane protein research. Thus, this study not only describes new detergent tools that are potentially useful for membrane protein structural study but also introduces plausible correlations between the chemical properties of detergents and membrane protein stabilization efficacy.
Ehsan M, Du Y, Mortensen JS, et al., 2019, Self-assembly behavior and application of terphenyl-cored trimaltosides for membrane-protein studies: impact of detergent hydrophobic group geometry on protein stability., Chemistry - A European Journal, ISSN: 0947-6539
Amphipathic agents are widely used in various fields including biomedical sciences. Micelle-forming detergents are particularly useful for in vitro membrane-protein characterization. As many conventional detergents are limited in their ability to stabilize membrane proteins, it is necessary to develop novel detergents to facilitate membrane-protein research. In the current study, we developed novel trimaltoside detergents with an alkyl pendant-bearing terphenyl unit as a hydrophobic group, designated terphenyl-cored maltosides (TPMs). We found that the geometry of the detergent hydrophobic group substantially impacts detergent self-assembly behavior, as well as detergent efficacy for membrane-protein stabilization. TPM-Vs, with a bent terphenyl group, were superior to the linear counterparts (TPM-Ls) at stabilizing multiple membrane proteins. The favorable protein stabilization efficacy of these bent TPMs is likely associated with a binding mode with membrane proteins distinct from conventional detergents and facial amphiphiles. When compared to n-dodecyl-β-d-maltoside (DDM), most TPMs were superior or comparable to this gold standard detergent at stabilizing membrane proteins. Notably, TPM-L3 was particularly effective at stabilizing the human β2 adrenergic receptor (β2 AR), a G-protein coupled receptor, and its complex with Gs protein. Thus, the current study not only provides novel detergent tools that are useful for membrane-protein study, but also suggests a critical role for detergent hydrophobic group geometry in governing detergent efficacy.
Das M, Du Y, Mortensen JS, et al., 2019, Trehalose-cored amphiphiles for membrane protein stabilization: importance of the detergent micelle size in GPCR stability (vol 17, pg 3249, 2019), ORGANIC & BIOMOLECULAR CHEMISTRY, Vol: 17, Pages: 4919-4920, ISSN: 1477-0520
Ehsan M, Kumar A, Mortensen JS, et al., 2019, Self-Assembly Behaviors of a Penta-Phenylene Maltoside and Its Application for Membrane Protein Study., Chem Asian J
We prepared an amphiphile with a penta-phenylene lipophilic group and a branched trimaltoside head group. This new agent, designated penta-phenylene maltoside (PPM), showed a marked tendency to self-assembly into micelles via strong aromatic-aromatic interactions in aqueous media, as evidenced by 1 H NMR spectroscopy and fluorescence studies. When utilized for membrane protein studies, this new agent was superior to DDM, a gold standard conventional detergent, in stabilizing multiple proteins long term. The ability of this agent to form aromatic-aromatic interactions is likely responsible for enhanced protein stabilization when associated with a target membrane protein.
Das M, Du Y, Mortensen JS, et al., 2019, Trehalose-cored amphiphiles for membrane protein stabilization: importance of the detergent micelle size in GPCR stability., Org Biomol Chem, Vol: 17, Pages: 3249-3257
Despite their importance in biology and medicinal chemistry, structural and functional studies of membrane proteins present major challenges. To study diverse membrane proteins, it is crucial to have the correct detergent to efficiently extract and stabilize the proteins from the native membranes for biochemical/biophysical downstream analyses. But many membrane proteins, particularly eukaryotic ones, are recalcitrant to stabilization and/or crystallization with currently available detergents and thus there are major efforts to develop novel detergents with enhanced properties. Here, a novel class of trehalose-cored amphiphiles are introduced, with multiple alkyl chains and carbohydrates projecting from the trehalose core unit are introduced. A few members displayed enhanced protein stabilization behavior compared to the benchmark conventional detergent, n-dodecyl-β-d-maltoside (DDM), for multiple tested membrane proteins: (i) a bacterial leucine transporter (LeuT), (ii) the R. capsulatus photosynthetic superassembly, and (iii) the human β2 adrenergic receptor (β2AR). Due to synthetic convenience and their favourable behaviors for a range of membrane proteins, these agents have potential for membrane protein research. In addition, the detergent property-efficacy relationship discussed here will guide future design of novel detergents.
