Rapid and Sequential Dual Oxime Ligation Enables De Novo Formation of Functional Synthetic Membranes from Water-Soluble Precursors.

Cell membranes define the boundaries of life and primarily consist of phospholipids. Living organisms assemble phospholipids by enzymatically coupling two hydrophobic tails to a soluble polar head group. Previous studies have taken advantage of micellar assembly to couple single-chain precursors, forming non-canonical phospholipids. However, biomimetic nonenzymatic coupling of two alkyl tails to a polar head-group remains challenging, likely due to the sluggish reaction kinetics of the initial coupling step. Here we demonstrate rapid de novo formation of biomimetic liposomes in water using dual oxime bond formation between two alkyl chains and a phosphocholine head group. Membranes can be generated from non-amphiphilic, water-soluble precursors at physiological conditions using micromolar concentrations of precursors. We demonstrate that functional membrane proteins can be reconstituted into synthetic oxime liposomes from bacterial extracts in the absence of detergent-like molecules.

1-nm-Wide Hydrated Dipole Arrays Regulate AuNW Assembly on Striped Monolayers in Nonpolar Solvent

A hallmark of biological membranes is the dynamic localization of lipids and proteins. Lipids respond to temperature reduction below a critical point with phase separation, and poikilothermic animals and also bacteria adapt their lipid content to prevent gel phase formation in membranes. In a new study, Gohrbandt etal (2022) show that reduced membrane fluidity in bacterial cells causes reversible phase separation without membrane rupture invivo, highlighting the physical robustness of biological membranes.

The solution structure of the phosphocholine (PC) head group in 1,2-dipropionyl-sn-glycero-3-phosphocholine (C(3)-PC) in 30 mol. % dimethylsulfoxide (DMSO)-water solutions has been determined by using neutron diffraction enhanced with isotopic substitution in combination with computer simulation techniques. By investigating the atomic scale hydration structure around the PC head group, a unique description of the displacement of water molecules by DMSO molecules is detailed around various locations of the head group. Specifically, DMSO molecules were found to be the most prevalent around the onium portion of the head group, with the dipoles of the DMSO molecules being aligned where the negatively charged oxygen can interact strongly with the positively charged lipid group. The phosphate group is also partially dehydrated by the presence of the DMSO molecules. However, around this group the bulkier positive end of the DMSO dipole is interacting with negatively charged groups of the lipid head group, the DMSO layer shows no obvious ordering as it cannot form hydrogen bonds with the oxygen atoms in the PO(4) group such as water molecules can. Interestingly, DMSO-water contacts have also increased in the presence of the lipid molecule relative to DMSO-water contacts observed in pure DMSO/water solutions at similar concentrations.

The lipid bilayer is a dynamic environment that consists of a mixture of lipids with different properties that regulate the function of membrane proteins; these lipids are either annular, masking the protein hydrophobic surface, or specific lipids, essential for protein function. In this study, using tandem mass spectrometry, we have identified specific lipids associated with the Escherichia coli ABC transporter McjD, which translocates the antibacterial peptide MccJ25. Using non-denaturing mass spectrometry, we show that McjD in complex with MccJ25 survives the gas phase. Partial delipidation of McjD resulted in reduced ATPase activity and thermostability as shown by circular dichroism, both of which could be restored upon addition of defined E. coli lipids. We have resolved a phosphatidylglycerol lipid associated with McjD at 3.4 Å resolution, whereas molecular dynamic simulations carried out in different lipid environments assessed the binding of specific lipids to McjD. Combined, our data show a synergistic effect of zwitterionic and negatively charged lipids on the activity of McjD; the zwitterionic lipids provide structural stability to McjD, whereas the negatively charged lipids are essential for its function.

