John F. Nagle

Structure of Fully Hydrated Fluid Phase Lipid Bilayers with Monounsaturated Chains

Quantitative structures are obtained at 30°C for the fully hydrated fluid phases of palmitoyloleoylphosphatidylcholine (POPC), with a double bond on the sn-2 hydrocarbon chain, and for dierucoylphosphatidylcholine (di22:1PC), with a double bond on each hydrocarbon chain. The form factors F(qz) for both lipids are obtained using a combination of three methods. (1) Volumetric measurements provide F(0). (2) X-ray scattering from extruded unilamellar vesicles provides ΙF(qz)Ι for low qz. (3) Diffuse X-ray scattering from oriented stacks of bilayers provides ΙF(qz)Ι for high qz. Also, data using method (2) are added to our recent data for dioleoylphosphatidylcholine (DOPC) using methods (1) and (3); the new DOPC data agree very well with the recent data and with (4) our older data obtained using a liquid crystallographic X-ray method. We used hybrid electron density models to obtain structural results from these form factors. The result for area per lipid (A) for DOPC 72.4 ± 0.5 Å2 agrees well with our earlier publications, and we find A = 69.3 ± 0.5 Å2 for di22:1PC and A = 68.3 ± 1.5 Å2 for POPC. We obtain the values for five different average thicknesses: hydrophobic, steric, head-head, phosphate-phosphate and Luzzati. Comparison of the results for these three lipids and for our recent dimyristoylphosphatidylcholine (DMPC) determination provides quantitative measures of the effect of unsaturation on bilayer structure. Our results suggest that lipids with one monounsaturated chain have quantitative bilayer structures closer to lipids with two monounsaturated chains than to lipids with two completely saturated chains.

X-ray structure determination of fully hydrated L alpha phase dipalmitoylphosphatidylcholine bilayers

Bilayer form factors obtained from x-ray scattering data taken with high instrumental resolution are reported for multilamellar vesicles of L alpha phase lipid bilayers of dipalmitoylphosphatidylcholine at 50 degrees C under varying osmotic pressure. Artifacts in the magnitudes of the form factors due to liquid crystalline fluctuations have been eliminated by using modified Caillé theory. The Caillé fluctuation parameter eta 1 increases systematically with increasing lamellar D spacing and this explains why some higher order peaks are unobservable for the larger D spacings. The corrected form factors fall on one smooth continuous transform F(q); this shows that the bilayer does not change shape as D decreases from 67.2 A (fully hydrated) to 60.9 A. The distance between headgroup peaks is obtained from Fourier reconstruction of samples with four orders of diffraction and from electron density models that use 38 independent form factors. By combining these results with previous gel phase results, area AF per lipid molecule and other structural quantities are obtained for the fluid L alpha phase. Comparison with results that we derived from previous neutron diffraction data is excellent, and we conclude from diffraction studies that AF = 62.9 +/- 1.3 A2, which is in excellent agreement with a previous estimate from NMR data.

Lipid Bilayer Structure Determined by the Simultaneous Analysis of Neutron and X-Ray Scattering Data

Quantitative structures were obtained for the fully hydrated fluid phases of dioleoylphosphatidylcholine (DOPC) and dipalmitoylphosphatidylcholine (DPPC) bilayers by simultaneously analyzing x-ray and neutron scattering data. The neutron data for DOPC included two solvent contrasts, 50% and 100% D(2)O. For DPPC, additional contrast data were obtained with deuterated analogs DPPC_d62, DPPC_d13, and DPPC_d9. For the analysis, we developed a model that is based on volume probability distributions and their spatial conservation. The models design was guided and tested by a DOPC molecular dynamics simulation. The model consistently captures the salient features found in both electron and neutron scattering density profiles. A key result of the analysis is the molecular surface area, A. For DPPC at 50 degrees C A = 63.0 A(2), whereas for DOPC at 30 degrees C A = 67.4 A(2), with estimated uncertainties of 1 A(2). Although A for DPPC agrees with a recently reported value obtained solely from the analysis of x-ray scattering data, A for DOPC is almost 10% smaller. This improved method for determining lipid areas helps to reconcile long-standing differences in the values of lipid areas obtained from stand-alone x-ray and neutron scattering experiments and poses new challenges for molecular dynamics simulations.

