This section deals with the phase behaviour of pure lipid bilayers. Several phases can be clearly distinguished by differences in the head and tail ordering. In the following, only a very simplified and generalised overview of the phases and the phase behaviour of pure lipid bilayers can be given. For detailed information, please refer to [Gen89, KC94a, KC94b, KC95, KC98].
Biomembranes consist of a complex mixture of different lipid types with very differ-ent characteristics. The phase behaviour of such mixtures is much more complex than the phase behaviour of a single lipid bilayer. In this work, only bilayers of a single lipid type are considered.
4.3.1. Temperature dependence
Figure 4.4 shows the temperature dependence of the molecular volume and the heat capacity of a DPPC bilayer. Both graphs show the signature of three distinct phase transitions in a temperature range between 10 and 450C.
At low temperatures, the bilayer is found to be in a so-called subgel phase, the lamel-lar crystalline phase Lc. In this phase, the aliphatic tail chains are mostly stretched, parallel to each other and very densely packed and well ordered. The long axis of the head group is oriented parallel to the bilayer plane, and the heads are closely packed in a lattice with presumably orthorombic cells.
When the temperature is increased, the so-called subtransition from the subgel phaseLcto the gel phasesLβ orLβ0 is encountered. The transition is mainly driven by the head group interaction: the polar head groups become hydrated and orient them-selves perpendicular to the bilayer plane, the distance between the heads increases
Figure 4.4: Temperature dependence of the molecular volume and the heat capacity of a DPPC bilayer (from [TNN04]).
4.3. Lipid bilayer phases
(a) Tilted gel phase Lβ0
(b) Untilted gel phaseLβ
(c) Interdigitated gel phaseLβI
(d) Fluid phaseLα
(e) Ripple phasePβ0
Figure 4.5: Sketch of the bilayer phases.
d*t dt
dh
θ θ
Figure 4.6: Sketch of the lipid tilt. By tilting the lipid tail chains with a cross sectiondtby the angleθ, the component in the bilayer planed∗t is increased to match the head cross sectiondh.
and the head group ordering changes to a hexagonal or quasi-hexagonal lattice. How-ever, the tail chains are still well ordered and densely packed (see figure 4.5).
The behaviour of the tail groups in the subgel phaseLc and the gel phasesLβ and Lβ0 again very much depends on the relation of the head surface area and the tail cross section, i.e. on the packing parameter S. When the cross section of the head group is larger than the cross section of the tails, the lipid tail chains become tilted with respect to the bilayer normal. This effectively increases the tail cross section parallel to the bilayer plane, so that the mismatch between head and tail size is reduced (see figure 4.6). Consequently, the tails can align and densly pack, and a stable bilayer can be maintained. This effect is mostly emphasised in the gel phases, leading to the distinction between the tilted gel phaseLβ0 and the untilted gel phaseLβ. While PCs generally exhibit the tilted gel phaseLβ0, PEs or sphingolipids possess an untilted gel phaseLβ.
Figure 4.7: Experimentalp-T phase diagram of a DMPC-bilayer (A) and a DPPC-bilayer (B) from [KC98]
Upon further increasing the temperature, many lipid systems that exhibit the tilted gel phaseLβ0 undergo the so-called pretransition to the ripple phases Pβ0 orPβ(mst). In these curious phases, the bilayer, that is flat in all other phases, exhibits a rippled struc-ture with a repeat distance of a few tens of lipids. The molecular strucstruc-ture of these phases is not well understood. Although there seems to be a certain degree of disor-der in the system, the bilayer is generally well ordisor-dered, comparable to the gel phase Lβ0. Only recently, molecular dynamics simulations of lecithin bilayers have unveiled the molecular structure of the phase [V YMM05], but the nature of the transition is still not well understood. Chapter 8 will shed further light onto the characteristics and the structure of the phase as well as the mechanisms driving the transition.
When heating up the system some more, the system undergoes a highly cooperative phase transition, the so-called main transition or chain order/disorder transition of lipid bilayers. Above the transition temperature, the liquid-crystalline or fluid phase Lα is found (see figure 4.5d on the previous page). Although the lipids are bound to the bilayer and can not escape into the watery environment, the heads and tails are quite disordered and fluid-like, and a high in-plane mobility can be observed (figure 4.5d).
The bilayer in this phase can be thought of as a two-dimensional fluid. With only a few exceptions, the lipid component of biomembranes in natural environment is found to be in this phase [Gen89].
4.3.2. Pressure dependence
Figure 4.7 shows experimental pressure–temperature phase diagrams of bilayers of DPPC and DMPC. From these diagrams it can be seen that the main and subtransition-temperatures increase with increasing pressure. However, what is more eye-catching is the fact that two new phases are observed at higher pressures.
In DPPC bilayers, at intermediate temperatures and high pressure, a new phase, the so called interdigitated phaseLβI, occurs. In this phase, the hydrophobic chains of the lipids interdigitate and create a bilayer with a very low thickness and no tilt.
The chains are very densely packed, however, there is a large head–head distance (see
4.3. Lipid bilayer phases
figure 4.5c on page 47). At normal pressures, the phase is absent in most PCs, as it has unfavourable voids between the heads and increases the tail–water interface.
However, the phase can be induced by adding alcohol or chaotropic salts to the lipid-water mixture[KS04]. As lipids, alcohols are amphiphilic molecules, that assemble at the hydrophilic-hydrophobic interface, i.e. they go between the head groups, effec-tively increasing the average head-head distance and resulting in the interdigitated phase.
At low temperature and high pressure, the system exhibits an additional subgel phase (Gel III) with monoclinic chain packing.
4.3.3. Lipid type dependence
Clearly, the lipid type has a profound influence on the phase behaviour of the lipid bilayer.
The length of the hydrophobic tails influences the transition temperature: the longer the hydrophobic tails, the higher the phase transition temperature. However, the tail length can also completely suppress some phases or transitions. For example, there is no difference between the gel and subgel phase (Lβ0 andLc) for PCs with chains that have less than 13 acyl groups. The interdigitated phase LβI strongly depends on the interaction of the head groups. For long tails, this interaction is less important, and therefore the interdigitated phase can be seen for DPPC bilayers at high pressures.
Furthermore, the ripple phase is strongly influenced by the chain length (see chapter 8).
When the lipid contains an unsaturated hydrocarbon tail, the normal zig-zag struc-ture of the chain is broken, as the bond angle of the groups surrounding the double C =C bond changes. Furthermore, the tail can not rotate around the bond. There-fore, a double bond can be thought of as a kink in the tail. Such a kink in the otherwise very regular aliphatic chain perturbs the packing of the chains in the ordered gel and subgel-phases and decreases the main transition temperature significantly. Branched tail chains have an effect very similar to unsaturated tail chains. However, branched tails also increase the average cross section of the tail and consequently change the packing parameterS.
The nature of the head group has a profound influence on the ordered gel and subgel phase structures and the subtransition temperature, as these strongly depend on the head group interaction. Additionally, the head group controls whether the gel phase is tilted or untilted and therefore whether a ripple phase exists or not. Also, the interdigitated gel phaseLβI can be observed in bilayers of lipids that have sufficiently large heads, e.g. dialkyl phosphatidylcholines.
Finally, the linkage between the tails and the glycerol backbone has an influence on the phase behaviour. When the tails are linked to the backbone by alkyl groups instead of acyl groups, the gel phaseLβis replaced by the interdigitated gel phaseLβI. This can be explained by the fact that while PCs with acyl linkage can build hydrogen bonds between the acyl oxygens that usually stabilises the gel phaseLβ, this can not happen in PCs with alkyl linkage.