The nanofiber formation in aqueous suspension of two classes of symmetric single-chain bolaamphiphiles with different polar headgroups and a diacetylene-modified alkyl chain with a length of 32, 34, and 36 C atoms was investigated by differential scanning calorimetry, transmission electron microscopy, and small-angle neutron scattering. As observed before for other bolalipids with phosphocholine (PC) and dimethyl-phosphoethanolamine (Me2PE) headgroups, the molecules form fibers when suspended in water at low temperatures but disassemble into micellar-like aggregates upon heating. The introduction of a diacetylene group in the middle of the long chain leads to a perturbation of chain packing so that this fiber-micelle transition occurs at lower temperature compared to the other bolalipids having unmodified alkyl chains. The aim of our project was the introduction of diacetylene groups into alkyl chains to be able to polymerize the fibers at low temperature. This should enhance the fiber stability and prevent the disassembly into micellar aggregates at higher temperature. Polymerization of aggregates containing diacetylene-modified bolaamphiphiles can be easily traced by UV/vis spectroscopy as colored products are formed. We found that polymerization of bolaamphiphiles with PC headgroups leads to a breakdown of most fibers into micellelike aggregates, and only some longer fibers segments are still detectable. In contrast, the use of Me2PE headgroups improves polymerizability and length of the polymerized fibers. The compound with 36 C atoms in the chain could be polymerized at low temperatures, and the fibers remained stable at least up to a temperature of 60 °C. This shows that the perturbation of the chain packing due to the diacetylene groups in the chains can be overcome by elongation of the chains, so that thermostable fibers with a diameter of the length of the bolalipid molecule can be successfully formed.

Mixing behaviour of asymmetrical glycerol diether bolalipids with saturated and unsaturated phosphatidylcholines

Various amphiphilic fullerene derivatives were prepared by functionalization of [5,6]fullerene-C60-Ih (C60) with malonate or bis-malonate derivatives obtained by esterification of the malonic acid mono-esters 5–7. Cyclopropafullerene 10 was obtained by protection of the carboxylic acid function of 6 as a tert-butyl ester, followed by Bingel addition to C60 and a deprotection step (Scheme 2). The preparation of 10 was also attempted directly from the malonic acid mono-ester 6 under Bingel conditions. Surprisingly, the corresponding 3′-iodo-3′H-cyclopropa[1,9][5,6]fullerene-C60-Ih-3′-carboxylate 11 was formed instead of 10 (Scheme 3). The general character of this new reaction was confirmed by the preparation of 15 and 16 from the malonic acid mono-esters 13 and 14, respectively (Scheme 4). All the other amphiphilic fullerene derivatives were prepared by taking advantage of the versatile regioselective reaction developed by Diederich and co-workers which led to macrocyclic bis-adducts of C60 by a cyclization reaction at the C-sphere with bis-malonate derivatives in a double Bingel cyclopropanation. The bis-adducts 37–39 with a carboxylic acid polar head group and four pendant long alkyl chains of different length were prepared from diol 22 and acids 5–7, respectively (Scheme 9). In addition, the amphiphilic fullerene derivatives 45, 46, 49, 54, and 55 bearing different polar head groups and compound 19 with no polar head group were synthesized (Schemes 11–13, 15, and 5, resp.). The ability of all these compounds to form Langmuir monolayers at the air-water interface was investigated in a systematic study. The films at the water surface were characterized by their surface pressure vs. molecular area isotherms, compression and expansion cycles, and Brewster-angle microscopy. The spreading behavior of compound 10 was not good, the two long alkyl chains in 10 being insufficient to prevent aggregation resulting from the strong fullerene-fullerene interactions. While no films could be obtained from compound 19 with no polar head group, all the corresponding amphiphilic fullerene bis-adducts showed good spreading characteristics and reversible behavior upon successive compression/expansion cycles. The encapsulation of the fullerene in a cyclic addend surrounded by four long alkyl chains is, therefore, an efficient strategy to prevent the irreversible aggregation resulting from strong fullerene-fullerene interactions usually observed for amphiphilic C60 derivatives at the air-water interface. The balance of hydrophobicity to hydrophilicity was modulated by changing the length of the surrounding alkyl chains or the nature of the polar head group. The best results in terms of film formation and stability were obtained with the compounds having the largest polar head group, i.e. 45 and 46, and dodecyl chains. Finally, the Langmuir films obtained from the amphiphilic fullerene bis-adducts were transferred onto solid substrates, yielding high-quality Langmuir-Blodgett films.