Cecchetti C, Pyle E, Byrne B, 2019, Transporter oligomerisation: roles in structure and function, Biochemical Society Transactions, Vol: 47, Pages: 433-440, ISSN: 0300-5127
Oligomerisation is a key feature of integral membrane transporters with roles in structure, function and stability. In this review, we cover some very recent advances in our understanding of how oligomerisation affects these key transporter features, with emphasis on a few groups of transporters, including the nucleobase ascorbate transporters, neurotransmitter sodium symporters and major facilitator superfamily members.
Hussain H, Helton T, Du Y, et al., 2018, A comparative study of branched and linear mannitol-based amphiphiles on membrane protein stability, ANALYST, Vol: 143, Pages: 5702-5710, ISSN: 0003-2654
Das M, Du Y, Mortensen JS, et al., 2018, Rationally Engineered Tandem Facial Amphiphiles for Improved Membrane Protein Stabilization Efficacy, CHEMBIOCHEM, Vol: 19, Pages: 2225-2232, ISSN: 1439-4227
Pyle E, Kalli AC, Amillis S, et al., 2018, Structural lipids enable the formation of Ffnctional oligomers of the eukaryotic purine symporter UapA, Cell Chemical Biology, Vol: 25, Pages: 840-848.e4, ISSN: 2451-9456
The role of membrane lipids in modulating eukaryotic transporter assembly and function remains unclear. We investigated the effect of membrane lipids in the structure and transport activity of the purine transporter UapA from Aspergillus nidulans. We found that UapA exists mainly as a dimer and that two lipid molecules bind per UapA dimer. We identified three phospholipid classes that co-purified with UapA: phosphatidylcholine, phosphatidylethanolamine (PE), and phosphatidylinositol (PI). UapA delipidation caused dissociation of the dimer into monomers. Subsequent addition of PI or PE rescued the UapA dimer and allowed recovery of bound lipids, suggesting a central role of these lipids in stabilizing the dimer. Molecular dynamics simulations predicted a lipid binding site near the UapA dimer interface. Mutational analyses established that lipid binding at this site is essential for formation of functional UapA dimers. We propose that structural lipids have a central role in the formation of functional, dimeric UapA.
Das M, Du Y, Mortensen JS, et al., 2018, An Engineered Lithocholate-Based Facial Amphiphile Stabilizes Membrane Proteins: Assessing the Impact of Detergent Customizability on Protein Stability, CHEMISTRY-A EUROPEAN JOURNAL, Vol: 24, Pages: 9860-9868, ISSN: 0947-6539
Ehsan M, Das M, Stern V, et al., 2018, Steroid-Based Amphiphiles for Membrane Protein Study: The Importance of Alkyl Spacers for Protein Stability, CHEMBIOCHEM, Vol: 19, Pages: 1433-1443, ISSN: 1439-4227
Ehsan M, Du Y, Molist I, et al., 2018, Vitamin E-based glycoside amphiphiles for membrane protein structural studies, ORGANIC & BIOMOLECULAR CHEMISTRY, Vol: 16, Pages: 2489-2498, ISSN: 1477-0520
Membrane proteins play critical roles in a variety of cellular processes. For a detailed molecular level understanding of their biological functions and roles in disease, it is necessary to extract them from the native membranes. While the amphipathic nature of these bio-macromolecules presents technical challenges, amphiphilic assistants such as detergents serve as useful tools for membrane protein structural and functional studies. Conventional detergents are limited in their ability to maintain the structural integrity of membrane proteins and thus it is essential to develop novel agents with enhanced properties. Here, we designed and characterized a novel class of amphiphiles with vitamin E (i.e., α-tocopherol) as the hydrophobic tail group and saccharide units as the hydrophilic head group. Designated vitamin E-based glycosides (VEGs), these agents were evaluated for their ability to solubilize and stabilize a set of membrane proteins. VEG representatives not only conferred markedly enhanced stability to a diverse range of membrane proteins compared to conventional detergents, but VEG-3 also showed notable efficacy toward stabilization and visualization of a membrane protein complex. In addition to hydrophile–lipophile balance (HLB) of detergent molecules, the chain length and molecular geometry of the detergent hydrophobic group seem key factors in determining detergent efficacy for membrane protein (complex) stability.