Hydrogen bonded chain mechanisms for proton conduction and proton pumping

This review focuses on the question of how protons are transported across membranes during bioenergetic processes. It has been found that proton transport through membranes is a central feature of several bioenergetic systems, such as bacteriorhodopsin, which pumps protons to higher free energies [115], and the ATP synthase, which consumes energetic protons [90]. In addition, it may be noted that proton transport has been implicated in transhydrogenase [28, 91], cytochrome oxidase, and the bcl redox loop in the mitochondrial respiratory chains [29, 90, 119]. ATP-driven proton pumps may be utilized in a variety of biological systems: evidence for them has been found in kidney [69] and liver [97] lysosomal vesicles, maize apical meristem plasmalemma [13], chromaffin granules [2], turtle bladder [1], Escherichia coli and Streptococcus lactis [68], and Neurospora crassa [36]. In addition, H + / K § antiport has been suggested to occur in alkaline tolerant strains of Bacillus firmus [38], in gastric parietal cells [102], and H § symport in E. coli [89]. While the question of proton transport mechanisms was stimulated by the delocalized chemiosmotic theory [72], it should be emphasized that the question is even more relevant for localized theories [120]. Indeed, for localized theories one requires proton transport for long distances along the membrane [49, 50, 86], not just proton transport across the membrane. It is also possible to imagine that even more extensive networks of proton pathways exist in the cell, perhaps utilizing the cytosol microstructure [5, 6]. Finally, proton transport may be a part of other bioenergetic mechanisms such as muscle action [75] or flagellar motion [34, 52]. It will not be our purpose to review the theories of Mitchell [72] and Williams [120]. These theories, which are very important, are phenomenological. They attempt to describe where various ions go and which enzymes and substrates are utilized, but they do not attempt to describe molecular mechanisms. One danger of purely phenomenological theories is that they may require processes for which no physical mechanisms exist. Our purpose will be to review theories of mechanisms for proton transport which can then be utilized within the framework of any phenomenological theories. Various kinds of active transmembrane proton transport can be envisaged (Fig. 1). One would involve a carrier molecule that would attach a proton and carry it across the membrane (Fig. 1 A). Another would involve a transmembrane protein

Structure and interactions of fully hydrated dioleoylphosphatidylcholine bilayers.

This study focuses on dioleoylphosphatidylcholine (DOPC) bilayers near full hydration. Volumetric data and high-resolution synchrotron x-ray data are used in a method that compares DOPC with well determined gel phase dipalmitoylphosphatidylcholine (DPPC). The key structural quantity obtained is fully hydrated area/lipid A0 = 72.2 +/- 1.1 A2 at 30 degrees C, from which other quantities such as thickness of the bilayer are obtained. Data for samples over osmotic pressures from 0 to 56 atmospheres give an estimate for the area compressibility of KA = 188 dyn/cm. Obtaining the continuous scattering transform and electron density profiles requires correction for liquid crystal fluctuations. Quantitation of these fluctuations opens an experimental window on the fluctuation pressure, the primary repulsive interaction near full hydration. The fluctuation pressure decays exponentially with water spacing, in agreement with analytical results for soft confinement. However, the ratio of decay length lambda(fl) = 5.8 A to hydration pressure decay length lambda = 2.2 A is significantly larger than the value of 2 predicted by analytical theory and close to the ratio obtained in recent simulations. We also obtain the traditional osmotic pressure versus water spacing data. Our analysis of these data shows that estimates of the Hamaker parameter H and the bending modulus Kc are strongly coupled.

Structure of gel phase DMPC determined by X-ray diffraction.