The structure of a zwitterionic phosphocholine (PC) surfactant monolayer adsorbed on the surface of water has been determined using neutron reflectivity in combination with H/D isotopic substitution. The most significant results of this study are the level of hydration of the PC headgroup and the lack of dehydration with increasing temperature and salt addition. The fraction of the alkyl chain (f(c)) immersed in water for all three chain isomers studied was found to be around 0.15, suggesting that the PC headgroup geometries influenced not only the headgroup hydration but also the degree of immersion of the alkyl chain in water. At the critical micelle concentration (CMC), the number of water molecules associated with the PC headgroup in C(m)PC (m = 12, 14, 16) was on order of 15. This value was significantly greater than that obtained for nonionic and ionic surfactants with similar limiting area per molecule at the CMC (A(cmc)). However, the fraction of the chain immersed in water for the ionic and nonionic surfactants was much greater. This suggests that the unique surface biocompatibility of PC surfactants arises from their strong affinity for water, and the relatively low fraction of mixing with the alkyl chain arises from the higher structural order within the PC monolayer. As surface coverage decreased, the number of water molecules associated with each PC headgroup increased, but f(c) remained constant for all the surfactants. This observation was consistent with the small variation in the thickness of the headgroup region, and the entire layer changed little with surfactant concentration. This is attributed to the role of PC headgroup geometries to maintain the conformational order within the layer as packing density varies. Further structural analysis based on a kinematic approach showed that, as the chain length was increased from C12 to C14 to C16 at the CMC, the angle of tilt for the alkyl chain increased from 40 degrees to 48 degrees to 53 degrees , respectively, whereas the thickness of the whole layer and that of the PC head region was largely constant. The almost vertical projection of the PC headgroup from these single alkyl chain surfactants is in sharp contrast to its strongly tilted conformation, as reported for dichain phospholipids such as dipalmitoyl glycerol phosphocholine (DPPC).

Membrane protein function is regulated by the lipid bilayer composition. In many cases the changes in function correlate with changes in the lipid intrinsic curvature (c0), and c0 is considered a determinant of protein function. Yet, water-soluble amphiphiles that cause either negative or positive changes in curvature have similar effects on membrane protein function, showing that changes in lipid bilayer properties other than c0 are important—and may be dominant. To further investigate the mechanisms underlying the bilayer regulation of protein function, we examined how maneuvers that alter phospholipid head groups effective “size”—and thereby c0—alter gramicidin (gA) channel function. Using dioleoylphospholipids and planar bilayers, we varied the head groups’ physical volume and the electrostatic repulsion among head groups (and thus their effective size). When 1,2-dioleyol-sn-glycero-3-phosphocholine (DOPC), was replaced by 1,2-dioleyol-sn-glycero-3-phosphoethanolamine (DOPE) with a smaller head group (causing a more negative c0), the channel lifetime (τ) is decreased. When the pH of the solution bathing a 1,2-dioleyol-sn-glycero-3-phosphoserine (DOPS) bilayer is decreased from 7 to 3 (causing decreased head group repulsion and a more negative c0), τ is decreased. When some DOPS head groups are replaced by zwitterionic head groups, τ is similarly decreased. These effects do not depend on the sign of the change in surface charge. In DOPE:DOPC (3:1) bilayers, pH changes from 5→9 to 5→0 (both increasing head group electrostatic repulsion, thereby causing a less negative c0) both increase τ. Nor do the effects depend on the use of planar, hydrocarbon-containing bilayers, as similar changes were observed in hydrocarbon-free lipid vesicles. Altering the interactions among phospholipid head groups may alter also other bilayer properties such as thickness or elastic moduli. Such changes could be excluded using capacitance measurements and single channel measurements on gA channels of different lengths. We conclude that changes gA channel function caused by changes in head group effective size can be predicted from the expected changes in c0.