Sadaf A, Du Y, Santillan C, et al., 2017, Dendronic trimaltoside amphiphiles (DTMs) for membrane protein study, CHEMICAL SCIENCE, Vol: 8, Pages: 8315-8324, ISSN: 2041-6520
Ehsan M, Ghani L, Du Y, et al., 2017, New penta-saccharide-bearing tripod amphiphiles for membrane protein structure studies, ANALYST, Vol: 142, Pages: 3889-3898, ISSN: 0003-2654
Pyle E, Kalli AC, Amillis S, et al., 2017, Structural lipids enable the formation of functional oligomers of the eukaryotic purine symporter UapA, Publisher: Cold Spring Harbor Laboratory
<jats:title>Abstract</jats:title><jats:p>The role of membrane lipids in modulating eukaryotic transporter structure and function remains poorly understood. We used native mass spectrometry in combination with molecular dynamics simulations and <jats:italic>in vivo</jats:italic> analyses to investigate the roles of membrane lipids in the structure and transport activity of the purine transporter, UapA, from <jats:italic>Aspergillus nidulans</jats:italic>. We revealed that UapA exists mainly as a dimer and that two lipid molecules bind per UapA dimer. We identified three classes of phospholipids: phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI) which co-purified with UapA. Delipidation of UapA caused dissociation of the dimer into individual protomers. Subsequent addition of PI or PE rescued the UapA dimer and allowed recovery of bound lipids, suggesting a central role of these lipids in stabilising the dimer. We predicted a putative lipid-binding site near the UapA dimer interface. Mutational analyses established that lipid binding at this site is essential for formation of functional UapA dimers. Our findings reveal unprecedented level of detail into the nature of UapA-lipid interactions and provide a framework for studying similar eukaryotic systems.</jats:p>
Byrne B, 2017, It takes two to transport via an elevator, CELL RESEARCH, Vol: 27, Pages: 965-966, ISSN: 1001-0602
Hussain H, Mortensen JS, Du Y, et al., 2017, Tandem malonate-based glucosides (TMGs) for membrane protein structural studies, Scientific Reports, Vol: 7, ISSN: 2045-2322
High-resolution membrane protein structures are essential for understanding the molecular basis of diverse biological events and important in drug development. Detergents are usually used to extract these bio-macromolecules from the membranes and maintain them in a soluble and stable state in aqueous solutions for downstream characterization. However, many eukaryotic membrane proteins solubilized in conventional detergents tend to undergo structural degradation, necessitating the development of new amphiphilic agents with enhanced properties. In this study, we designed and synthesized a novel class of glucoside amphiphiles, designated tandem malonate-based glucosides (TMGs). A few TMG agents proved effective at both stabilizing a range of membrane proteins and extracting proteins from the membrane environment. These favourable characteristics, along with synthetic convenience, indicate that these agents have potential in membrane protein research.
Hussain H, Du Y, Tikhonova E, et al., 2017, Resorcinarene-Based Facial Glycosides: Implication of Detergent Flexibility on Membrane-Protein Stability, CHEMISTRY-A EUROPEAN JOURNAL, Vol: 23, Pages: 6724-6729, ISSN: 0947-6539
As a membrane-mimetic system, detergent micelles are popularly used to extract membrane proteins from lipid environments and to maintain their solubility and stability in an aqueous medium. However, many membrane proteins encapsulated in conventional detergents tend to undergo structural degradation during extraction and purification, thus necessitating the development of new agents with enhanced properties. In the current study, two classes of new amphiphiles are introduced, resorcinarene-based glucoside and maltoside amphiphiles (designated RGAs and RMAs, respectively), for which the alkyl chains are facially segregated from the carbohydrate head groups. Of these facial amphiphiles, two RGAs (RGA-C11 and RGA-C13) conferred markedly enhanced stability to four tested membrane proteins compared to a gold-standard conventional detergent. The relatively high water solubility and micellar stability of the RGAs compared to the RMAs, along with their generally favourable behaviours for membrane protein stabilisation described here, are likely to be, at least in part, a result of the high conformational flexibility of these glucosides. This study suggests that flexibility could be an important factor in determining the suitability of new detergents for membrane protein studies.
