Determination of DMPC hydration in the Lα and Lβ′ phases by 2H solid state NMR of D2O

AbstractThe number of water molecules bound to the dimyristoylphosphatidylcholine (DMPC) interface was investigated both in the fluid (Lα) and gel (Lβ′) phases by solid state deuterium NMR of D2O. We determined that each DMPC molecule binds 9.7±0.5 and less than 4.3±0.5 D2O in the fluid and gel phases respectively. These results are accounted for by considering the number of DMPC binding sites as well as the molecular organization in each phase. ©1997 Federation of European Biochemical Societies

Key Role of Polyphosphoinositides in Dynamics of Fusogenic Nuclear Membrane Vesicles

The role of phosphoinositides has been thoroughly described in many signalling and membrane trafficking events but their function as modulators of membrane structure and dynamics in membrane fusion has not been investigated. We have reconstructed models that mimic the composition of nuclear envelope precursor membranes with naturally elevated amounts of phosphoinositides. These fusogenic membranes (membrane vesicle 1(MV1) and nuclear envelope remnants (NER) are critical for the assembly of the nuclear envelope. Phospholipids, cholesterol, and polyphosphoinositides, with polyunsaturated fatty acid chains that were identified in the natural nuclear membranes by lipid mass spectrometry, have been used to reconstruct complex model membranes mimicking nuclear envelope precursor membranes. Structural and dynamic events occurring in the membrane core and at the membrane surface were monitored by solid-state deuterium and phosphorus NMR. “MV1-like” (PC∶PI∶PIP∶PIP2, 30∶20∶18∶12, mol%) membranes that exhibited high levels of PtdIns, PtdInsP and PtdInsP2 had an unusually fluid membrane core (up to 20% increase, compared to membranes with low amounts of phosphoinositides to mimic the endoplasmic reticulum). “NER-like” (PC∶CH∶PI∶PIP∶PIP2, 28∶42∶16∶7∶7, mol%) membranes containing high amounts of both cholesterol and phosphoinositides exhibited liquid-ordered phase properties, but with markedly lower rigidity (10–15% decrease). Phosphoinositides are the first lipids reported to counterbalance the ordering effect of cholesterol. At the membrane surface, phosphoinositides control the orientation dynamics of other lipids in the model membranes, while remaining unchanged themselves. This is an important finding as it provides unprecedented mechanistic insight into the role of phosphoinositides in membrane dynamics. Biological implications of our findings and a model describing the roles of fusogenic membrane vesicles are proposed

Membrane structure and dynamics by 2H- and 31P-NMR. Effects of amphipatic peptidic toxins on phospholipid and biological membranes

The actions of bee venom melittin and delta-lysin from Staphylococcus aureus on membranes have been monitored by solid-state deuterium and phosphorus NMR and shown to differ depending on temperature and on the lipid-to-peptide molar ratio Ri. In the gel phase of phosphatidylcholine model membranes, for lipid-to-peptide ratios Ri greater than 15, melittin induces isotropic lines interpreted as reflecting the presence of small discoidal structures, whereas delta-lysin does not. These small objects are metastable, that is, within a time-scale of hours they return to large lipid bilayers. The kinetics of this process depend on the lecithin chain length. In the fluid phases, at temperatures greater than that of the gel-to-fluid transition Tc, analysis of the quadruplar splittings in terms of chain ordering indicates that both melittin and delta-lysin similarly disorder the membrane. At temperatures above but close to Tc, melittin preferentially orders the center of the bilayer, while delta-lysin promotes ordering throughout the entire bilayer thickness. These effects are interpreted as reflecting different locations of the peptides with respect to the membrane surface. The addition of greater amounts of toxins, Ri = 4, on phosphatidylcholine model membranes induces very small structures irrespective of the temperature in the case of melittin, but only above Tc for delta-lysin. NMR spectral features similar to those characterizing the small fast-tumbling objects with phosphatidylcholine are also observed with egg phosphatidylethanolamine and erythrocyte membranes. The formation of small structures is thus inferred as a general process which reflects membrane supramolecular reorganization

Interaction of the Neuropeptide Met-Enkephalin with Zwitterionic and Negatively Charged Bicelles as Viewed by (31)P and (2)H Solid-State NMR

The interaction of the neuropeptide methionine-enkephalin (Menk) with bicelles was investigated by solid-state NMR. Bicelles composed of dimyristoylphosphatidylcholine (DMPC) and dicaproylphosphatidylcholine (DCPC) were modified to investigate the effect of the lipid headgroup and electrostatic charges on the association with Menk. A total of 10 mol % of DMPC was replaced by zwitterionic phosphatidylethanolamine (DMPE), anionic phosphatidylglycerol (DMPG), or phosphatidylserine (DMPS). The preparation of DMPE-doped bicelles (Bic/PE) is reported for the first time. The (31)P and (2)H NMR results revealed changes in the lipid dynamics when Menk interacts with the bicellar systems. (2)H NMR experiments showed a disordering effect of Menk on the lipid chains in all the bicelles except Bic/PG, whereas the study of the choline headgroups indicated a decreased order of the lipids only in Bic/PE and Bic/PG. Our results suggest that the insertion depth of Menk into bicelles is modulated by their composition, more specifically by the balance between hydrophobic and electrostatic interactions. Menk would be buried at the lipid polar/apolar interface, the depth of penetration into the hydrophobic membrane core following the scaling Bic > Bic/PE > Bic/PS at the slightly acidic pH used in this study. The peptide would not insert into the bilayer core of Bic/PG and would rather remain at the surface

Cation modulation of bicelle size and magnetic alignment as revealed by solid-state NMR and electron microscopy.

The influence of salts (KCl, NaCl, CaCl(2), and MgCl(2)) on bicelles (bilayered micelles) made of dimyristoylphosphatidylcholine (DMPC, molar fraction X = 78%) and dicaproylphosphatidylcholine (DCPC) was investigated by solid-state (31)P- and (2)H NMR as well as by freeze-fracture electron microscopy. Sizes were determined from (2)H- and (31)P NMR on the basis of a model that incorporated a planar bilayer and a (half-torus) curved rim representing the DMPC and DCPC regions of the bicelle, respectively. Good agreement was shown with sizes determined independently from freeze-fracture electron microscopy images. In the presence of K(+) and Na(+), bicelles have diameters of approximately 300 A while in the presence of Ca(2+) and Mg(2+); their diameter increases to approximately 500 A. Bicelle magnetic alignment is considerably improved by the presence of salts. The optimum salt concentration for such an effect ranges from 50 to 200 mM. Bicelles are magnetically aligned for temperatures roughly ranging from 30 degrees C to 40 degrees C with monovalent cations; this range is slightly extended in the presence of divalent salts. In this temperature range, the dynamics of the long-chain hydrocarbon region of the bicelle (leading to a bicelle thickness of 38 A) and of water is about the same independently of cation nature and concentration. However, at higher temperatures, considerable differences in water dynamics are observed between systems with monovalent and divalent cations. In these conditions, the system consists of a mixture of micelles and extended bilayers, which show residual macroscopic alignment in the magnetic field

Amphotericin and model membranes: The effect of amphotericin B on cholesterol-containing systems as viewed by 2 H-NMR

NRC publication: Ye