On how hydrogen bonds affect foam stability

A shotgun lipidomics approach that allowed the analysis of eight lipid classes directly from crude extracts of the soil bacterium Sinorhizobium meliloti is presented. New MS-MS transitions are reported for the analysis of monomethylphosphatidylethanolamines, dimethylphosphatidylethanolamines, and three bacterial non-phosphorus-containing lipid classes [sulfoquinovosyldiacylglycerols, ornithines, and diacylglyceryl-(N,N,N-trimethyl)-homoserines]. Unique MS-MS transitions allowed the analysis of isomeric species from various lipid classes without chromatography. Analyses required small sample amounts and minimal preparation; thus, this methodology has excellent potential to be used as a screening tool for the analysis of large numbers of samples in functional genomics studies. FA distributions within lipid classes of S. meliloti are described for the first time, and the relative positions of fatty acyl substituents (sn-1, sn-2) in phospholipids are presented. FA distributions in diacylglyceryl-(N,N,N-trimethyl)-homoserines were identical to those of phospholipids, indicating a common biosynthetic origin for these lipids. The method was applied to the analysis of mutants deficient in the PhoB regulator protein. Increased lipid cyclopropanation was observed in PhoB-deficient mutants under P(i) starvation.

Mouse fibroblast L-M cells were grown in tissue culture medium containing selectively deuterated choline or ethanolamine. Both compounds were incorporated into the corresponding phospholipids at levels greater than 50% thus leading to a selective deuteration of these phospholipid head groups. Choline and ethanolamine were labeled at either the alpha- or the beta-carbon atom and well-resolved deuterium and phosphorus n.m.r. spectra were obtained from intact cells, crude plasma membranes and lipid extracts, leading to the following conclusions. (i) A large fraction, if not all, of the phospholipids in the intact L-M cell membranes were organized in a liquid crystalline bilayer. (ii) The phosphoethanolamine and the phosphocholine head group conformation were found to be remarkably similar in pure lipid bilayers and in intact L-M cell membranes with the head group dipoles being oriented parallel to the membrane surface. (iii) The deuterium T1 spin lattice relaxation times fell in the range of 7-25 ms and were similar in intact L-M cells and in pure lipid model membranes, suggesting that the two head groups are not involved in strong interactions with membrane proteins. The rotational diffusion rate of the two head groups was reduced by at least a factor of 10 compared to molecules of the same size in aqueous solution. (iv) The phosphocholine head group was sensitive to the size and sign of membrane surface charges as verified in mixing experiments with charged lipids. In L-M cell membranes the phosphocholine appeared to sense an electrically neutral environment in spite of the fact that L-M cell membranes contain 10-20% negatively charged lipids.

Role of ceramide structure and its microenvironment on the conformational order of model stratum corneum lipids mixtures: an approach by FTIR spectroscopy

Surfactants are a class of chemical compounds with unique properties, structure of which is consisted of hydrophobic and hydrophilic parts. This double structure gives rise to their unique behaviors in various conditions. Bolaforms are organic molecules compounds of two charged groups which are connected via a linear poly methylen chain.Bolaforms\' surfactants don\'t work as well as Conventional surfactant (having a head group and one alkyl chain) in reducing surface tension. Bolaforms are absorbed in water – air interface, however, their effectiveness in terms of reducing water surface tension is not considerable compared to common surfactants. In this study, synthesis of two kinds of bolaforms which the same spacer length and different heads is investigated. The structural formula for synthesized kinds is as follows: bolaform A with structural formula Br-(CH3)3N+(CH2)3 N+(CH3)­3 Br – ­with methyl side branches in head group and bolaform B with structural formula of Br-(CH3 CH2)3N+(CH2)3 N+(CH3 CH2)3Br –. IR and NMR identification techniques approves obtained product. Using surface tension measurement device it can be observed that synthesized bolaforms have high CMC; in other words, bolaform\'s surfactants with short spacer play electrolytic role.