Das M, Du Y, Ribeiro O, et al., 2017, Conformationally Preorganized Diastereomeric Norbornane-Based Maltosides for Membrane Protein Study: Implications of Detergent Kink for Micellar Properties, Journal of the American Chemical Society, Vol: 139, Pages: 3072-3081, ISSN: 1520-5126
Detergents are essential tools for functional and structural studies of membrane proteins. However, conventional detergents are limited in their scope and utility, particularly for eukaryotic membrane proteins. Thus, there are major efforts to develop new amphipathic agents with enhanced properties. Here, a novel class of diastereomeric agents with a preorganized conformation, designated norbornane-based maltosides (NBMs), were prepared and evaluated for their ability to solubilize and stabilize membrane proteins. Representative NBMs displayed enhanced behaviors compared to n-dodecyl-β-d-maltoside (DDM) for all membrane proteins tested. Efficacy of the individual NBMs varied depending on the overall detergent shape and alkyl chain length. Specifically, NBMs with no kink in the lipophilic region conferred greater stability to the proteins than NBMs with a kink. In addition, long alkyl chain NBMs were generally better at stabilizing membrane proteins than short alkyl chain agents. Furthermore, use of one well-behaving NBM enabled us to attain a marked stabilization and clear visualization of a challenging membrane protein complex using electron microscopy. Thus, this study not only describes novel maltoside detergents with enhanced protein-stabilizing properties but also suggests that overall detergent geometry has an important role in determining membrane protein stability. Notably, this is the first systematic study on the effect of detergent kinking on micellar properties and associated membrane protein stability.
Transporters are integral membrane proteins with central roles in the efficient movement of molecules across biological membranes. Many transporters exist as oligomers in the membrane. Depending on the individual transport protein, oligomerization can have roles in membrane trafficking, function, regulation and turnover. For example, our recent studies on UapA, a nucleobase ascorbate transporter, from Aspergillus nidulans, have revealed both that dimerization of this protein is essential for correct trafficking to the membrane and the structural basis of how one UapA protomer can affect the function of the closely associated adjacent protomer. Here we review roles of oligomerisation in a number of particularly well-studied transporters and transporter families.
Cho KH, Hariharan P, Mortensen JS, et al., 2016, Isomeric Detergent Comparison for Membrane Protein Stability: Importance of Inter-Alkyl-Chain Distance and Alkyl Chain Length, ChemBioChem, Vol: 17, Pages: 2334-2339, ISSN: 1439-4227
Membrane proteins encapsulated by detergent micelles are widely used for structural study. Because of their amphipathic property, detergents have the ability to maintain protein solubility and stability in an aqueous medium. However, conventional detergents have serious limitations in their scope and utility, particularly for eukaryotic membrane proteins and membrane protein complexes. Thus, a number of new agents have been devised; some have made significant contributions to membrane protein structural studies. However, few detergent design principles are available. In this study, we prepared meta and ortho isomers of the previously reported para-substituted xylene-linked maltoside amphiphiles (XMAs), along with alkyl chain-length variation. The isomeric XMAs were assessed with three membrane proteins, and the meta isomer with a C12 alkyl chain was most effective at maintaining solubility/stability of the membrane proteins. We propose that interplay between the hydrophile–lipophile balance (HLB) and alkyl chain length is of central importance for high detergent efficacy. In addition, differences in inter-alkyl-chain distance between the isomers influence the ability of the detergents to stabilise membrane proteins.