The structure of fully hydrated gel phase dimyristoylphosphatidylcholine lipid bilayers was obtained at 10°C. Oriented lipid multilayers were used to obtain high signal-to-noise intensity data. The chain tilt angle and an estimate of the methylene electron density were obtained from wide angle reflections. The chain tilt angle is measured to be 32.3 0.6 o near full hydration, and it does not change as the sample is mildly dehydrated from a repeat spacing of D 59.9 A to D 56.5 A. Low angle diffraction peaks were obtained up to the tenth order for 17 samples with variable D and prepared by three different methods with different geometries. In addition to the usual Fourier reconstructions of the electron density profiles, model electron density profiles were fit to all the low angle data simultaneously while constraining the model to include the wide-angle data and the measured lipid volume. Results are obtained for area/lipid (A 47.2 0.5 A 2 ), the compressibility modulus (KA 500 100 dyn/cm), various thicknesses, such as the hydrocarbon thickness (2DC 30.3 0.2 A), and the head-to-head spacing (DHH 40.1 0.1 A).

Area/lipid of bilayers from NMR.

Values of area per lipid A ranging from 56 to 72 A 2 have been reported from essentially the same SCD data from DPPC in the L alpha phase. The differences are due primarily to three separate binary choices in interpretation. It is argued that one particular combination is best; this yields A = 62 +/- 2 A 2 for DPPC at 50 degrees C. Each preceding interpretation agrees with at least one of the three present choices and disagrees with at least one.

Theory of Lipid Monolayer and Bilayer Phase Transitions" Effect of Headgroup Interactions

SummaryHeadgroup and soft core interactions are added to a lipid monolayer-bilayer model and the surface pressure-area phase diagrams are calculated. The results show that quite small headgroup interactions can have biologically significant effects on the transition temperature and the phase diagram. In particular, the difference in transition temperatures of lecithins and phosphatidyl ethanolamines is easy to reproduce in the model. The phosphatidic acid systems seem to require weak transient hydrogen bonding which is also conjectured to play a role in most of the lipid systems. By a simple surface free energy argument it is shown that monolayers under a surface pressure of 50 dynes/cm should behave as bilayers, in agreement with experiment. Although the headgroup interactions are biologically very significant, in fundamental studies of the main phase transition in lipids they are secondary in importance to the hydrocarbon chain interactions (including the excluded volume interaction, the rotational isomerism, and the attractive van der Waals interaction).

Theory of biomembrane phase transitions

Experiments on the chain melting thermal transition in simple biological membranes are briefly reviewed and shown to indicate that a microscopic order‐disorder model may be appropriate to describe the thermodynamics of the transition. To test this conclusion further a pair of two dimensional lattice models (A and B) are introduced and the statistical mechanics is solved exactly using dimer techniques. The phenomenological parameters required by the models are evaluated from experiments on other systems. The more realistic of the two models (model A) has a second order transition at a temperature of 353°K compared to 315°K for dipalmitoyl‐L‐α‐lecithin membranes. However, the specific heat peaks do not have the same shape and the transition is broader for model A than for the experiment. In comparison, the less realistic model B has a first order transition at 925°K, considerably higher than the experimental transition temperature. From these results it seems likely that the points of disagreement between m...

Structural Determinants of Water Permeability through the Lipid Membrane

Despite intense study over many years, the mechanisms by which water and small nonelectrolytes cross lipid bilayers remain unclear. While prior studies of permeability through membranes have focused on solute characteristics, such as size, polarity, and partition coefficient in hydrophobic solvent, we focus here on water permeability in seven single component bilayers composed of different lipids, five with phosphatidylcholine headgroups and different chain lengths and unsaturation, one with a phosphatidylserine headgroup, and one with a phosphatidylethanolamine headgroup. We find that water permeability correlates most strongly with the area/lipid and is poorly correlated with bilayer thickness and other previously determined structural and mechanical properties of these single component bilayers. These results suggest a new model for permeability that is developed in the accompanying theoretical paper in which the area occupied by the lipid is the major determinant and the hydrocarbon thickness is a secondary determinant. Cholesterol was also incorporated into DOPC bilayers and X-ray diffuse scattering was used to determine quantitative structure with the result that the area occupied by DOPC in the membrane decreases while bilayer thickness increases in a correlated way because lipid volume does not change. The water permeability decreases with added cholesterol and it correlates in a different way from pure lipids with area per lipid, bilayer thickness, and also with area compressibility.