Abstract:Macrocycles made of cholate building blocks were previously found to transport glucose readily across lipid bilayers. In this study, an 15N, 13Cα-labeled glycine was inserted into a cyclic cholate trimer and attached at the end of a linear trimer, respectively. The isotopic labeling allowed us to use solid-state NMR spectroscopy to study the dynamics, aggregation, and depth of insertion of these compounds in lipid membranes. The cyclic compound was found to be mostly immobilized in DLPC, POPC/POPG, and POPC/PO… Show more
“…With only a moderate 1 H mixing time of 100-225 ms, many peptide–lipid cross peaks already appeared, confirming the membrane insertion of hBD-3 analogs. In contrast, if a peptide only binds membrane surface, the paramyxovirus fusion peptide in DMPC bilayers for example, no peptide–lipid cross peaks are observed even at longer mixing times up to 900 ms 43,44 .Fig. 3Site-specific depth of insertion of hBD-3 analog in POPC/POPG bilayers.…”
Section: Resultsmentioning
confidence: 99%
“…For the lipid–peptide interface, D I as 0.00125 nm 2 /ms for H 2 O and 0.0025 nm 2 /ms for CH 2 were used. These values have been widely used for antimicrobial peptides, DNA, and membrane channels 44,45,71 .…”
Human β-defensins (hBD) play central roles in antimicrobial activities against various microorganisms and in immune-regulation. These peptides perturb phospholipid membranes for function, but it is not well understood how defensins approach, insert and finally disrupt membranes on the molecular level. Here we show that hBD-3 analogs interact with lipid bilayers through a conserved surface that is formed by two adjacent loops in the solution structure. By integrating a collection of 13C, 1H and 31P solid-state NMR methods with long-term molecular dynamic simulations, we reveal that membrane-binding rigidifies the peptide, enhances structural polymorphism, and promotes β-strand conformation. The peptide colocalizes with negatively charged lipids, confines the headgroup motion, and deforms membrane into smaller, ellipsoidal vesicles. This study designates the residue-specific, membrane-bound topology of hBD-3 analogs, serves as the basis for further elucidating the function-relevant structure and dynamics of other defensins, and facilitates the development of defensin-mimetic antibiotics, antifungals, and anti-inflammatories.
“…With only a moderate 1 H mixing time of 100-225 ms, many peptide–lipid cross peaks already appeared, confirming the membrane insertion of hBD-3 analogs. In contrast, if a peptide only binds membrane surface, the paramyxovirus fusion peptide in DMPC bilayers for example, no peptide–lipid cross peaks are observed even at longer mixing times up to 900 ms 43,44 .Fig. 3Site-specific depth of insertion of hBD-3 analog in POPC/POPG bilayers.…”
Section: Resultsmentioning
confidence: 99%
“…For the lipid–peptide interface, D I as 0.00125 nm 2 /ms for H 2 O and 0.0025 nm 2 /ms for CH 2 were used. These values have been widely used for antimicrobial peptides, DNA, and membrane channels 44,45,71 .…”
Human β-defensins (hBD) play central roles in antimicrobial activities against various microorganisms and in immune-regulation. These peptides perturb phospholipid membranes for function, but it is not well understood how defensins approach, insert and finally disrupt membranes on the molecular level. Here we show that hBD-3 analogs interact with lipid bilayers through a conserved surface that is formed by two adjacent loops in the solution structure. By integrating a collection of 13C, 1H and 31P solid-state NMR methods with long-term molecular dynamic simulations, we reveal that membrane-binding rigidifies the peptide, enhances structural polymorphism, and promotes β-strand conformation. The peptide colocalizes with negatively charged lipids, confines the headgroup motion, and deforms membrane into smaller, ellipsoidal vesicles. This study designates the residue-specific, membrane-bound topology of hBD-3 analogs, serves as the basis for further elucidating the function-relevant structure and dynamics of other defensins, and facilitates the development of defensin-mimetic antibiotics, antifungals, and anti-inflammatories.
“…Membrane insertion depth of a residue corresponds to a restraint on the z i coordinate in such a frame. Such insertion depths can be obtained, for instance, by continuous‐wave EPR power saturation measurements, electron spin echo envelope modulation (ESEEM) measurements of water accessibility, or solid‐state NMR spin diffusion measurements . They can be specified as restraints on either the Cα atom coordinate or the spin label coordinate.…”
Conformational ensembles of intrinsically disordered peptide chains are not fully determined by experimental observations. Uncertainty due to lack of experimental restraints and due to intrinsic disorder can be distinguished if distance distributions restraints are available. Such restraints can be obtained from pulsed dipolar electron paramagnetic resonance (EPR) spectroscopy applied to pairs of spin labels. Here, we introduce a Monte Carlo approach for generating conformational ensembles that are consistent with a set of distance distribution restraints, backbone dihedral angle statistics in known protein structures, and optionally, secondary structure propensities or membrane immersion depths. The approach is tested with simulated restraints for a terminal and an internal loop and for a protein with 69 residues by using sets of sparse restraints for underlying well-defined conformations and for published ensembles of a premolten globule-like and a coil-like intrinsically disordered protein.
“…Similar conclusions were drawn in a solid-state NMR study. 62 With the pore-forming mechanism firmly established, we became interested in using additional noncovalent interactions to tune the stacking. The idea was that, once we learned to control the stacking, we would be able to open and close the pore on demand.…”
Controlled translocation of molecules and ions across lipid membranes is the basis of numerous biological functions. Because synthetic systems can help researchers understand the more complex biological ones, many chemists have developed synthetic mimics of biological transporters. Both systems need to deal with similar fundamental challenges. In addition to providing mechanistic insights into transport mechanisms, synthetic transporters are useful in a number of applications including separation, sensing, drug delivery, and catalysis.
Disciplines
Chemistry
CommentsReprinted (adapted) C O N S P E C T U S C ontrolled translocation of molecules and ions across lipid membranes is the basis of numerous biological functions. Because synthetic systems can help researchers understand the more complex biological ones, many chemists have developed synthetic mimics of biological transporters. Both systems need to deal with similar fundamental challenges. In addition to providing mechanistic insights into transport mechanisms, synthetic transporters are useful in a number of applications including separation, sensing, drug delivery, and catalysis.In this Account, we present several classes of membrane transporters constructed in our laboratory from a facially amphiphilic building block, cholic acid. Our "molecular baskets" can selectively shuttle glucose across lipid membranes without transporting smaller sodium ions. We have also built oligocholate foldamers that transiently fold into helices with internal hydrophilic binding pockets to transport polar guests. Lastly, we describe amphiphilic macrocycles, which form transmembrane nanopores in lipid bilayers through the strong associative interactions of encapsulated water molecules.In addition to presenting the different transport properties of these oligocholate transporters, we illustrate how fundamental studies of molecular behavior in solution facilitate the creation of new and useful membrane transporters, despite the large difference between the two environments. We highlight the strong conformational effect of transporters. Because the conformation of a molecule often alters its size and shape, and the distribution of functional groups, conformational control can be used rationally to tune the property of a transporter. Finally, we emphasize that, whenever water is the solvent, its unique properties;small size, strong solvation for ionic functionalities, and an extraordinary cohesive energy density (i.e., total intermolecular interactions per unit volume);tend to become critical factors to be considered. Purposeful exploitation of these solvent properties may be essential to the success of the supramolecular process involved;this is also the reason for the "learning through water play" in the title of this Account.
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