Bae HE, Mortensen JS, Ribeiro O, et al., 2016, Tandem neopentyl glycol maltosides (TNMs) for membrane protein stabilisation, CHEMICAL COMMUNICATIONS, Vol: 52, Pages: 12104-12107, ISSN: 1359-7345
Cho KH, Ribeiro O, Du Y, et al., 2016, Mesitylene-cored glucoside amphiphiles (MGAs) for membrane protein studies: importance of alkyl chain density in detergent efficacy., Chemistry - A European Journal, Vol: 22, ISSN: 0947-6539
Detergents serve as useful tools for membrane protein structural and functional studies. Their amphipathic nature allows detergents to associate with the hydrophobic regions of membrane proteins whilst maintaining the proteins in aqueous solution. However, widely used conventional detergents are limited in their ability to maintain the structural integrity of membrane proteins and thus there are major efforts underway to develop novel agents with improved properties. We prepared mesitylene-cored glucoside amphiphiles (MGAs) with three alkyl chains and compared these agents with previously developed xylene-linked maltoside agents (XMAs) with two alkyl chains and a conventional detergent (DDM). When these agents were evaluated for four membrane proteins including a G protein-coupled receptor (GPCR), some agents such as MGA-C13 and MGA-C14 resulted in markedly enhanced stability of membrane proteins compared to both DDM and the XMAs. This favourable behaviour is due likely to the increased hydrophobic density provided by the extra alkyl chain. Thus, this study not only describes new glucoside agents with potential for membrane protein research, but also introduces a new detergent design principle for future development.
Das M, Du Y, Mortensen JS, et al., 2016, Butane-1,2,3,4-tetraol-based amphiphilic stereoisomers for membrane protein study: importance of chirality in the linker region, CHEMICAL SCIENCE, Vol: 8, Pages: 1169-1177, ISSN: 2041-6520
Boulet-Audet M, Kazarian SG, Byrne B, 2016, In-column ATR-FTIR spectroscopy to monitor affinity chromatography purification of monoclonal antibodies, Scientific Reports, Vol: 6, ISSN: 2045-2322
In recent years many monoclonal antibodies (mAb) have entered the biotherapeutics market, offering new treatments for chronic and life-threatening diseases. Protein A resin captures monoclonal antibody (mAb) effectively, but the binding capacity decays over repeated purification cycles. On an industrial scale, replacing fouled Protein A affinity chromatography resin accounts for a large proportion of the raw material cost. Cleaning-in-place (CIP) procedures were developed to extend Protein A resin lifespan, but chromatograms cannot reliably quantify any remaining contaminants over repeated cycles. To study resin fouling in situ, we coupled affinity chromatography and Fourier transform infrared (FTIR) spectroscopy for the first time by embedding an attenuated total reflection (ATR) sensor inside a micro-scale column while measuring the UV 280 nm and conductivity. Our approach quantified the in-column protein concentration in the resin bed and determined protein conformation. Our results showed that Protein A ligand leached during CIP. We also found that host cell proteins bound to the Protein A resin even more strongly than mAbs and that typical CIP conditions do not remove all fouling contaminants. The insights derived from in-column ATR-FTIR spectroscopy monitoring could contribute to mAb purification quality assurance as well as guide the development of more effective CIP conditions to optimise resin lifespan.
Carlsson E, Thwaite JE, Jenner DC, et al., 2016, Bacillus anthracis TIR Domain-Containing Protein Localises to Cellular Microtubule Structures and Induces Autophagy, PLOS One, Vol: 11, ISSN: 1932-6203
Toll-like receptors (TLRs) recognise invading pathogens and mediate downstream immune signalling via Toll/IL-1 receptor (TIR) domains. TIR domain proteins (Tdps) have been identified in multiple pathogenic bacteria and have recently been implicated as negative regulators of host innate immune activation. A Tdp has been identified in Bacillus anthracis, the causative agent of anthrax. Here we present the first study of this protein, designated BaTdp. Recombinantly expressed and purified BaTdp TIR domain interacted with several human TIR domains, including that of the key TLR adaptor MyD88, although BaTdp expression in cultured HEK293 cells had no effect on TLR4- or TLR2- mediated immune activation. During expression in mammalian cells, BaTdp localised to microtubular networks and caused an increase in lipidated cytosolic microtubule-associated protein 1A/1B-light chain 3 (LC3), indicative of autophagosome formation. In vivo intra-nasal infection experiments in mice showed that a BaTdp knockout strain colonised host tissue faster with higher bacterial load within 4 days post-infection compared to the wild type B. anthracis. Taken together, these findings indicate that BaTdp does not play an immune suppressive role, but rather, its absence increases virulence. BaTdp present in wild type B. anthracis plausibly interact with the infected host cell, which undergoes autophagy in self-defence.
Byrne B, Alguel Y, Scull NJ, et al., 2016, Structure of eukaryotic purine/Hþ symporter UapA suggests a role for homodimerization in transport activity, Nature Communications, Vol: 7, ISSN: 2041-1723
The uric acid/xanthine H+ symporter, UapA, is a high-affinity purine transporter from the filamentous fungus Aspergillus nidulans. Here we present the crystal structure of a genetically stabilized version of UapA (UapA-G411VΔ1–11) in complex with xanthine. UapA is formed from two domains, a core domain and a gate domain, similar to the previously solved uracil transporter UraA, which belongs to the same family. The structure shows UapA in an inward-facing conformation with xanthine bound to residues in the core domain. Unlike UraA, which was observed to be a monomer, UapA forms a dimer in the crystals with dimer interactions formed exclusively through the gate domain. Analysis of dominant negative mutants is consistent with dimerization playing a key role in transport. We postulate that UapA uses an elevator transport mechanism likely to be shared with other structurally homologous transporters including anion exchangers and prestin.
Hussain H, Du Y, Scull NJ, et al., 2016, Accessible mannitol-based amphiphiles (MNAs) for membrane protein solubilisation and stabilisation, Chemistry, Vol: 22, ISSN: 0861-9255
Integral membrane proteins are amphipathic molecules crucial for all cellular life. The structural study of these macromolecules starts with protein extraction from the native membranes, followed by purification and crystallisation. Detergents are essential tools for these processes, but detergent-solubilised membrane proteins often denature and aggregate, resulting in loss of both structure and function. In this study, a novel class of agents, designated mannitol-based amphiphiles (MNAs), were prepared and characterised for their ability to solubilise and stabilise membrane proteins. Some of MNAs conferred enhanced stability to four membrane proteins including a G protein-coupled receptor (GPCR), the β2 adrenergic receptor (β2AR), compared to both n-dodecyl-d-maltoside (DDM) and the other MNAs. These agents were also better than DDM for electron microscopy analysis of the β2AR. The ease of preparation together with the enhanced membrane protein stabilisation efficacy demonstrates the value of these agents for future membrane protein research.
Ehsan M, Du Y, Scull NJ, et al., 2016, Highly branched pentasaccharide-bearing amphiphiles for membrane protein studies, Journal of the American Chemical Society, Vol: 138, Pages: 3789-3796, ISSN: 1520-5126
Detergents are essential tools for membrane protein manipulation. Micelles formed by detergent molecules have the ability to encapsulate the hydrophobic domains of membrane proteins. The resulting protein–detergent complexes (PDCs) are compatible with the polar environments of aqueous media, making structural and functional analysis feasible. Although a number of novel agents have been developed to overcome the limitations of conventional detergents, most have traditional head groups such as glucoside or maltoside. In this study, we introduce a class of amphiphiles, the PSA/Es with a novel highly branched pentasaccharide hydrophilic group. The PSA/Es conferred markedly increased stability to a diverse range of membrane proteins compared to conventional detergents, indicating a positive role for the new hydrophilic group in maintaining the native protein integrity. In addition, PDCs formed by PSA/Es were smaller and more suitable for electron microscopic analysis than those formed by DDM, indicating that the new agents have significant potential for the structure–function studies of membrane proteins.
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